Plasmids: Extrachromosomal, usually CIRCULAR DNA molecules that are self-replicating and transferable from one organism to another. They are found in a variety of bacterial, archaeal, fungal, algal, and plant species. They are used in GENETIC ENGINEERING as CLONING VECTORS.Conjugation, Genetic: A parasexual process in BACTERIA; ALGAE; FUNGI; and ciliate EUKARYOTA for achieving exchange of chromosome material during fusion of two cells. In bacteria, this is a uni-directional transfer of genetic material; in protozoa it is a bi-directional exchange. In algae and fungi, it is a form of sexual reproduction, with the union of male and female gametes.R Factors: A class of plasmids that transfer antibiotic resistance from one bacterium to another by conjugation.Bacteriocin Plasmids: Plasmids encoding bacterial exotoxins (BACTERIOCINS).DNA, Bacterial: Deoxyribonucleic acid that makes up the genetic material of bacteria.Escherichia coli: A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.Base Sequence: The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.Genes, Bacterial: The functional hereditary units of BACTERIA.Extrachromosomal Inheritance: Vertical transmission of hereditary characters by DNA from cytoplasmic organelles such as MITOCHONDRIA; CHLOROPLASTS; and PLASTIDS, or from PLASMIDS or viral episomal DNA.Molecular Sequence Data: Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.DNA Restriction Enzymes: Enzymes that are part of the restriction-modification systems. They catalyze the endonucleolytic cleavage of DNA sequences which lack the species-specific methylation pattern in the host cell's DNA. Cleavage yields random or specific double-stranded fragments with terminal 5'-phosphates. The function of restriction enzymes is to destroy any foreign DNA that invades the host cell. Most have been studied in bacterial systems, but a few have been found in eukaryotic organisms. They are also used as tools for the systematic dissection and mapping of chromosomes, in the determination of base sequences of DNAs, and have made it possible to splice and recombine genes from one organism into the genome of another. EC 3.21.1.Transformation, Bacterial: The heritable modification of the properties of a competent bacterium by naked DNA from another source. The uptake of naked DNA is a naturally occuring phenomenon in some bacteria. It is often used as a GENE TRANSFER TECHNIQUE.Cloning, Molecular: The insertion of recombinant DNA molecules from prokaryotic and/or eukaryotic sources into a replicating vehicle, such as a plasmid or virus vector, and the introduction of the resultant hybrid molecules into recipient cells without altering the viability of those cells.DNA Transposable Elements: Discrete segments of DNA which can excise and reintegrate to another site in the genome. Most are inactive, i.e., have not been found to exist outside the integrated state. DNA transposable elements include bacterial IS (insertion sequence) elements, Tn elements, the maize controlling elements Ac and Ds, Drosophila P, gypsy, and pogo elements, the human Tigger elements and the Tc and mariner elements which are found throughout the animal kingdom.Replicon: Any DNA sequence capable of independent replication or a molecule that possesses a REPLICATION ORIGIN and which is therefore potentially capable of being replicated in a suitable cell. (Singleton & Sainsbury, Dictionary of Microbiology and Molecular Biology, 2d ed)DNA, Recombinant: Biologically active DNA which has been formed by the in vitro joining of segments of DNA from different sources. It includes the recombination joint or edge of a heteroduplex region where two recombining DNA molecules are connected.F Factor: A plasmid whose presence in the cell, either extrachromosomal or integrated into the BACTERIAL CHROMOSOME, determines the "sex" of the bacterium, host chromosome mobilization, transfer via conjugation (CONJUGATION, GENETIC) of genetic material, and the formation of SEX PILI.Chromosomes, Bacterial: Structures within the nucleus of bacterial cells consisting of or containing DNA, which carry genetic information essential to the cell.Drug Resistance, Microbial: The ability of microorganisms, especially bacteria, to resist or to become tolerant to chemotherapeutic agents, antimicrobial agents, or antibiotics. This resistance may be acquired through gene mutation or foreign DNA in transmissible plasmids (R FACTORS).DNA Replication: The process by which a DNA molecule is duplicated.Transformation, Genetic: Change brought about to an organisms genetic composition by unidirectional transfer (TRANSFECTION; TRANSDUCTION, GENETIC; CONJUGATION, GENETIC, etc.) and incorporation of foreign DNA into prokaryotic or eukaryotic cells by recombination of part or all of that DNA into the cell's genome.Bacterial Proteins: Proteins found in any species of bacterium.Restriction Mapping: Use of restriction endonucleases to analyze and generate a physical map of genomes, genes, or other segments of DNA.Nucleic Acid Hybridization: Widely used technique which exploits the ability of complementary sequences in single-stranded DNAs or RNAs to pair with each other to form a double helix. Hybridization can take place between two complimentary DNA sequences, between a single-stranded DNA and a complementary RNA, or between two RNA sequences. The technique is used to detect and isolate specific sequences, measure homology, or define other characteristics of one or both strands. (Kendrew, Encyclopedia of Molecular Biology, 1994, p503)Genetic Vectors: DNA molecules capable of autonomous replication within a host cell and into which other DNA sequences can be inserted and thus amplified. Many are derived from PLASMIDS; BACTERIOPHAGES; or VIRUSES. They are used for transporting foreign genes into recipient cells. Genetic vectors possess a functional replicator site and contain GENETIC MARKERS to facilitate their selective recognition.Sequence Analysis, DNA: A multistage process that includes cloning, physical mapping, subcloning, determination of the DNA SEQUENCE, and information analysis.Plant Tumor-Inducing Plasmids: Plasmids coding for proteins which induce PLANT TUMORS. The most notable example of a plant tumor inducing plasmid is the Ti plasmid found associated with AGROBACTERIUM TUMEFACIENS.Recombination, Genetic: Production of new arrangements of DNA by various mechanisms such as assortment and segregation, CROSSING OVER; GENE CONVERSION; GENETIC TRANSFORMATION; GENETIC CONJUGATION; GENETIC TRANSDUCTION; or mixed infection of viruses.DNA, Circular: Any of the covalently closed DNA molecules found in bacteria, many viruses, mitochondria, plastids, and plasmids. Small, polydisperse circular DNA's have also been observed in a number of eukaryotic organisms and are suggested to have homology with chromosomal DNA and the capacity to be inserted into, and excised from, chromosomal DNA. It is a fragment of DNA formed by a process of looping out and deletion, containing a constant region of the mu heavy chain and the 3'-part of the mu switch region. Circular DNA is a normal product of rearrangement among gene segments encoding the variable regions of immunoglobulin light and heavy chains, as well as the T-cell receptor. (Riger et al., Glossary of Genetics, 5th ed & Segen, Dictionary of Modern Medicine, 1992)Gene Transfer, Horizontal: The naturally occurring transmission of genetic information between organisms, related or unrelated, circumventing parent-to-offspring transmission. Horizontal gene transfer may occur via a variety of naturally occurring processes such as GENETIC CONJUGATION; GENETIC TRANSDUCTION; and TRANSFECTION. It may result in a change of the recipient organism's genetic composition (TRANSFORMATION, GENETIC).Replication Origin: A unique DNA sequence of a replicon at which DNA REPLICATION is initiated and proceeds bidirectionally or unidirectionally. It contains the sites where the first separation of the complementary strands occurs, a primer RNA is synthesized, and the switch from primer RNA to DNA synthesis takes place. (Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed)Electrophoresis, Agar Gel: Electrophoresis in which agar or agarose gel is used as the diffusion medium.Mutation: Any detectable and heritable change in the genetic material that causes a change in the GENOTYPE and which is transmitted to daughter cells and to succeeding generations.Tetracycline: A naphthacene antibiotic that inhibits AMINO ACYL TRNA binding during protein synthesis.Genes: A category of nucleic acid sequences that function as units of heredity and which code for the basic instructions for the development, reproduction, and maintenance of organisms.beta-Lactamases: Enzymes found in many bacteria which catalyze the hydrolysis of the amide bond in the beta-lactam ring. Well known antibiotics destroyed by these enzymes are penicillins and cephalosporins.Transfection: The uptake of naked or purified DNA by CELLS, usually meaning the process as it occurs in eukaryotic cells. It is analogous to bacterial transformation (TRANSFORMATION, BACTERIAL) and both are routinely employed in GENE TRANSFER TECHNIQUES.Operon: In bacteria, a group of metabolically related genes, with a common promoter, whose transcription into a single polycistronic MESSENGER RNA is under the control of an OPERATOR REGION.Amino Acid Sequence: The order of amino acids as they occur in a polypeptide chain. This is referred to as the primary structure of proteins. It is of fundamental importance in determining PROTEIN CONFORMATION.Anti-Bacterial Agents: Substances that reduce the growth or reproduction of BACTERIA.Promoter Regions, Genetic: DNA sequences which are recognized (directly or indirectly) and bound by a DNA-dependent RNA polymerase during the initiation of transcription. Highly conserved sequences within the promoter include the Pribnow box in bacteria and the TATA BOX in eukaryotes.Transcription, Genetic: The biosynthesis of RNA carried out on a template of DNA. The biosynthesis of DNA from an RNA template is called REVERSE TRANSCRIPTION.DNA, Superhelical: Circular duplex DNA isolated from viruses, bacteria and mitochondria in supercoiled or supertwisted form. This superhelical DNA is endowed with free energy. During transcription, the magnitude of RNA initiation is proportional to the DNA superhelicity.Blotting, Southern: A method (first developed by E.M. Southern) for detection of DNA that has been electrophoretically separated and immobilized by blotting on nitrocellulose or other type of paper or nylon membrane followed by hybridization with labeled NUCLEIC ACID PROBES.Genetic Complementation Test: A test used to determine whether or not complementation (compensation in the form of dominance) will occur in a cell with a given mutant phenotype when another mutant genome, encoding the same mutant phenotype, is introduced into that cell.Rhizobium: A genus of gram-negative, aerobic, rod-shaped bacteria that activate PLANT ROOT NODULATION in leguminous plants. Members of this genus are nitrogen-fixing and common soil inhabitants.Escherichia coli Proteins: Proteins obtained from ESCHERICHIA COLI.Enterobacteriaceae: A family of gram-negative, facultatively anaerobic, rod-shaped bacteria that do not form endospores. Its organisms are distributed worldwide with some being saprophytes and others being plant and animal parasites. Many species are of considerable economic importance due to their pathogenic effects on agriculture and livestock.Sequence Homology, Nucleic Acid: The sequential correspondence of nucleotides in one nucleic acid molecule with those of another nucleic acid molecule. Sequence homology is an indication of the genetic relatedness of different organisms and gene function.Colicins: Bacteriocins elaborated by strains of Escherichia coli and related species. They are proteins or protein-lipopolysaccharide complexes lethal to other strains of the same species.Tetracycline Resistance: Nonsusceptibility of bacteria to the action of TETRACYCLINE which inhibits aminoacyl-tRNA binding to the 30S ribosomal subunit during protein synthesis.Plant Tumors: A localized proliferation of plant tissue forming a swelling or outgrowth, commonly with a characteristic shape and unlike any organ of the normal plant. Plant tumors or galls usually form in response to the action of a pathogen or a pest. (Holliday, P., A Dictionary of Plant Pathology, 1989, p330)Virulence: The degree of pathogenicity within a group or species of microorganisms or viruses as indicated by case fatality rates and/or the ability of the organism to invade the tissues of the host. The pathogenic capacity of an organism is determined by its VIRULENCE FACTORS.Polymerase Chain Reaction: In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships.Vaccines, DNA: Recombinant DNA vectors encoding antigens administered for the prevention or treatment of disease. The host cells take up the DNA, express the antigen, and present it to the immune system in a manner similar to that which would occur during natural infection. This induces humoral and cellular immune responses against the encoded antigens. The vector is called naked DNA because there is no need for complex formulations or delivery agents; the plasmid is injected in saline or other buffers.Chromosome Mapping: Any method used for determining the location of and relative distances between genes on a chromosome.Electroporation: A technique in which electric pulses of intensity in kilovolts per centimeter and of microsecond-to-millisecond duration cause a temporary loss of the semipermeability of CELL MEMBRANES, thus leading to ion leakage, escape of metabolites, and increased uptake by cells of drugs, molecular probes, and DNA.Salmonella: A genus of gram-negative, facultatively anaerobic, rod-shaped bacteria that utilizes citrate as a sole carbon source. It is pathogenic for humans, causing enteric fevers, gastroenteritis, and bacteremia. Food poisoning is the most common clinical manifestation. Organisms within this genus are separated on the basis of antigenic characteristics, sugar fermentation patterns, and bacteriophage susceptibility.Pseudomonas: A genus of gram-negative, aerobic, rod-shaped bacteria widely distributed in nature. Some species are pathogenic for humans, animals, and plants.Streptomycin: An antibiotic produced by the soil actinomycete Streptomyces griseus. It acts by inhibiting the initiation and elongation processes during protein synthesis.Klebsiella pneumoniae: Gram-negative, non-motile, capsulated, gas-producing rods found widely in nature and associated with urinary and respiratory infections in humans.Open Reading Frames: A sequence of successive nucleotide triplets that are read as CODONS specifying AMINO ACIDS and begin with an INITIATOR CODON and end with a stop codon (CODON, TERMINATOR).Kanamycin: Antibiotic complex produced by Streptomyces kanamyceticus from Japanese soil. Comprises 3 components: kanamycin A, the major component, and kanamycins B and C, the minor components.DNA: A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine).Repetitive Sequences, Nucleic Acid: Sequences of DNA or RNA that occur in multiple copies. There are several types: INTERSPERSED REPETITIVE SEQUENCES are copies of transposable elements (DNA TRANSPOSABLE ELEMENTS or RETROELEMENTS) dispersed throughout the genome. TERMINAL REPEAT SEQUENCES flank both ends of another sequence, for example, the long terminal repeats (LTRs) on RETROVIRUSES. Variations may be direct repeats, those occurring in the same direction, or inverted repeats, those opposite to each other in direction. TANDEM REPEAT SEQUENCES are copies which lie adjacent to each other, direct or inverted (INVERTED REPEAT SEQUENCES).Cell Line: Established cell cultures that have the potential to propagate indefinitely.Species Specificity: The restriction of a characteristic behavior, anatomical structure or physical system, such as immune response; metabolic response, or gene or gene variant to the members of one species. It refers to that property which differentiates one species from another but it is also used for phylogenetic levels higher or lower than the species.Penicillinase: A beta-lactamase preferentially cleaving penicillins. (Dorland, 28th ed) EC 3.5.2.-.Gene Expression Regulation, Bacterial: Any of the processes by which cytoplasmic or intercellular factors influence the differential control of gene action in bacteria.DNA, Viral: Deoxyribonucleic acid that makes up the genetic material of viruses.Drug Resistance, Bacterial: The ability of bacteria to resist or to become tolerant to chemotherapeutic agents, antimicrobial agents, or antibiotics. This resistance may be acquired through gene mutation or foreign DNA in transmissible plasmids (R FACTORS).Genome, Bacterial: The genetic complement of a BACTERIA as represented in its DNA.Drug Resistance, Multiple, Bacterial: The ability of bacteria to resist or to become tolerant to several structurally and functionally distinct drugs simultaneously. This resistance may be acquired through gene mutation or foreign DNA in transmissible plasmids (R FACTORS).Chloramphenicol: An antibiotic first isolated from cultures of Streptomyces venequelae in 1947 but now produced synthetically. It has a relatively simple structure and was the first broad-spectrum antibiotic to be discovered. It acts by interfering with bacterial protein synthesis and is mainly bacteriostatic. (From Martindale, The Extra Pharmacopoeia, 29th ed, p106)Lactose Factors: Plasmids which determine the ability of a bacterium to ferment lactose.Electrophoresis, Gel, Pulsed-Field: Gel electrophoresis in which the direction of the electric field is changed periodically. This technique is similar to other electrophoretic methods normally used to separate double-stranded DNA molecules ranging in size up to tens of thousands of base-pairs. However, by alternating the electric field direction one is able to separate DNA molecules up to several million base-pairs in length.Salmonella typhimurium: A serotype of Salmonella enterica that is a frequent agent of Salmonella gastroenteritis in humans. It also causes PARATYPHOID FEVER.Nucleic Acid Conformation: The spatial arrangement of the atoms of a nucleic acid or polynucleotide that results in its characteristic 3-dimensional shape.Genes, Regulator: Genes which regulate or circumscribe the activity of other genes; specifically, genes which code for PROTEINS or RNAs which have GENE EXPRESSION REGULATION functions.Transduction, Genetic: The transfer of bacterial DNA by phages from an infected bacterium to another bacterium. This also refers to the transfer of genes into eukaryotic cells by viruses. This naturally occurring process is routinely employed as a GENE TRANSFER TECHNIQUE.Microbial Sensitivity Tests: Any tests that demonstrate the relative efficacy of different chemotherapeutic agents against specific microorganisms (i.e., bacteria, fungi, viruses).Chloramphenicol Resistance: Nonsusceptibility of bacteria to the action of CHLORAMPHENICOL, a potent inhibitor of protein synthesis in the 50S ribosomal subunit where amino acids are added to nascent bacterial polypeptides.DNA-Binding Proteins: Proteins which bind to DNA. The family includes proteins which bind to both double- and single-stranded DNA and also includes specific DNA binding proteins in serum which can be used as markers for malignant diseases.Phenotype: The outward appearance of the individual. It is the product of interactions between genes, and between the GENOTYPE and the environment.DNA, Fungal: Deoxyribonucleic acid that makes up the genetic material of fungi.Gene Order: The sequential location of genes on a chromosome.Chloramphenicol O-Acetyltransferase: An enzyme that catalyzes the acetylation of chloramphenicol to yield chloramphenicol 3-acetate. Since chloramphenicol 3-acetate does not bind to bacterial ribosomes and is not an inhibitor of peptidyltransferase, the enzyme is responsible for the naturally occurring chloramphenicol resistance in bacteria. The enzyme, for which variants are known, is found in both gram-negative and gram-positive bacteria. EC 2.3.1.28.Phylogeny: The relationships of groups of organisms as reflected by their genetic makeup.Enterococcus faecalis: A species of gram-positive, coccoid bacteria commonly isolated from clinical specimens and the human intestinal tract. Most strains are nonhemolytic.Nebramycin: A complex of antibiotic substances produced by Streptomyces tenebrarius.Molecular Weight: The sum of the weight of all the atoms in a molecule.Kanamycin Resistance: Nonsusceptibility of bacteria to the antibiotic KANAMYCIN, which can bind to their 70S ribosomes and cause misreading of messenger RNA.Bacteriophages: Viruses whose hosts are bacterial cells.Escherichia coli Infections: Infections with bacteria of the species ESCHERICHIA COLI.Ampicillin: Semi-synthetic derivative of penicillin that functions as an orally active broad-spectrum antibiotic.Deoxyribonuclease EcoRI: One of the Type II site-specific deoxyribonucleases (EC 3.1.21.4). It recognizes and cleaves the sequence G/AATTC at the slash. EcoRI is from E coliRY13. Several isoschizomers have been identified. EC 3.1.21.-.Streptomyces: A genus of bacteria that form a nonfragmented aerial mycelium. Many species have been identified with some being pathogenic. This genus is responsible for producing a majority of the ANTI-BACTERIAL AGENTS of practical value.Deoxyribonucleases, Type II Site-Specific: Enzyme systems containing a single subunit and requiring only magnesium for endonucleolytic activity. The corresponding modification methylases are separate enzymes. The systems recognize specific short DNA sequences and cleave either within, or at a short specific distance from, the recognition sequence to give specific double-stranded fragments with terminal 5'-phosphates. Enzymes from different microorganisms with the same specificity are called isoschizomers. EC 3.1.21.4.Genetic Engineering: Directed modification of the gene complement of a living organism by such techniques as altering the DNA, substituting genetic material by means of a virus, transplanting whole nuclei, transplanting cell hybrids, etc.Trimethoprim: A pyrimidine inhibitor of dihydrofolate reductase, it is an antibacterial related to PYRIMETHAMINE. It is potentiated by SULFONAMIDES and the TRIMETHOPRIM, SULFAMETHOXAZOLE DRUG COMBINATION is the form most often used. It is sometimes used alone as an antimalarial. TRIMETHOPRIM RESISTANCE has been reported.Bacteriocins: Substances elaborated by specific strains of bacteria that are lethal against other strains of the same or related species. They are protein or lipopolysaccharide-protein complexes used in taxonomy studies of bacteria.Genes, Viral: The functional hereditary units of VIRUSES.Mercury: A silver metallic element that exists as a liquid at room temperature. It has the atomic symbol Hg (from hydrargyrum, liquid silver), atomic number 80, and atomic weight 200.59. Mercury is used in many industrial applications and its salts have been employed therapeutically as purgatives, antisyphilitics, disinfectants, and astringents. It can be absorbed through the skin and mucous membranes which leads to MERCURY POISONING. Because of its toxicity, the clinical use of mercury and mercurials is diminishing.Trimethoprim Resistance: Nonsusceptibility of bacteria to the action of TRIMETHOPRIM.Integrons: DNA elements that include the component genes and insertion site for a site-specific recombination system that enables them to capture mobile gene cassettes.Saccharomyces cerevisiae: A species of the genus SACCHAROMYCES, family Saccharomycetaceae, order Saccharomycetales, known as "baker's" or "brewer's" yeast. The dried form is used as a dietary supplement.Genes, Reporter: Genes whose expression is easily detectable and therefore used to study promoter activity at many positions in a target genome. In recombinant DNA technology, these genes may be attached to a promoter region of interest.Gene Expression Regulation: Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control (induction or repression) of gene action at the level of transcription or translation.Spiroplasma citri: The type species of gram-negative bacteria in the genus SPIROPLASMA, family SPIROPLASMATACEAE, causing citrus stubborn disease.Agrobacterium tumefaciens: A species of gram-negative, aerobic bacteria isolated from soil and the stems, leafs, and roots of plants. Some biotypes are pathogenic and cause the formation of PLANT TUMORS in a wide variety of higher plants. The species is a major research tool in biotechnology.DNA Helicases: Proteins that catalyze the unwinding of duplex DNA during replication by binding cooperatively to single-stranded regions of DNA or to short regions of duplex DNA that are undergoing transient opening. In addition DNA helicases are DNA-dependent ATPases that harness the free energy of ATP hydrolysis to translocate DNA strands.DNA Primers: Short sequences (generally about 10 base pairs) of DNA that are complementary to sequences of messenger RNA and allow reverse transcriptases to start copying the adjacent sequences of mRNA. Primers are used extensively in genetic and molecular biology techniques.Gene Transfer Techniques: The introduction of functional (usually cloned) GENES into cells. A variety of techniques and naturally occurring processes are used for the gene transfer such as cell hybridization, LIPOSOMES or microcell-mediated gene transfer, ELECTROPORATION, chromosome-mediated gene transfer, TRANSFECTION, and GENETIC TRANSDUCTION. Gene transfer may result in genetically transformed cells and individual organisms.Recombinant Fusion Proteins: Recombinant proteins produced by the GENETIC TRANSLATION of fused genes formed by the combination of NUCLEIC ACID REGULATORY SEQUENCES of one or more genes with the protein coding sequences of one or more genes.RNA, Messenger: RNA sequences that serve as templates for protein synthesis. Bacterial mRNAs are generally primary transcripts in that they do not require post-transcriptional processing. Eukaryotic mRNA is synthesized in the nucleus and must be exported to the cytoplasm for translation. Most eukaryotic mRNAs have a sequence of polyadenylic acid at the 3' end, referred to as the poly(A) tail. The function of this tail is not known for certain, but it may play a role in the export of mature mRNA from the nucleus as well as in helping stabilize some mRNA molecules by retarding their degradation in the cytoplasm.Rhodococcus equi: A species of RHODOCOCCUS found in soil, herbivore dung, and in the intestinal tract of cows, horses, sheep, and pigs. It causes bronchopneumonia in foals and can be responsible for infection in humans compromised by immunosuppressive drug therapy, lymphoma, or AIDS.Viral Proteins: Proteins found in any species of virus.Tellurium: Tellurium. An element that is a member of the chalcogen family. It has the atomic symbol Te, atomic number 52, and atomic weight 127.60. It has been used as a coloring agent and in the manufacture of electrical equipment. Exposure may cause nausea, vomiting, and CNS depression.Mutagenesis, Insertional: Mutagenesis where the mutation is caused by the introduction of foreign DNA sequences into a gene or extragenic sequence. This may occur spontaneously in vivo or be experimentally induced in vivo or in vitro. Proviral DNA insertions into or adjacent to a cellular proto-oncogene can interrupt GENETIC TRANSLATION of the coding sequences or interfere with recognition of regulatory elements and cause unregulated expression of the proto-oncogene resulting in tumor formation.beta-Galactosidase: A group of enzymes that catalyzes the hydrolysis of terminal, non-reducing beta-D-galactose residues in beta-galactosides. Deficiency of beta-Galactosidase A1 may cause GANGLIOSIDOSIS, GM1.Gene Expression: The phenotypic manifestation of a gene or genes by the processes of GENETIC TRANSCRIPTION and GENETIC TRANSLATION.Bacillus subtilis: A species of gram-positive bacteria that is a common soil and water saprophyte.Coliphages: Viruses whose host is Escherichia coli.Penicillin Resistance: Nonsusceptibility of an organism to the action of penicillins.Lactococcus lactis: A non-pathogenic species of LACTOCOCCUS found in DAIRY PRODUCTS and responsible for the souring of MILK and the production of LACTIC ACID.Pseudomonas aeruginosa: A species of gram-negative, aerobic, rod-shaped bacteria commonly isolated from clinical specimens (wound, burn, and urinary tract infections). It is also found widely distributed in soil and water. P. aeruginosa is a major agent of nosocomial infection.Mutagenesis: Process of generating a genetic MUTATION. It may occur spontaneously or be induced by MUTAGENS.Shigella sonnei: A lactose-fermenting bacterium causing dysentery.Bacteriophage lambda: A temperate inducible phage and type species of the genus lambda-like viruses, in the family SIPHOVIRIDAE. Its natural host is E. coli K12. Its VIRION contains linear double-stranded DNA with single-stranded 12-base 5' sticky ends. The DNA circularizes on infection.Gentamicins: A complex of closely related aminoglycosides obtained from MICROMONOSPORA purpurea and related species. They are broad-spectrum antibiotics, but may cause ear and kidney damage. They act to inhibit PROTEIN BIOSYNTHESIS.Interspersed Repetitive Sequences: Copies of transposable elements interspersed throughout the genome, some of which are still active and often referred to as "jumping genes". There are two classes of interspersed repetitive elements. Class I elements (or RETROELEMENTS - such as retrotransposons, retroviruses, LONG INTERSPERSED NUCLEOTIDE ELEMENTS and SHORT INTERSPERSED NUCLEOTIDE ELEMENTS) transpose via reverse transcription of an RNA intermediate. Class II elements (or DNA TRANSPOSABLE ELEMENTS - such as transposons, Tn elements, insertion sequence elements and mobile gene cassettes of bacterial integrons) transpose directly from one site in the DNA to another.Chromosome Deletion: Actual loss of portion of a chromosome.Deoxyribonuclease HindIII: One of the Type II site-specific deoxyribonucleases (EC 3.1.21.4). It recognizes and cleaves the sequence A/AGCTT at the slash. HindIII is from Haemophilus influenzae R(d). Numerous isoschizomers have been identified. EC 3.1.21.-.Staphylococcus aureus: Potentially pathogenic bacteria found in nasal membranes, skin, hair follicles, and perineum of warm-blooded animals. They may cause a wide range of infections and intoxications.Bacteria: One of the three domains of life (the others being Eukarya and ARCHAEA), also called Eubacteria. They are unicellular prokaryotic microorganisms which generally possess rigid cell walls, multiply by cell division, and exhibit three principal forms: round or coccal, rodlike or bacillary, and spiral or spirochetal. Bacteria can be classified by their response to OXYGEN: aerobic, anaerobic, or facultatively anaerobic; by the mode by which they obtain their energy: chemotrophy (via chemical reaction) or PHOTOTROPHY (via light reaction); for chemotrophs by their source of chemical energy: CHEMOLITHOTROPHY (from inorganic compounds) or chemoorganotrophy (from organic compounds); and by their source for CARBON; NITROGEN; etc.; HETEROTROPHY (from organic sources) or AUTOTROPHY (from CARBON DIOXIDE). They can also be classified by whether or not they stain (based on the structure of their CELL WALLS) with CRYSTAL VIOLET dye: gram-negative or gram-positive.Bacterial Toxins: Toxic substances formed in or elaborated by bacteria; they are usually proteins with high molecular weight and antigenicity; some are used as antibiotics and some to skin test for the presence of or susceptibility to certain diseases.Sequence Homology, Amino Acid: The degree of similarity between sequences of amino acids. This information is useful for the analyzing genetic relatedness of proteins and species.Thiamphenicol: A methylsulfonyl analog of CHLORAMPHENICOL. It is an antibiotic and immunosuppressive agent.Enterobacteriaceae Infections: Infections with bacteria of the family ENTEROBACTERIACEAE.Klebsiella Infections: Infections with bacteria of the genus KLEBSIELLA.Hemolysin Factors: Plasmids controlling the synthesis of hemolysin by bacteria.RNA Phages: Bacteriophages whose genetic material is RNA, which is single-stranded in all except the Pseudomonas phage phi 6 (BACTERIOPHAGE PHI 6). All RNA phages infect their host bacteria via the host's surface pili. Some frequently encountered RNA phages are: BF23, F2, R17, fr, PhiCb5, PhiCb12r, PhiCb8r, PhiCb23r, 7s, PP7, Q beta phage, MS2 phage, and BACTERIOPHAGE PHI 6.Clostridium perfringens: The most common etiologic agent of GAS GANGRENE. It is differentiable into several distinct types based on the distribution of twelve different toxins.Virus Replication: The process of intracellular viral multiplication, consisting of the synthesis of PROTEINS; NUCLEIC ACIDS; and sometimes LIPIDS, and their assembly into a new infectious particle.Ampicillin Resistance: Nonsusceptibility of a microbe to the action of ampicillin, a penicillin derivative that interferes with cell wall synthesis.Virulence Factors: Those components of an organism that determine its capacity to cause disease but are not required for its viability per se. Two classes have been characterized: TOXINS, BIOLOGICAL and surface adhesion molecules that effect the ability of the microorganism to invade and colonize a host. (From Davis et al., Microbiology, 4th ed. p486)Lac Operon: The genetic unit consisting of three structural genes, an operator and a regulatory gene. The regulatory gene controls the synthesis of the three structural genes: BETA-GALACTOSIDASE and beta-galactoside permease (involved with the metabolism of lactose), and beta-thiogalactoside acetyltransferase.Klebsiella oxytoca: A species of gram-negative bacteria causing URINARY TRACT INFECTIONS and SEPTICEMIA.beta-Lactam Resistance: Nonsusceptibility of bacteria to the action of the beta-lactam antibiotics. Mechanisms responsible for beta-lactam resistance may be degradation of antibiotics by BETA-LACTAMASES, failure of antibiotics to penetrate, or low-affinity binding of antibiotics to targets.Genes, Fungal: The functional hereditary units of FUNGI.Serotyping: Process of determining and distinguishing species of bacteria or viruses based on antigens they share.

Stabilization of poly-L-lysine/DNA polyplexes for in vivo gene delivery to the liver. (1/43669)

We are developing a self-assembling non-viral in vivo gene delivery vehicle based on poly-l-lysine and plasmid DNA. We have characterized poly-l-lysines of different chain lengths for DNA condensation and strength of DNA binding. Poly-l-lysine chains >20 residues bound DNA efficiently in physiological saline, while shorter chains did not. Attachment of asialoorosomucoid to PLL increased the PLL chain length required for efficient DNA binding in saline and for efficient DNA condensation. By electron microscopy, poly-l-lysine/DNA polyplexes appeared as toroids 25-50 nm in diameter or rods 40-80 nm long; conjugation of asialoorosomucoid to the polylysine component increased the size of resulting polyplexes to 50-90 nm. In water, poly-l-lysine and asialoorosomucoid-PLL polyplexes have effective diameters of 46 and 87.6 nm, respectively. Polyplexes containing only poly-l-lysine and DNA aggregated in physiological saline at all charge ratios and aggregated at neutral charge ratios in water. Attachment of asialoorosomucoid lessened, but did not eliminate, the aggregation of PLL polyplexes, and did not result in efficient delivery of polyplexes to hepatocytes. Conjugation of polyethylene glycol to poly-l-lysine sterically stabilized resulting polyplexes at neutral charge ratios by shielding the surfaces. For efficient in vivo gene delivery, polyplexes will need to be sterically stabilized to prevent aggregation and interaction with serum components.  (+info)

Cloning and characterisation of a novel ompB operon from Vibrio cholerae 569B. (2/43669)

The ompB operon of Vibrio cholerae 569B has been cloned and fully sequenced. The operon encodes two proteins, OmpR and EnvZ, which share sequence identity with the OmpR and EnvZ proteins of a variety of other bacteria. Although the order of the ompR and envZ genes of V. cholerae is similar to that of the ompB operon of E. coli, S. typhimurium and X. nematophilus, the Vibrio operon exhibits a number of novel features. The structural organisation and features of the V. cholerae ompB operon are described.  (+info)

Co-expression of glutathione S-transferase with methionine aminopeptidase: a system of producing enriched N-terminal processed proteins in Escherichia coli. (3/43669)

We describe here an Escherichia coli expression system that produces recombinant proteins enriched in the N-terminal processed form, by using glutathione S-transferase cGSTM1-1 and rGSTT1-1 as models, where c and r refer to chick and rat respectively. Approximately 90% of the cGSTM1-1 or rGSTT1-1 overexpressed in E. coli under the control of a phoA promoter retained the initiator methionine residue that was absent from the mature isoenzymes isolated from tissues. The amount of initiator methionine was decreased to 40% of the expressed cGSTM1-1 when the isoenzyme was co-expressed with an exogenous methionine aminopeptidase gene under the control of a separate phoA promoter. The recombinant proteins expressed were mainly methionine aminopeptidase. The yield of cGSTM1-1 was decreased to 10% of that expressed in the absence of the exogenous methionine aminopeptidase gene. By replacing the phoA with its natural promoter, the expression of methionine aminopeptidase decreased drastically. The yield of the co-expressed cGSTM1-1 was approx. 60% of that in the absence of the exogenous methionine aminopeptidase gene; approx. 65% of the initiator methionine residues were removed from the enzyme. Under similar conditions, N-terminal processing was observed in approx. 70% of the recombinant rGSTT1-1 expressed. By increasing the concentration of phosphate in the growth medium, the amount of initiator methionine on cGSTM1-1 was decreased to 14% of the overexpressed isoenzymes, whereas no further improvement could be observed for rGSTT1-1. The initiator methionine residue does not affect the enzymic activities of either cGSTM1-1 or rGSTT1-1. However, the epoxidase activity and the 4-nitrobenzyl chloride-conjugating activity of the purified recombinant rGSTT1-1 are markedly higher that those reported recently for the same isoenzyme isolated from rat livers.  (+info)

Expression of the plague plasminogen activator in Yersinia pseudotuberculosis and Escherichia coli. (4/43669)

Enteropathogenic yersiniae (Yersinia pseudotuberculosis and Yersinia enterocolitica) typically cause chronic disease as opposed to the closely related Yersinia pestis, the causative agent of bubonic plague. It is established that this difference reflects, in part, carriage by Y. pestis of a unique 9.6-kb pesticin or Pst plasmid (pPCP) encoding plasminogen activator (Pla) rather than distinctions between shared approximately 70-kb low-calcium-response, or Lcr, plasmids (pCD in Y. pestis and pYV in enteropathogenic yersiniae) encoding cytotoxic Yops and anti-inflammatory V antigen. Pla is known to exist as a combination of 32.6-kDa (alpha-Pla) and slightly smaller (beta-Pla) outer membrane proteins, of which at least one promotes bacterial dissemination in vivo and degradation of Yops in vitro. We show here that only alpha-Pla accumulates in Escherichia coli LE392/pPCP1 cultivated in enriched medium and that either autolysis or extraction of this isolate with 1.0 M NaCl results in release of soluble alpha and beta forms possessing biological activity. This process also converted cell-bound alpha-Pla to beta-Pla and smaller forms in Y. pestis KIM/pPCP1 and Y. pseudotuberculosis PB1/+/pPCP1 but did not promote solubilization. Pla-mediated posttranslational hydrolysis of pulse-labeled Yops in Y. pseudotuberculosis PB1/+/pPCP1 occurred more slowly than that in Y. pestis but was otherwise similar except for accumulation of stable degradation products of YadA, a pYV-mediated fibrillar adhesin not encoded in frame by pCD. Carriage of pPCP by Y. pseudotuberculosis did not significantly influence virulence in mice.  (+info)

The virulence plasmid-encoded impCAB operon enhances survival and induced mutagenesis in Shigella flexneri after exposure to UV radiation. (5/43669)

Upon exposure to UV radiation, Shigella flexneri SA100 displayed survival and mutation frequencies comparable to those of Escherichia coli AB1157, which contains a functional UmuDC error-prone DNA repair system. Survival of SA100 after UV irradiation was associated with the presence of the 220-kb virulence plasmid, pVP. This plasmid encodes homologues of ImpA and ImpB, which comprise an error-prone DNA repair system encoded on plasmid TP110 that was initially identified in Salmonella typhimurium, and ImpC, encoded upstream of ImpA and ImpB. Although the impB gene was present in representatives of all four species of Shigella, not all isolates tested contained the gene. Shigella isolates that lacked impB were more sensitive to UV radiation than isolates that contained impB. The nucleotide sequence of a 2.4-kb DNA fragment containing the imp operon from S. flexneri SA100 pVP was 96% identical to the imp operon from the plasmid TP110. An SA100 derivative with a mutation in the impB gene had reduced survival following UV irradiation and less UV-induced mutagenesis relative to the parental strain. We also found that S. flexneri contained a chromosomally encoded umuDC operon; however, the umuDC promoter was not induced by exposure to UV radiation. This suggests that the imp operon but not the umuDC operon contributes to survival and induced mutagenesis in S. flexneri following exposure to UV radiation.  (+info)

Molecular and evolutionary analysis of Borrelia burgdorferi 297 circular plasmid-encoded lipoproteins with OspE- and OspF-like leader peptides. (6/43669)

We previously described two OspE and three OspF homologs in Borrelia burgdorferi 297 (D. R. Akins, S. F. Porcella, T. G. Popova, D. Shevchenko, S. I. Baker, M. Li, M. V. Norgard, and J. D. Radolf, Mol. Microbiol. 18:507-520, 1995; D. R. Akins, K. W. Bourell, M. J. Caimano, M. V. Norgard, and J. D. Radolf, J. Clin. Investig. 101:2240-2250, 1998). In this study, we characterized four additional lipoproteins with OspE/F-like leader peptides (Elps) and demonstrated that all are encoded on plasmids homologous to cp32 and cp18 from the B31 and N40 strains, respectively. Statistical analysis of sequence similarities using the binary comparison algorithm revealed that the nine lipoproteins from strain 297, as well as the OspE, OspF, and Erp proteins from the N40 and B31 strains, fall into three distinct families. Based upon the observation that these lipoproteins all contain highly conserved leader peptides, we now propose that the ancestors of each of the three families arose from gene fusion events which joined a common N terminus to unrelated proteins. Additionally, further sequence analysis of the strain 297 circular plasmids revealed that rearrangements appear to have played an important role in generating sequence diversity among the members of these three families and that recombinational events in the downstream flanking regions appear to have occurred independently of those within the lipoprotein-encoding genes. The association of hypervariable regions with genes which are differentially expressed and/or subject to immunological pressures suggests that the Lyme disease spirochete has exploited recombinatorial processes to foster its parasitic strategy and enhance its immunoevasiveness.  (+info)

Reverse transcription-nested polymerase chain reaction for detecting p40 RNA of Borna disease virus, without risk of plasmid contamination. (7/43669)

Several methods for the detection of Borna disease virus (BDV) RNA have been reported, one being the reverse transcription-nested polymerase chain reaction (RT-nested PCR) method. However, due to the possibility of contamination of the cloned DNA in a reaction tube, false-positive results might be obtained by RT-nested PCR. To detect only BDV RNA without anxiety of contamination, we developed an RT-nested PCR system using "mRNA selective PCR kit". Using this system, cDNA of BDV p40 in the plasmid (up to 5 x 10(7) molecules) was not amplified. BDV specific sequence was amplified from total RNA (more than 50 pg) of MDCK/BDV cells, which were persistently infected with BDV. These results indicate that this mRNA selective RT-nested PCR system can specifically amplify target RNA as distinguished from plasmid contaminated.  (+info)

Diversity of rhizobia associated with Amorpha fruticosa isolated from Chinese soils and description of Mesorhizobium amorphae sp. nov. (8/43669)

Fifty-five Chinese isolates from nodules of Amorpha fruticosa were characterized and compared with the type strains of the species and genera of bacteria which form nitrogen-fixing symbioses with leguminous host plants. A polyphasic approach, which included RFLP of PCR-amplified 16S rRNA genes, multilocus enzyme electrophoresis (MLEE), DNA-DNA hybridization, 16S rRNA gene sequencing, electrophoretic plasmid profiles, cross-nodulation and a phenotypic study, was used in the comparative analysis. The isolates originated from several different sites in China and they varied in their phenotypic and genetic characteristics. The majority of the isolates had moderate to slow growth rates, produced acid on YMA and harboured a 930 kb symbiotic plasmid (pSym). Five different RFLP patterns were identified among the 16S rRNA genes of all the isolates. Isolates grouped by PCR-RFLP of the 16S rRNA genes were also separated into groups by variation in MLEE profiles and by DNA-DNA hybridization. A representative isolate from each of these DNA homology groups had a separate position in a phylogenetic tree as determined from sequencing analysis of the 16S rRNA genes. A new species, Mesorhizobium amorphae, is proposed for the majority of the isolates, which belonged to a moderately slow- to slow-growing, acid-producing group based upon their distinct phylogenetic position, their unique electrophoretic type, their low DNA homology with reference strains representing the species within the genus Mesorhizobium and their distinct phenotypic features. Strain ACCC 19665 was chosen as the type strain for M. amorphae sp. nov.  (+info)

Antibiotic resistance represents a significant public health problem. When resistance genes are mobile, being carried on plasmids or phages, their spread can be greatly accelerated. Plasmids in particular have been implicated in the spread of antibiotic resistance genes. However, the selective pressures which favour plasmid-carried resistance genes have not been fully established. Here we address this issue with mathematical models of plasmid dynamics in response to different antibiotic treatment regimes. We show that transmission of plasmids is a key factor influencing plasmid-borne antibiotic resistance, but the dosage and interval between treatments is also important. Our results also hold when plasmids carrying the resistance gene are in competition with other plasmids that do not carry the resistance gene. By altering the interval between antibiotic treatments, and the dosage of antibiotic, we show that different treatment regimes can select for either plasmid-carried, or chromosome-carried,
Individual bacterial cells may contain several different types of plasmids and in some cases more than 10 at a time. Plasmids are generally isolated from the bacterial cells in the supercoiled configuration. So far, thousands of different types of plasmids have been isolated. More than 300 different types of naturally occurring plasmids have been isolated from E.coli alone. Though, plasmids are not considered as part of the cells genome, when a bacterial cell divides each daughter cells receives a copy of each plasmid. Plasmids can also be transferred from one bacterial cell to another by the process called conjugation. Plasmids that govern their own transfer by conjugation are called conjugative plasmids but not all plasmids are conjugative.. ...
Götting C, Thierbach G, Pühler A, Kalinowski J. Versatile low-copy-number plasmids for temperature-inducible overexpression of bacterial genes in Escherichia coli. BIOTECHNIQUES. 1998;24(3):362 ...
The prevalence of 20-30 kb plasmids, almost half of which belong to only three restriction types by RFLP analysis, in staphylococci isolated from sources very distant in time and space suggests that these nonconjugative plasmids are surprisingly widespread for non-self-mobile plasmids. Plasmids in this size range can potentially be transferred by transducing phages (Lindsay and Holden 2006; Malachowa and Deleo 2010; Smillie et al. 2010); most phage genomes identified in staphylococci are ,40 kb, and transduction is thought to be restricted by phage genome size (Smillie et al. 2010). More of these 20-30 kb plasmids may be mobilizable than is apparent with current genome data if they contain mob genes not yet identified as such. However, the scarcity of conjugative plasmids (only 12 in total) implies that mobilization is rare and that staphylococcal plasmid transfer occurs mainly by transduction (Lindsay and Holden 2004; Lindsay 2010). The now larger dataset makes the mechanism of intercellular ...
Although IncP-1 plasmids are important for horizontal gene transfer among bacteria, in particular antibiotic resistance spread, so far only three plasmids from the subgroup IncP-1 alpha have been completely sequenced. In this study we doubled this number. The three IncP-1 alpha plasmids pB5, pB11 and pSP21 were isolated from bacteria of two different sewage treatment plants and sequenced by a combination of next-generation and capillary sequencing technologies. A comparative analysis including the previously analysed IncP-1 alpha plasmids RK2, pTB11 and pBS228 revealed a highly conserved plasmid backbone (at least 99.9% DNA sequence identity) comprising 54 core genes. The accessory elements of the plasmid pB5 constitute a class 1 integron interrupting the parC gene and an IS6100 copy inserted into the integron. In addition, the tetracycline resistance genes tetAR and the ISTB11-like element are located between the klc operon and the trfA-ssb operon. Plasmid pB11 is loaded with a Tn5053-like ...
OBJECTIVES: To investigate the diversity of plasmids that carry blaTEM-52 genes among Escherichia coli and Salmonella enterica originating from animals, meat products and humans. METHODS: A collection of 22 blaTEM-52-encoding plasmids was characterized by restriction fragment length polymorphism (RFLP), replicon typing (by PCR or replicon sequencing), susceptibility testing, assessment of plasmid ability to self-transfer by conjugation and typing of the genetic environment of the blaTEM-52 gene. Detected IncI1 plasmids underwent further plasmid multilocus sequence typing. RESULTS: RFLP profiles demonstrated dissemination of blaTEM-52 in Denmark (imported meat from Germany), France, Belgium and the Netherlands from 2000 to 2006 by mainly two different plasmids, one encoding blaTEM-52b (IncX1A, 45 kb) and the other blaTEM-52c (IncI1, 80 kb). In addition, blaTEM-52b was also found to be located on various other plasmids belonging to IncA/C and IncL/M, while blaTEM-52c was found on IncN-like as well ...
In this semirural community, we found that numerically dominant commensal E. coli strains (showing similar antimicrobial resistance and same antibiotic resistance genes) colonizing children and domestic animals in the same period of time and in the same community are genotypically diverse. We also found that plasmids carrying the same antibiotic resistance genes were distinct, which is consistent with recent reports showing that AMR genes move frequently among different plasmids (28, 29). Our research suggests that a common pool of AMR genes could be cocirculating on different plasmids among different E. coli clones in a community (Table 2)-probably through dissemination mediated by transposons, integrons, or gene cassettes (28, 30). Even when the same resistance gene alleles and same plasmid replicon types were identified across isolates, the plasmids harboring these traits were still distinct. We also found potential evidence of Tn2 participation in mobility of the gene blaTEM-1B, as we found ...
Plasmids, DNA (or rarely RNA) molecules which replicate in cells autonomously (independently of chromosomes) as non-essential genetic elements, play important roles for microbes grown under specific environmental conditions as well as in scientific laboratories and in biotechnology. For example, bacterial plasmids are excellent models in studies on regulation of DNA replication, and their derivatives are the most commonly used vectors in genetic engineering. Detailed mechanisms of replication initiation, which is the crucial process for efficient maintenance of plasmids in cells, have been elucidated for several plasmids. However, to understand plasmid biology, it is necessary to understand regulation of plasmid DNA replication in response to different environmental conditions in which host cells exist. Knowledge of such regulatory processes is also very important for those who use plasmids as expression vectors to produce large amounts of recombinant proteins. Variable conditions in large-scale
Plasmids are important members of the bacterial mobile gene pool, and are among the most important contributors to horizontal gene transfer between bacteria. They typically harbour a wide spectrum of host beneficial traits, such as antibiotic resistance, inserted into their backbones. Although these inserted elements have drawn considerable interest, evolutionary information about the plasmid backbones, which encode plasmid related traits, is sparse. Here we analyse 25 complete backbone genomes from the broad-host-range IncP-1 plasmid family. Phylogenetic analysis reveals seven clades, in which two plasmids that we isolated from a marine biofilm represent a novel clade. We also found that homologous recombination is a prominent feature of the plasmid backbone evolution. Analysis of genomic signatures indicates that the plasmids have adapted to different host bacterial species. Globally circulating IncP-1 plasmids hence contain mosaic structures of segments derived from several parental plasmids that
A plasmid partition system is a mechanism that assures the stable transmission of plasmids during bacterial cell division. Each plasmid has its independent replication system which controls the number of copies of the plasmid in a cell. The higher the copy number is, the more likely the two daughter cells will contain the plasmid. Generally, each molecule of plasmid diffuses randomly, so the probability of having a plasmid-less daughter cell is 21−N, where N is the number of copies. For instance, if there are 2 copies of a plasmid in a cell, there is a 50% chance of having one plasmid-less daughter cell. However, high-copy number plasmids have a cost for the hosting cell. This metabolic burden is lower for low-copy plasmids, but those have a higher probability of plasmid loss after a few generations. To control vertical transmission of plasmids, in addition to controlled-replication systems, bacterial plasmids use different maintenance strategies, such as multimer resolution systems, ...
Plasmid-mediated resistance is the transfer of antibiotic resistance genes which are carried on plasmids. The plasmids can be transferred between bacteria within the same species or between different species via conjugation. Plasmids often carry multiple antibiotic resistance genes, contributing to the spread of multidrug-resistance (MDR). Antibiotic resistance mediated by MDR plasmids severely limits the treatment options for the infections caused by Gram-negative bacteria, especially Enterobacteriaceae family. The global spread of MDR plasmids has been enhanced by selective pressure from antibiotic usage in human and veterinary medicine. Resistance plasmids by definition carry one or more antibiotic resistance genes. They are frequently accompanied by the genes encoding virulence determinants, specific enzymes or resistance to toxic heavy metals. Multiple resistance genes are commonly arranged in the resistance cassettes. The antibiotic resistance genes found on the plasmids confer resistance ...
Fragments ofCandida boidinii chromosomal DNA were inserted into the integrative vector YIp-kanr and examined for the presence of sequences promoting autonomous replication of plasmids inSaccharomyces cerevisiae. Restriction maps of two plasmids, designated S6/4 and S6/5, originating from the sameS. cerevisiae transformant, were constructed. Southern hybridization data confirmed that the plasmids carry sequences from theC. boidinii chromosome. Both plasmids transformS. cerevisiae strains at 4-5-fold higher frequency than cloning vectors based on the replication origin of the 2μm plasmid. Mitotic stability of the constructed plasmids is similar to that of the 2μ-based vector pNF2 inS. cerevisiae.
To study thermal adaptations in the cyanobacterium, Synechococcus sp. PCC 7002, we screened about 3,000 mutants for their tolerance to high temperature, and found one, SHT1, that is sensitive to high-temperature stress. The mutant had a modified gene construct in the endogenous plasmid, pAQ1. One of the four ORFs, ORF93, was duplicated, and its mRNA level was higher than in the wild type. At 38°C, the growth of SHT1 was retarded as compared with the wild type, and above 38°C, almost all the cells of SHT1 died. This temperature is much lower than that required for induction of heat shock proteins. Interestingly, in both the wild type and SHT1, the thermal stability of oxygen-evolving machinery increased upon acclimation to high temperatures. These findings indicate that the lack of thermal tolerance in the SHT1 strain is likely independent of the adaptation of the PSII complex and heat shock responses, whereas there are essential contributions of genes in the endogenous plasmid to the ...
Recreate original plasmid by cutting out insert - posted in Molecular Cloning: Maybe this is a stupid question, but I am going to ask it anyway: I would like to cut out an insert from my plasmid (pET28c) between restriction sites Nhe1 and Not1 and ligate the plasmid back to its orginial state without the insert (no I dont have the original plasmid anymore). However, obviously these restriction sites dont generate compatible sticky ends. Does anyone know how to recreate this...
Despite the near-ubiquity of plasmids in bacterial populations and the profound contribution of plasmid-borne genes and infectious gene transfer to the adaptation and evolution of bacteria, the mechanisms responsible for the maintenance of plasmids in bacterial populations are poorly understood. In this report, we address the question of how plasmids manage to persist over evolutionary time. Previous explanations have typically relied upon the ability of plasmids to deliver occasionally-useful genes to the right place at the right time. In contrast, we present a general mathematical proof that if (as suggested by several empirical studies) plasmids are not infectiously transmitted at a rate high enough to be maintained as genetic parasites in single populations, they will not be able to persist indefinately in these populations by carrying genes that are beneficial or sometimes beneficial to their host bacteria. Using more specific mathematical models, along with computer simulations, we ...
uncut plasmid dna vs linearized plasmid gel - posted in Molecular Cloning: Hello, I am going to run a gel comparing my uncut plasmid dna vs linearized plasmid. My insert is 135 bps and my vector is 3Kb. What can I expect to see on my gel, and how many bands can I expect to see. I am assuming the uncut plasmid will have several bands at different sizes and the linearized will have only one, is that right ? I am assuming the total size of my product will now be 3135 bps. Pls advise.
Figure 20. The pBR322 plasmid. Two genes of pBR322 confer resistance to antibiotics to any cell that contains the plasmid. AmpR confers resistance to amplicillin and tetR confers resistance to tetracycline to cells containing the plasmids. AmpR is a gene that codes for the periplasmic enzyme beta-lactamase that cleaves the ring structure found in amphicillin, which is a penicillin antibiotic. TetR is a gene that codes for a protein that modifies the bacterial cell wall and prevents tetracycline from entering the cell.. Multiple restriction endonuclease sites are present where foreign DNA fragments may be inserted.. Relaxed plasmid DNA replication continues in the presence of chloramphenicol. An interesting feature of this plasmid is that relaxed plasmid DNA replication continues even in the presence of an inhibitor of protein synthesis such as chloramphenicol. This feature allows increased yields of plasmid/cell of up to 100-fold ...
Easy Yeast Plasmid Isolation Kit: rescue plasmids from yeast using spin columns. Simple and highly efficient method. Zymolyase included.
One of the disadvantages of circular plasmids and chromosomes is their high sensitivity to rearrangements caused by homologous recombination. Odd numbers of crossing-over occurring during or after replication of a circular replicon result in the formation of a dimeric molecule in which the two copies of the replicon are fused. If they are not converted back to monomers, the dimers of replicons may fail to correctly segregate at the time of cell division. Resolution of multimeric forms of circular plasmids and chromosomes is mediated by site-specific recombination, and the enzymes that catalyze this type of reaction fall into two families of proteins: the serine and tyrosine recombinase families. Here we give an overview of the variety of site-specific resolution systems found on circular plasmids and chromosomes.
Santa Cruz Biotechnology now offers target-specific CRISPR/Cas9 Knockout (KO) Plasmids, CRISPR Double Nickase Plasmids and CRISPR/dCas9 Activation Plasmids for over 18,910 human and 18,340 mouse protein encoding genes. CRISPR/Cas9 KO Plasmids and CRISPR Double Nickase Plasmids enable the identification and cleavage of specific genes encoding a protein of interest thereby eliminating production of that gene product (protein). CRISPR/dCas9 Activation Plasmids activate endogenous gene transcription using a robust synergistic activation mediator (SAM) system. This exciting new technology is a useful tool for studying protein function and signalling pathways ...
In spite of the importance of plasmids in bacterial adaptation we have a poor understanding of their dynamics. It is not known if or how plasmids persist in and spread through (invade) a bacterial population when there is no selection for plasmid-encoded traits. Moreover, the differences in dynamics between spatially structured and mixed populations are poorly understood. Through a joint experimental/theoretical approach we tested the hypothesis that self-transmissible IncP-1 plasmids can invade a bacterial population in the absence of selection when initially very rare, but only in spatially structured habitats and when nutrients are regularly replenished. Using protocols that differed in degree of spatial structure and nutrient levels, the invasiveness of plasmid pB10 in E. coli was monitored during at least 15 days, with an initial fraction of plasmid-bearing (p+) cells as low as 1E-7. To further explore the mechanisms underlying plasmid dynamics we developed a spatially explicit mathematical ...
Mono- and Stereopictres of 5.0 Angstrom coordination sphere of Manganese atom in PDB 3dkx: Crystal Structure of the Replication Initiator Protein Encoded on Plasmid PMV158 (Repb), Trigonal Form, to 2.7 Ang Resolution
Difficult to Express Proteins: Novel Plasmid Technology to Significantly Increase Product Yield in CHO Cells, by Marco Cacciutolo
Mcr-1-harboring Enterobacteriaceae are reported worldwide since their first discovery in 2015. However, a limited number of studies are available that compared full-length plasmid sequences of human and animal origins. In this study, mcr-1-bearing plasmids from seven Escherichia coli isolates recovered from patients (n = 3), poultry meat (n = 2) and turkey meat (n = 2) in Switzerland were further analyzed and compared. Isolates were characterized by multilocus sequence typing (MLST). The mcr-1-bearing plasmids were transferred by transformation into reference strain E. coli DH5α and MCR-1-producing transformants were selected on LB-agar supplemented with 2 mg/L colistin. Purified plasmids were then sequenced and compared. MLST revealed six distinct STs, illustrating the high clonal diversity among mcr-1-positive E. coli isolates of different origins. Two different mcr-1-positive plasmids were identified from a single E. coli ST48 human isolate. All other isolates possessed a
The reason that we stock plasmids that only contain one terminator is because sometimes, despite the increased versatility, researchers may not want unnecessary terminators in their plasmids, for example when size constraints are an issue. The button below takes you to the plasmids with single terminators. If you prefer to use a plasmid with triple terminator, we suggest you search using the Plasmid Seach tool since nearly all our plasmids have triple terminators. The Plasmid Search tool will give you access to our full plasmid catalogue.. Finally, please try the Design Your Own Plasmid Online button below, to see what our cloning system can do for you. And remember that, while our plasmids are designed for easy cloning modifications, we are happy to do it for you if you prefer to outsource the cloning work.. ...
Prokaryotic transcriptomes change not only in response to physiological parameters but also to genetic rearrangements mediated by mobile elements. Plasmids are extrachromosomal genetic elements that replicate autonomously, and many can be transmitted between different strains through conjugation. Plasmids provide benefits to their hosts, such as resistance to antibiotics or degradation of recalcitrant aromatic compounds [1]; however, in several cases, the carriage of a large plasmid results in changes in the transcriptome of the host chromosome [2-4]. Similar to the effects of plasmid carriage on the transcriptional network of the host chromosome, differences in host background can alter the transcription patterns of backbone and accessory genes on a plasmid. Many plasmid backbone genes essential for conjugative transfer, replication initiation, and active partitioning are regulated both autogenously and by host factors [5]. Additionally, a number of plasmid-encoded degradative accessory genes ...
Synthetic biology heavily depends on rapid and simple techniques for DNA engineering, such as Ligase Cycling Reaction (LCR), Gibson assembly and Golden Gate assembly, all of which allow for fast, multi-fragment DNA assembly. A major enhancement of Golden Gate assembly is represented by the Modular Cloning (MoClo) system that allows for simple library propagation and combinatorial construction of genetic circuits from reusable parts. Yet, one limitation of the MoClo system is that all circuits are assembled in low- and medium copy plasmids, while a rapid route to chromosomal integration is lacking. To overcome this bottleneck, here we took advantage of the conditional-replication, integration, and modular (CRIM) plasmids, which can be integrated in single copies into the chromosome of Escherichia coli and related bacteria by site-specific recombination at different phage attachment (att) sites. By combining the modularity of the MoClo system with the CRIM plasmids features we created a set of 32 novel
ID YEP367 preliminary; circular DNA; SYN; 8400 BP. XX AC ATCC37735; XX DT 01-JUL-1993 (Rel. 7, Created) DT 01-JUL-1995 (Rel. 12, Last updated, Version 1) XX DE Saccharomyces/E.coli plasmid vector YEp367 - incomplete. XX KW cloning vector. XX OS Cloning vector OC Artificial sequences; Cloning vehicles. XX RN [1] RC YEp352E from YEp352 & linker RC YEp363A from pNM480 & YEp351 RC YEp353A from pNM480 & YEp352 RC YEp353 from YEp353A & YEp352E RC YEp354A from pNM481 & YEp352 RC YEp354 from YEp354A & YEp352E RC YEp355A from pNM482 & YEp352 RC YEp355 from YEp355A & YEp352E RC YEp356, YEp356R from YEp353 & pUC18 RC YEp357, YEp357R from YEp354 & pUC18 RC YEp358, YEp358R from YEp355 & pUC18 RC YEp363 from YEp363A & YEp353 RC YEp364 from YEp363A & YEp354 RC YEp365 from YEp363A & YEp355 RC YEp366 from YEp363A & YEp356 RC YEp367 from YEp363A & YEp357 RC YEp368 from YEp363A & YEp358 RC YEp366R from YEp363A & YEp356R RC YEp367R from YEp363A & YEp357R RC YEp368R from YEp363A & YEp358R RC YIp353 from YEp353 & ...
Background Prokaryotic plasmids have played out significant roles in the evolution of bacterial genomes and have a great impact on the metabolic functions of the host cell. variable genes (distributed genes and unique genes) than to the chromosomal core genes. Although all the functional categories of the chromosomal genes were exhibited by the plasmid genes, the proportions of each category differed between these two gene sets. The 598 gene families shared between chromosomes and plasmids displayed a uniform distribution between the two groups. A phylogenetic analysis of the shared genes, including the chromosomal core gene set, indicated that gene exchange events between plasmids and chromosomes occurred frequently during the evolutionary histories of the strains and species in this group. Moreover, the shared genes between plasmids and chromosomes usually had different promoter and terminator sequences, suggesting that they are regulated by different elements at the transcriptional level. ...
www.MOLUNA.de Plasmids in Bacteria [4191995] - Structure and Evolution.- Plasmids as Organisms.- Report on a Workshop: Structure and Function.- Evolutionary Relevance of Genetic Rearrangements Involving Plasmids.- Mechanisms of Transposition in Bacteria.- Insertion of Transcriptional Elements Outside the Replication Region Can Interfere with Replication, Maintenance, and Stability of ColE1-Derived Plasmids.- Studies on the Transposition of IS1.- On
ID YEP353 preliminary; circular DNA; SYN; 7944 BP. XX AC U03500; ATCC37725; XX DT 01-JUL-1993 (Rel. 7, Created) DT 01-JUL-1995 (Rel. 12, Last updated, Version 1) XX DE Saccharomyces/E.coli plasmid vector YEp353 - complete. XX KW cloning vector. XX OS Cloning vector OC Artificial sequences; Cloning vehicles. XX RN [1] RP 1-7944 RC YEp352E from YEp352 & linker RC YEp363A from pNM480 & YEp351 RC YEp353A from pNM480 & YEp352 RC YEp353 from YEp353A & YEp352E RC YEp354A from pNM481 & YEp352 RC YEp354 from YEp354A & YEp352E RC YEp355A from pNM482 & YEp352 RC YEp355 from YEp355A & YEp352E RC YEp356, YEp356R from YEp353 & pUC18 RC YEp357, YEp357R from YEp354 & pUC18 RC YEp358, YEp358R from YEp355 & pUC18 RC YEp363 from YEp363A & YEp353 RC YEp364 from YEp363A & YEp354 RC YEp365 from YEp363A & YEp355 RC YEp366 from YEp363A & YEp356 RC YEp367 from YEp363A & YEp357 RC YEp368 from YEp363A & YEp358 RC YEp366R from YEp363A & YEp356R RC YEp367R from YEp363A & YEp357R RC YEp368R from YEp363A & YEp358R RC YIp353 ...
DNA plasmids. Coloured atomic force micrograph of numerous pGL3 plasmids of DNA (deoxyribonucleic acid). A plasmid is a loop of DNA that can exist in a cell independently of the cells chromosomal DNA. Plasmids are important as they can replicate and express genes despite being outside the organisms chromosomes. This means that they can be used to introduce genes to organisms, a process used in gene therapy and genetic engineering. Genes, specific lengths of DNA, determine the development and characteristics of an organism. Atomic force microscopes make an image by moving a sensitive probe over a surface. Magnification: x24,000 at 6x7cm size. - Stock Image G110/0752
The present results concern the recombinant bacteria Escherichia coliHB101(GAPDH) which produces glyceraldehyde 3-phosphate dehydrogenase. An unusual phenomenon was noticed concerning the plasmid...
Plasmids are found across bacteria, archaea, and eukaryotes and play an important role in evolution. Plasmids exist at different copy numbers, the number of copies of the plasmid per cell, ranging from a single plasmid per cell to hundreds of plasmids per cell. This feature of a copy number greater than one can lead to a population of plasmids within a single cell that are not identical clones of one another, but rather have individual mutations that make a given plasmid unique. During cell division, this population of plasmids is partitioned into the two daughter cells, resulting in a random distribution of different plasmid variants in each daughter. In this study, we use stochastic simulations to investigate how random plasmid partitioning compares to a perfect partitioning model. Our simulation results demonstrate that random plasmid partitioning accelerates mutant allele fixation when the allele is beneficial and the selection is in an additive or recessive regime where increasing the copy ...
Duncan Clark wrote: , , I was always curious about the replicons used in some clinical E.colis , that carry umpteen different plasmids. I remember many many years ago , that one such E.coli was used for plasmid markers and had may 10 , different plasmids in it. They didnt appear to lose those plasmids , despite no selection. May have been published in Plasmid back in the , late 70s or early 80s. It all depends on the replication control mechanism of the replicons I suppose. ColE1 replicon is the standard one taught to students and I havent really look into others, so Im speaking in complete ignorance here, but there is no reason why a plasmid with different replicon would necessary result in plasmid incompatibility (because ColE1 has an unusual replication control mechanism?), although I understand that many would. , Duncan ...
View Notes - BMI_Lab8DNAPurification from PHS 2301 at St. Johns Duplicate. Laboratory VIII Topic: Purification of DNA, plasmid isolation using alkaline method Introduction: Genetic transformation
SimPlot analysis.Similarity plots with plasmids R751, pBP136 and pB3 as reference plasmids. Each coloured plot corresponds to a specific plasmid depicted in the
The isolation method is optimized for cultures grown in LB media; other rich media may require increased volumes of Suspension-, Lysis-, and Neutralization Buffer, and an additional wash step. The isolation procedure is suitable for all plasmid sizes; lysates of larger constructs (up to 100 kb) should be cleared by filtration to avoid shearing.. The yield of plasmid DNA preparations is dependent on several parameters, e.g., quality of the bacterial culture growth, amount of used culture suspension for the preparation, plasmid type used etc. As a rule of thumb the typical yield of a high copy number plasmid is about 3 - 5 µg of DNA per ml of original bacterial culture (pUC, pTZ, pGEM in common host strains like XL-1 blue, HB101, JM 109). The typical yield of low copy number plasmids is about 0.2 - 1 µg of DNA per ml of original bacterial culture.. The Genopure kits are supplied with folded filters to eliminate the time-consuming centrifugation step after the alkaline lysis. In approximately 2 ...
Plasmids, strains and primers used in this study are listed in Additional file 7: Table S1, Additional file 8: Table S2, Additional file 9: S3 1. Oligonucleotides and gBlocks were ordered from IDT and Eurofins. All fragments obtained by PCR were gel- or column purified (Nucleospin® Gel and PCR Clean-up columns) before cloning, and resulting plasmids were verified by sequencing (Eurofins). Yeast transformations were done using lithium acetate and PEG3350, and genomic integrations were performed with various helper plasmids and pre-expressed iCas9 from plasmid pCT (Addgene #60620) and plated on Sc-Leu+cloNAT. Strains were cured for pCT and helper plasmids after genome engineering and before proceeding to transcriptional regulation using dCas9.. EasyClone-MarkerFree vectors pCfB2909, pCfB3035, pCfB3037 and helper plasmids pCfB3042, pCfB3046, and pCfB3050 as well as genomic integration verification primers were adapted as previously described [46]. Yeast strains were plated according to ...
Genetic reagents for generating plasmids containing multiples of a DNA segment are prepared by modification of the restriction sites of plasmid rings. The modified plasmids permit cloning of DNA segments which can be recovered with sticky ends that are complementary but not rotationally equivalent. Such segments will polymerize in a head-to-tail conformation. The plasmid rings with modified restriction sites are also used in linear form to obtain head-to-tail joining of multiple DNA segments into plasmid rings for further cloning and/or expression of the DNA segment-directed protein. The critical restriction site sequences of the modified plasmid rings can be prepared as a reagent which permits the sequence to be introduced into any plasmid. The reagents have utility in preparing multiples of protein-forming genes, and in preparing large amounts of homogeneous DNAs which can themselves be used as reagents.
Fig 3. Scheme of construction of 4 isogenic biosensor strains. It was a pyramiding process during which the phenotypes of bacterial reporter developed previously in our project were combined, in order to implement strains with different mercury sensitivity. (E) BBa_J23103, BBa_J23101 and BBa_J23117 belong to a constitutive promoter library from partsregistry, among which BBa_J23103 is a very weak one, BBa_J23117 is medium and BBa_J23101 is the second strongest in the library. (A)Expression of MerR driven by weak constitutive promoter BBa_J23103 on low copy number plasmid backbone pSB3K3 was pyramided with wild type PmerT which is the most mercury-sensitive promoter combined with LacZ alpha fragment as the reporter gene on plasmid backbone pSB1A3. It is an extreme to confer the bioreporter the most sensitivity, based on our previous results. (B) Strong promoter BBa_J23101 drive the high expression intensity of MerR on plasmid backbone pSB1A2, while PmerT 88 on pSB3K3 is the most insensitive ...
Replication of the miniF plasmid pML31 was examined during the division cycle of Escherichia coli growing with doubling times between 40 and 90 min at 37°C and compared to the replication of plasmid pBR322 and the minichromosome pAL70. The replication pattern of pML31 was indistinguishable from that of pBR322 at all growth rates and very different from the cell- cycle-specific replication of the minichromosome. It is concluded that both pML31 and pBR322 plasmids can replicate at all stages of the division cycle, with a probability of replication that increases gradually, but perhaps nut exponentially, during the cycle. In contrast, the modes of segregation of pML31 and pBR322 plasmids into daughter cells at division appeared to differ, raising the possibility that pML31 may segregate in a nonrandom fashion similar to that of chromosomes and minichromosomes ...
Our method for disrupting E. coli chromosomal genes is analogous to one that has been used for many years in yeast (8). It is based on results of K. Murphy (18) who provided us with his materials before publication. Because multicopy plasmids might interfere with recombination by acting as competitive inhibitors (18), we cloned the Red genes (γ, β, and exo) into a low copy number plasmid. We used a vector which shows temperature-sensitive replication (37) to permit its easy curing from the resultant mutants. The plasmids pKD20 and pKD46 (Fig. 2) express the Red system under control of a well-regulated promoter to avoid unwanted recombinational events under noninducing conditions. They differ in that the latter has the native tL3 terminator downstream of exo. Although all recombinants described here were made by using pKD20, we now use pKD46 instead because we recently discovered that pKD46 yields a greatly enhanced number of recombinants. The reason is unknown. Curiously, tL3 encodes a small ...
Like the above examples, pSC101s replication is positively regulated by RepA binding the origin. RepA is also used to control copy number, by two mechanisms. Firstly, RepA negatively regulates its own transcription, thus the RepA protein levels (and its ability to promote replication) is confined to narrow limits. Secondly, The plasmid contains several (3-7) repeats of a 17-22bp sequence called iteron sequences. RepA binds the iterons, and at higher plasmid conncentration, this can lead to "handcuffing" of two plasmids. Interestingly, adding extra iteron sequences on other plasmids can reduce the copy number by this handcuffing mechanism. F, RK6, P1, RK2, and RP4 also use iterons, but the regulating protein and origins differ. pETcoco is an interesting plasmid, made by Novagen. It can be maintained as a single copy plasmid using the origin and positive regulator from the F plasmid (oriS and RepE). It can be swiched to a medium copy plasmid using the machinery from the RK2 plasmid (oviV and ...
Plasmid replication control is usually controlled by balancing the levels of a positive and a negative regulator of replication. For some plasmids (pMB1/colE1 replicons) the positive regulator is an RNA and in others (e.g. pSC101) it is a protein. Plasmids with a protein positive regulator will not replicate in the abscence of protein production - stringent control (although not the same as the stringent response due to a shortage of loaded tRNAs). Plasmids with an RNA positive regulator will continue to replicate in the abscence of protein production. This is termed relaxed control. High yields of plasmid may be obtained by halting protein production (via chloroamphenicol) when the culture reaches a high density and then continuing incubation for a number of hours. This might be of practical relevance when prepping the 1 and 3 series of Synthetic Biology plasmids.--BC 19:05, 3 Sep 2005 (EDT) ...
Plasmid Construction. The pGL3-promoter and pGL3-basic plasmids were purchased from Promega. The pL6 plasmid was generated by ligation of the 1.3-kb KpnI and XhoI (-1326 to +1; the translation start site was designated as +1) DNA fragment of mouse MOR into the polylinker sites of a promoterless luciferase vector, pGL3-basic (Promega, Madison, WI). The sequence of the insertion was confirmed by sequencing. The DNA fragment flanking the poly (A) site was generated by PCR from mouse genomic DNA with a pair of primers (sense, 5′-AATAGGCCGGCCGCATTAGGAGCATTGCTGAG-3′; antisense, 5′-ACGCGTCGACCCTAACTCTGGGATGGCAAG-3′; the underlined nucleotides indicate the overhanging restriction enzyme sites for FseI and SalI, respectively). The pL6PA and pGL3pPA plasmids were constructed by subcloning this DNA fragment after digestion with FseI and SalI into pL6 and pGL3-promoter plasmid, respectively. The pL6 and pGL3-promoter plasmids also have been digested by FseI and SalI, to replace the original SV40 ...
This two part exercise provides state-of-the-art information and practical experience with a variety of techniques that form the foundation of the biotechnology industry. In part A, students create a strain of E. coli that is resistant to the antibiotic ampicillin by introducing a plasmid that contains an ampicillin-resistance gene. The success of the transformation is monitored by growing the bacteria on an ampicillin-containing media. This experiment provides sufficient sterile materials for sixteen platings. In Part B, students isolate the amplified plasmid from the bacteria without the use of toxic chemicals. Then they digest the isolated DNA with EcoR1 and compare the electrophoretic properties of linear and circular plasmid DNA molecules. By inserting desired DNA segments into plasmids, this procedure has enabled scientists to amplify more than 1000 specific genes including those for human interferon, insulin, and growth hormone. Part B of the exercise requires a centrifuge that can be ...
No, RNase A should not be omitted from buffer P1. It is required to prevent RNA contamination of the purified plasmid DNA. RNase A will not interfere with downstream in-vitro transcription experiments, since it will be efficiently removed during the plasmid purification procedures using QIAGEN Plasmid Kits ...
In article ,[email protected][142.20.8.21], jlight at RESUNIX.RI.SICKKIDS.ON.CA writes: ,From: jlight at RESUNIX.RI.SICKKIDS.ON.CA ,Subject: Denaturation of plasmid DNA ,Date: 30 Nov 1995 16:52:45 -0800 ,Heres a basic question for which I should know the answer. Basically, Im ,working with a mixed population of cDNAs cloned into a plasmid and I want ,to anneal a primer (with an affinity tag) to the specific constructs in ,solution to try and recover the homologous constructs intact. When you ,want to separate the strands of plasmid DNA and you have mostly ccc DNA ,(say, from a Qiagen prep), whats the best way to separate the strands only ,partially so that when you reanneal, one strand from any particular plasmid ,will reanneal with the other strand from the same plasmid molecule, rather ,than those of other homologous plasmid molecules? Please dont flame me if ,you think this is stupid - Im trying to be serious. Thanks in advance for ,your help. I really do not think that it is ...
Plasmid pCLIPf-NK1R Control Plasmid from Dr. Ana Eganas lab contains the insert Neurokinin-1 Receptor, Truncated and is published in Unpublished This plasmid is available through Addgene.
This video explain plasmid cloning. I am also adding here background information: Plasmids are circular, double-stranded DNA (dsDNA) molecules that are separate from a cells chromosomal DNA. These...
Plasmid constructions.The plasmids used for mapping assays were constructed as described below. All of the plasmids were sequenced to verify the expected genotypes.. Full-length UL31 was cloned into the pCDNA3 vector such that UL31 expression was driven by the human cytomegalovirus immediate early promoter-enhancer. The construct was designated pJB261.. The full-length UL34 sequence (encoding amino acids [aa] 1 to 275) was PCR amplified from HSV-1(F) viral DNA by use of the primers 5′ GTA GTC GAC ATA TGG CGG GAC TGG GCA AG 3′ and 5′ GCG GTC GAC AGG GCT GTG TGG GGC GAA GGC GTC 3′. The PCR product was then removed with SalI and ligated into the pGEX4T-1 SalI restriction site. This plasmid was designated pJB253. The UL34 fragment was then cut from pJB253 with the SalI enzyme and ligated into the XhoI site of pCDNA3, and the construct was designated pJB280a. To include sequences sufficient to promote translation of the gene, we PCR amplified a full-length UL34 fragment from pJB253 by using ...
Vector properties of plasmid pNH602, a higher-copy-number deletion mutant of plasmid R6K, were tested by cloning the 6.5 Mg/molBam HI pSa fragment carrying determinants of resistance to four antibiotics in the uniqueBam HI site of pNH602. The resultingin vitro constructed recombinant plasmid pNH606 was found to be stable, conjugative, multicopy (20 copies of pNH606 perE. coli chromosome were estimated) and to ensure the increased expression of different genes responsible for the antibiotic resistance. The pSa fragment inserted in theBam HI site of plasmid pNH602 (located in Tn2660) was proved to be transposable to other replicons. Recombinant plasmid pNH606 was analyzed using restriction enzymesBam HI andEco RI and its physical and genetic map was constructed.
This exercise was designed to provide an exciting introduction to specific gene structure and function. In the experiment, students are given two plasmids (A and B) which are identified in the instructors guide. One plasmid (A) has a functional gene for the enzyme ß-galactosidase. The ß-galactosidase gene in the other plasmid (B) is inactive because it contains a segment of foreign DNA. In the first part of the exercise, students analyze restriction digests of both plasmids in order to determine which plasmid should have a functional ß-galactosidase gene. In the second part of the exercise, the plasmids are introduced into E. coli by transformation and the color of the resulting colonies (blue or white) is then used to assess the functional status of the ß-galactosidase gene. This exercise can be completed within a single 3-hour laboratory session or two 2-hour laboratory periods.. ...
GENETIC BASIS OF RESISTANCE:. Most drug resistance is due to a genetic change in the organism, either as a chromosomal mutation or the acquisition of a plasmid ortransposon. a) CHROMOSOME-MEDIATED-RESISTANCE:. Chromosomal resistance is due to a mutation in the gene that codes for either the target of the drug or the transport system in the membrane that controls the uptake of the drug. The frequency of spontaneous mutations usually ranges from 10~ 7 to 10 ~ 9 which is much lower then the frequency of acquisition of resistance plasmids. Therefore, chromosomal resilience is less of a clinical problem than plasmid. Mediated resistance. b) PLASMID-MED1ATED RESISTANCE:. Plasmid-mediated resistance is very important from clinical point of view for 3 reasons: 1) It occurs in many different species, especially gram negative rods. 2) Plasmida frequently madiate resistance to multiple drugs. 3) Plasmids have a high rate of trans. for from one cell to another, usually by conjugation. Resistance plasmids ...
Study Flashcards On restriction enzymes/plasmids at Cram.com. Quickly memorize the terms, phrases and much more. Cram.com makes it easy to get the grade you want!
QIAGEN has established a partnership with Aldevron, a company that specializes in custom plasmid DNA preparation services. Through this partnership, Aldevron is offering customers the possibility to order standard plasmid DNA preparation services performed using high-quality QIAGEN plasmid purification products. This service offers three different scales of endotoxin-free* plasmid DNA. This standardized service is only available via the Web site |span style=text-decoration: underline;>www.plasmid.com|/span>. |br /> |br /> Learn more:
February 8, 2011 at 2:28 pm , nick , cool results, everyday science Few experiments in science are conclusive. So, it is very exciting when an experiment provides completely unambiguous results. Today that happened.. Molecular biologists use bacteria-specifically strains of E. coli that dont make people sick (i.e., non-pathogenic)-to make lots of copies of circular DNA, called plasmids; one can think of E. coli as a photocopier for plasmids. Bacteria love to take up plasmids, copy them, and express them to make proteins with unique functions, and it is this ability that makes them so evolutionarily successful. Bacteria can transfer plasmids containing antibiotic resistance genes to each other, for example, and in this manner, can become "superbugs." Staph is one pathogen that has done this so successfully in hospitals that we will soon run out of antibiotics to treat it.. Interestingly, molecular biologists give E. coli antibiotic resistance genes on purpose. Its not because were ...
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InvivoGen provides two different E. coli competent strains : GT115 cells are specifically designed for cloning and propagation of plasmids containing hairpin structures and the R6K gamma origin of replication, such as pCpG-siRNA plasmids, GT116 cells are specifically designed for cloning and propagation of shRNA-expressing plasmids which contain hairpin structures, such as psiRNA plasmids.
Hi, in this agarose gel photo (70 volts, 3.5uL Gel Red in 30ml of TBE, 1% Agarose, 1 hour run), Ive checked the digestion of a large plasmid (15Kbp, one cut site). The theory says that the digested plasmid (linearized) should migrate slower than the supercoiled form of the uncut one (bigger band of the Undigested Plasmid). In my case the digested plasmid is faster than the undigested one. Ive found that my digested plasmid could be single-stranded (http://bitesizebio.com/13524/how-to-identify-supercoils-nicks-and-circles-in-plasmid-preps/). Do you think that could be the case?. ...
OriGene offers the largest collection of cDNA expression plasmids, ready for transfecting into mammalian cells. Genome-wide coverage of tagged ORF vectors, non-tagged genes and lentiviral vectors.
This could be a few things: 1. Bad antibiotic - check to see if bacteria that dont harbor the same resistance will grow/not grow in the media or on the plate. 2. Something wrong with the method of DNA prep - try to prep other plasmids using the same kit/reagents. Try to prep plasmids with the same antibiotic resistance. 3. No plasmid to prep - Use a method of colony cracking where you take some of the culture, pop the bacteria and run it straight on a gel without prepping for clean plasmid. This will let you know if anything is there to prep. In general, clones should not be able to grow on a particular antibiotic if there is no plasmid expressing resistance ...
The mKate2 Optimization plasmid is similar to the Edit-R Cas9 nuclease expression plasmid with the Blasticidin resistance marker, where the mKate2 fluorescent gene sequence has replaced the Cas9 nuclease gene. The mKate2 Optimization plasmid can be used to perform transfection optimization in a similar manner as described in the ��Transfection Optimization�� section of the technical manual. However, if you are working with a SMARTCas9 nuclease expression plasmid, then we recommend performing transfection optimization with a corresponding SMARTCas9-mKate2 expression plasmid under the control of a promoter you know to be active in your cells ...
Chapter 19) (a) Minimal defective prophage created by removal of the N through kil genes in the P L operon and replacement of rexA and rexB with a drug resistance cassette, either ampicillin (bla) or chloramphenicol (cat). With this system, raising the temperature induces the operon directly without N-mediated antitermination. (b) Minimal prophage moved onto a high-copy-number vector. The pBR322 origin of DNA replication between nucleotide coordinates 2348 and 3296 was amplified.This linear PCR product was used to clone the minimal prophage in a gap repair reaction. This linear pBR322 fragment contains ori but lacks an antibiotic resistance gene, so only those plasmid clones that have undergone successful recombineering will contain an antibiotic resistance marker inherited from the prophage.The high-copy-number plasmids thus generated, pSIM2 and pSIM4 (Cmr and Ampr, respectively),were used as targets in subsequent recombineering reactions.The pBR322 segment was replaced precisely with a linear ...
5121 Telomeres, centromeres, and other highly repetitive genomic regions are typically under-represented in shotgun libraries, and they are often absent from partial-digest BAC libraries. Accurate cloning of isolated fragments containing short tandem repeats, AT-rich DNA, or regions with strong secondary structure likewise can be difficult or impossible using conventional supercoiled plasmids. We have developed vectors and methods to alleviate these types of cloning bias. A transcription-free, linear plasmid was derived from the linear coliphage N15 for cloning into E.coli. The ends of the vector are free to rotate during replication, minimizing formation of secondary structures that are substrates for deletion or rearrangement. The "pJAZZ" linear vectors are shown to provide unprecedented ability to maintain regions of up to 30 kb that are unclonable in circular plasmids. Examples include regions of di-, tri-, and tetra-nucleotide repeats up to several kb, as well as highly AT-rich fragments of ...
Wein, Tanita, Hülter, Nils F., Mizrahi, Itzhak and Dagan, Tal (2019) Emergence of plasmid stability under non-selective conditions maintains antibiotic resistance Nature Communications, 10 (1). DOI 10.1038/s41467-019-10600-7. Full text not available from this repository ...
... : Found or purchased all 11 basic Plasmid types - worth 20 GamerScore. Find guides to this achievement here.
Plasmids can be submitted in either form. For DNA, aliquot 15 µL of DNA into a 1.5 mL microfuge tube (at a concentration of 0.1 -1µg/µL)...
Description: Small toxic membrane polypeptide, Qin prophage; homologous to plasmid-encoded plasmid stabilization toxins regulated by antisense RNA; functional relevance of chromosomal homologs is ...
Agarosegelelectrophoresis with the digestions (CMV, eGFP, pDS7, 190-6), 125V for 30 minutes and then for 20 minutes; expected DNA bands: 190-6 (4840bp, 1903bp), pDS7 (8027bp, 6bp), CMV (654 bp (Insert), 2079bp (Plasmid)), eGFP (720bp (Insert), 2750bp (Plasmid)) - Correct DNA bands for 190-6 (~4800bp, ~1900bp, ~6700bp (undigested plasmid)) and eGFP (~2000bp (Plasmid), ~750 bp (Insert)); CMV probably not digested (two bands; one probably normal, one supercoiled) and pDS7 not clear - Restriction digest from CMV (EcoR1, Pst1; 6µl DNA, buffer H) and pDS7 (EcoR1, Spe1; 2µl DNA, buffer B) Expected DNA bands: CMV see above, pDS7 (3647bp, 3369bp, 1011bp, 6bp) -, Protocol (5 Restriction digest) - Agarosegelelectorphoresis for 30 minutes, 150V ...
Basically, plasmid DNA can be isolated from cells on the basis of size and their conformation. As the plasmid DNA is smaller than chromosomal DNA.
Transfections were performed in six-well culture plates (Corning Inc., Corning, NY). Each well has approximately 10-cm2 surface for cell culture growth. Cells in each well were transfected with 2 μg of total plasmid DNA. To test the transactivating activity of wild-type and mutant hPXRs, the cells were transfected with 50 ng of pcDNA3, wild-type or mutant hPXR, and 1950 ng of pGL3-CYP3A4-luc plasmids. To examine the protein expression and localization, the cells were transfected with 2 μg of pcDNA3 and wild-type or mutant hPXR plasmids. For the rapamycin experiments, the cells were transfected with 50 ng of pcDNA3 or hPXR and 1950 ng of pGL3-CYP3A4-luc plasmids. In p70 S6K overexpression experiments, the cells were transfected with 100 ng of FLAG-pcDNA3, pcDNA3-FLAG-hPXR, or pcDNA3-FLAG-hPXRT57A; 500 ng of constitutively active p70 S6K; and 1000 ng of pGL3-CYP3A4-luc and 100 ng of pGL4-hRluc; FLAG-pcDNA3 was used to make up the total plasmid DNA to 2 μg in each transfection. In mammalian ...
A plasmid is a small DNA molecule that is physically separate from, and can replicate independently of, chromosomal DNA within a cell. Plasmids are commonly used to multiply (make many copies of) …
Vector ConstructionHuman Akt-Myr cDNA with myristolation signal for greater activity, also known as PKB, was cloned into the multiple cloning site of pAC-CMV-pLpA. The resulting plasmid was cotransfected into HEK293 cells with plasmid pJM17 which contains the Ad5 genome. Homologous recombination between the two plasmids resulted in the replacement of the Ad5 early region 1 with the human Akt cDNA expression cassette, generating a replication deficient recombinant virus. ...
Existing methods for cloning and recombination of DNA enable construction of arbitrary sequences. However, the sequential nature of these techniques makes them time-consuming and expensive. Furthermore, while the transformation of an existing plasmid into a host strain can be reliable when a selection marker is used, there are many current limitations: the number of different plasmids that can be co-transformed is limited by the choice of markers and compatible origins of replication; plasmids are less stable than chromosomal DNA and are difficult to maintain indefinitely without mutation; and cistronic interactions cannot be designed since each new nucleotide sequence added is on an unconnected DNA molecule. To overcome these limitations, we are designing reconfigurable chromosomes consisting of both fixed and variable regions. While the fixed region is carefully optimized and tuned ahead of time, the variable region can be modified in the field, at the point-of-use, leading to rapid and ...
Existing methods for cloning and recombination of DNA enable construction of arbitrary sequences. However, the sequential nature of these techniques makes them time-consuming and expensive. Furthermore, while the transformation of an existing plasmid into a host strain can be reliable when a selection marker is used, there are many current limitations: the number of different plasmids that can be co-transformed is limited by the choice of markers and compatible origins of replication; plasmids are less stable than chromosomal DNA and are difficult to maintain indefinitely without mutation; and cistronic interactions cannot be designed since each new nucleotide sequence added is on an unconnected DNA molecule. To overcome these limitations, we are designing reconfigurable chromosomes consisting of both fixed and variable regions. While the fixed region is carefully optimized and tuned ahead of time, the variable region can be modified in the field, at the point-of-use, leading to rapid and ...
Vector ConstructionHuman cDNA for apoptosis signaling kinase 1 with a mutation in catalytic lysine (presumed dominant negative). It was cloned into the multiple cloning site of pAC-CMV-pLpA. The resulting plasmid was cotransfected into HEK293 cells with plasmid pJM17 which contains the Ad5 genome. Homologous recombination between the two plasmids resulted in the replacement of the Ad5 early region 1 with the mouse cDNA for ASK1-KM, generating a replication deficient recombinant virus. ...
Transgenic mice. To make the 7X-tetOp A-FOS transgene, the plasmid pUHD 10-3 ( 19) was linearized with SacII and BamHI and ligated to a 62-bp polylinker: top strand, 5′-GGCCACCATGGCGTATCCCTACGACGTGCCCGATTATGCCCGATTATGCCCATATGCAGGAATTCAAGCTTG-3′ that encodes a Kozak consensus and a Hemagglutinin tag (YPYDVPDYA). An SV40 poly(A) fragment containing the small T-antigen intron that preceded the poly(A) site was obtained from the plasmid pRSVNeo as a 1,026-bp BamHI-SmaI fragment. This fragment, when ligated to the BamHI-NaeI-digested 2.7-kb vector (pTRE 850/85-ds) backbone, produced a plasmid with the SV40 poly(A) in reverse orientation (pTRE850/851 ds reversed). To obtain a plasmid with the SV40 poly(A) in the correct orientation, "pTRE850/851-ds-reversed" was digested with HindIII. The 2,704-bp vector and the 1,067-bp insert were then religated, and recombinants were selected in which the insert fragment [representing the SV40 poly(A)] is reversed when compared with the parental plasmid. This ...
To test whether ORF14 is cotranscribed with gumM, we employed a method consisting of the application of lacZtranscriptional fusions combined with plasmid integration (13). A promoterless lacZ-aacC1 interposon (4) was inserted into the unique BamHI site of the SmaI-ClaI fragment (nucleotides 14056 [GenBank accession no. U22511] and 2071 [GenBank accession no.U70053], respectively), which was previously subcloned into pK19mobGII. The hybrid plasmid was transferred from the broad-host-range mobilizing strain E. coli S17-1 (21) to wild-type X. campestris FC2 (14), as described previously (19). Exconjugants were selected in agar medium containing gentamicin, rifampin, and X-gluc. Yellow colonies appeared with a frequency of 10−4 and were sensitive to kanamycin. Correct gene replacement was verified by Southern hybridization. The strain mutated in ORF14 (XcORF14) produced normal amounts of xanthan, as judged by precipitation of the polymer from the broth as previously described (6) (not shown). ...
GenScripts industrial-grade Plasmid DNA can help your experiments achieve highly efficient cell transfection, helping to improve experimental outcomes in research areas such as protein expression, antibody production, and other research projects.
There are numerous methods for detecting protein-protein associations, each with advantages and disadvantages. Advantages of the FLS platform technology presented here are its versatility and simplicity with the capacity to evaluate proteins from many different sources yet without the need for purified proteins, unusual technical expertise, or highly specialized equipment. The most demanding requirement is for a fluorescence microscope with standard imaging capabilities. Regarding ease of use, it is also significant that this technology is designed to work upon transient expression of proteins from transfected plasmids and does not require the generation of stably expressing cell lines. Another advantage is that associations are visualized inside cells in a setting that approximates the native environment of the proteins. In addition to CV-1 and COS-7 cells, we have shown the technology to work as well in other cell types (BHK-21 and NIH 3T3) (data not shown) so that cell type restrictions seem ...
Read about how mammalian plasmids differ from their bacterial counterparts, including how replication occurs and whether selection is necessary for transfected cells.
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Miniprep plasmid DNA extraction kit is used when the starting E. coli culture volume is 1~5 ml of LB broth and the expected DNA yield is 20~30 μg. Other extraction and purification plasmid DNA Kits are available from varying manufacturers, named by size of bacterial culture, includes gigaprep, megaprep, and midiprep - Plasmid DNA Extraction (Miniprep) - AbVideo™ - Support - Abnova
Those sneaky file searchers over at the 2K Games forums have uncovered a few text strings embedded in the PC version of BioShocks game code which hint at a new set of plasmids coming our way. After looking through the games install files for a reference to a PC game editor, forum member Zemlor stumbled across a reference to a piece of content called PlasmidPack1. ...
Creative Biogene offers a wide range of plasmid preparation services for many applications, including research, preclinical, clinical, and diagnostic applications.
Bacterial plasmids are nucleotide sequences floating in the cytoplasm of bacteria. These molecules replicate independently from the main chromosomal DNA and are not essential to the survival or replication of their host. Plasmids are thought to be part of the bacterial domains mobilome (for overview, see Siefert, 2009), a sort of genetic commonwealth which most,…
The VTC4 gene (At3g02870) was amplified by PCR with primers 5′-ATGGCGGACAATGATTCTCT-3′ (forward) and 5′-TCATGCCCCTGTAAGCCGCA-3′ (reverse). The template was generated by RT (Omniscript RT kit) of RNA extracted from wild-type plants using the RNAeasy Plant Mini kit (both kits from Qiagen) according to the manufacturers instructions. The resulting PCR product was cloned into plasmid pCRT7/NT-TOPO using the pCR T7 TOPO TA Expression kit (Invitrogen). The newly created plasmid pAtIMPH contains the amplified At3g02870 gene (816 nucleotides) from the start ATG to the stop TGA along with upstream plasmid regions including a T7 promoter, ribosome binding site, ATG start site, 6xHIS region, Xpress epitope, and EK cleavage site. Because of the extra upstream plasmid sequences, the theoretical size of the protein is 36 amino acids longer than the 271 amino acids of IMP. pAtIMPH was used to transform TOP10F′ Escherichia coli cells, and the gene sequence was verified. Similarly, plasmids containing ...
RNase, Purification, Plasmid DNA, CIM® Columns, Plasmids, episomes, eukaryotic, prokaryotic, genetic vectors, gene manipulation, ribonuclease A, hydrophobic chromatography, ion-exchange, size exclusion, affinity
Bars = estimated 16S copies based on nicked-circular plasmid standard. Black and white striped bars = estimated 16S copies based on linearized plasmid standard.
Domain architecture and assignment details (superfamily, family, region, evalue) for gi|532433720|ref|YP_008470220.1|NC_022123 from NCBI plasmid sequences. Plus protein sequence and external database links.
C-terminal 6His and EGFP dual affinity and reporter tag yeast expression plasmid. This vector allows the creation of fusion proteins with an EKT cleavage tag.
Clone #1 was selected from each of the screened clones.. A sterile pipette tip was used to inoculate 5mL of 1x LBAmp100 in a 15mL conical tube. The tubes were incubated O/N @ 37C on a rocker.. 3mL of liquid culture was used as input for plasmid isolation with the QIAprep Spin Mini Kit (Qiagen) according to the manufacturers protocol.. 1mL of each culture was combined with 1mL of 50% sterile glycerol (25% glycerol final concentration) and stored @ -80C with existing bacterial stocks.. Plasmid DNA was eluted with 50μL of Buffer EB and quantified on the Roberts Lab NanoDrop1000 (ThermoFisher) in order to have a rough idea of concentrations to submit for Sanger sequencing. A dye-based quantification will be performed after sequencing results are back in order to obtain a more accurate assessment for use in ISH and/or qPCR standard curve creation.. Results:. ...
Cell Biologics provides custom services to our customers upon request. If you have any special needs in molecular biology products, please contact us: [email protected] Thank you!. Cell Biologics. ...
Cell Biologics provides custom services to our customers upon request. If you have any special needs in molecular biology products, please contact us: [email protected] Thank you!. Cell Biologics. ...
Biotechnical production processes often operate with plasmid-based expression systems in well-established prokaryotic and eukaryotic hosts such as Escherichia coli or Saccharomyces cerevisiae, respectively. Genetically engineered organisms produce important chemicals, biopolymers, biofuels and high-value proteins like insulin. In those bioprocesses plasmids in recombinant hosts have an essential impact on productivity. Plasmid-free cells lead to losses in the entire product recovery and decrease the profitability of the whole process. Use of antibiotics in industrial fermentations is not an applicable option to maintain plasmid stability. Especially in pharmaceutical or GMP-based fermentation processes, deployed antibiotics must be inactivated and removed. Several plasmid addiction systems (PAS) were described in the literature. However, not every system has reached a full applicable state. This review compares most known addiction systems and is focusing on biotechnical applications.. ...
TY - JOUR. T1 - Use of transmissible plasmids as cloning vectors in Caulobacter crescentus. AU - Schoenlein, Patricia V. AU - Gallman, Lilly M.. AU - Ely, Bert. PY - 1988/10/30. Y1 - 1988/10/30. N2 - Cloning vectors for studies of Caulobacter crescentus genes should be transferable between Escherichia coli and C. crescentus since a transformation system has not been developed for C. crescentus. We have tested a large number of vectors containing IncP or IncQ replicons and found that many of the vectors containing IncQ replicons, and all but one of the vectors containing IncP replicons, are readily transferred by conjugation into C. crescentus. All of the plasmids tested were maintained in C. crescentus at 1 to 5 copies per cell, but plasmids containing IncP replicons were more stable than plasmids containing IncQ replicons. Further studies with a derivative of the IncQ plasmid R300B showed that when a promoterless kanamycin (Km)-resistance gene (npt2) was inserted into the intercistronic region ...
TY - JOUR. T1 - Large plasmids in ruminal strains of Selenomonas ruminantium. AU - Fliegerova, K AU - Benada, O AU - Flint, H J PY - 1998/4. Y1 - 1998/4. N2 - The plasmid content of six different isolates of Selenomonas ruminantium from the rumen of sheep, cows or goats was examined by electron microscopy. In addition to small plasmids (, 12 kb) studied previously, all six strains contained at least one plasmid larger than 20 kb. Plasmid sizes of 1.4, 2.1, 2.4, 5.0, 6.2, 20.4, 20.8, 22.7, 33.3, 29.3, 30.7, 34.4 and 42.6 kb were estimated from contour length measurements. DNA-DNA hybridization experiments revealed homology among the large plasmids from five strains, while the 20.8 kb plasmid from a sixth isolate showed no apparent relationship with the plasmids of the other strains.. AB - The plasmid content of six different isolates of Selenomonas ruminantium from the rumen of sheep, cows or goats was examined by electron microscopy. In addition to small plasmids (, 12 kb) studied previously, ...
Plasmids play a key role in the genetic plasticity and survival of Staphylococcus aureus in challenging environments. Although many S. aureus plasmids have been described, still few studies portray the plasmid content of a given S. aureus population. The aim of this work was to characterize the plasmids carried by a collection of 53 S. aureus isolates collected in a large hospital in Lisbon, Portugal, and investigate their role in conferring resistance to several antimicrobial agents. Plasmids were present in 44 out of the 53 isolates and were grouped into eleven AccI restriction profiles. Plasmid curing of representative strains and comparison of antimicrobial susceptibility profiles between pairs of isogenic strains proved to be a valuable guidance tool in the identification of plasmid-located resistance genes. The plasmids harbored several resistance genes, namely blaZ (resistance to β-lactams), erm(C) (resistance to macrolides, lincosamides, and streptogramin B), cadA (resistance to cadmium ...
TY - JOUR. T1 - Plasmids of the pRM/pRF family occur in diverse Rickettsia species. AU - Baldridge, Gerald D. AU - Burkhardt, Nicole Y.. AU - Felsheim, Roderick F.. AU - Kurtti, Timothy J. AU - Munderloh, Ulrike G. PY - 2008/2/1. Y1 - 2008/2/1. N2 - The recent discoveries of the pRF and pRM plasmids of Rickettsia felis and R. monacensis have contravened the long-held dogma that plasmids are not present in the bacterial genus Rickettsia (Rickettsiales; Rickettsiaceae). We report the existence of plasmids in R. helvetica, R. peacockii, R. amblyommii, and R. massiliae isolates from ixodid ticks and in an R. hoogstraalii isolate from an argasid tick. R. peacockii and four isolates of R. amblyommii from widely separated geographic locations contained plasmids that comigrated with pRM during pulsed-field gel electrophoresis and larger plasmids with mobilities similar to that of pRF. The R. peacockii plasmids were lost during long-term serial passage in cultured cells. R. montanensis did not contain a ...
Tetracyclines have been among the most widely used antibiotics worldwide. Plasmid-mediated tetracycline resistance among hospital strains of bacteria has continued to rise and of major concern has been the transfer of resistance to pathogenic organisms. Bacteraemia due to hospital acquired S. typhimurium has been a major cause of morbidity at Kenyatta National Hospital (KNH), hence the need to study drug susceptibility pattern of this organism. This study also characterized the tetracycline resistance genes using oligonucleotide probes. Ninety seven S. typhimurium strains isolated from patients at KNH were used. Agar dilution method was used to determine minimum inhibitory concentration (MIC). Plasmids were isolated from each strain and the different plasmid profiles were grouped by their molecular weights into 6 patterns. Out of 97, 87 (88%) strains were resistant. MIC ranged from 1 microgram/ml to 128 micrograms/ml. Genes encoding for tetracycline resistance were located on plasmids of ...
Summary. Enterotoxigenic Escherichia coli (ETEC) strains of serotype O153:H45 have been found recently to be a frequent cause of sporadic cases and outbreaks of neonatal diarrhoea in Spain and the most important cause of infant diarrhoea in Chile. Relationships between sugar fermentation patterns, resistance to antibiotics and plasmid profiles were analysed in nine E. coli O153:H45 strains isolated in Spain that synthesised CFA/I antigen and STa enterotoxin. Derivative strains obtained by curing with acridine orange, and transconjugants rendered antibiotic resistant, were characterised phenotypically and analysed for plasmid content. Two fermentation patterns were recognised: rhamnose fermenters (four strains) and rhamnose non-fermenters (five strains). The ability to ferment rhamnose was the only differential characteristic found among 49 carbohydrate fermentation tests used to establish fermentation patterns. All nine strains possessed similar plasmid profiles of three or four plasmids of 52-87 Mda. A
Metapyrocatechase which catalyzes the oxygenative ring cleavage of catechol to form alpha-hydroxymuconic epsilon-semialdehyde is encoded by the xylE gene on the TOL plasmid of Pseudomonas putida mt-2. We have cloned the xylE region in Escherichia coli and determined the nucleotide sequence of the DNA fragment of 985 base pairs around the gene. The fragment included only one open translational frame of sufficient length to accommodate the enzyme. The predicted amino acid sequence consisted of 307 residues, and its NH2- and COOH-terminal sequences were in perfect agreement with those of the enzyme recently determined (Nakai, C., Hori, K., Kagamiyama, H., Nakazawa, T., and Nozaki, M. (1983) J. Biol. Chem. 258, 2916-2922). A mutant plasmid was isolated which did not direct the synthesis of the active enzyme. This plasmid had a DNA region corresponding to the NH2-terminal two-thirds of the polypeptide. From the deduced amino acid sequence, the secondary structure was predicted. Around 10 base pairs upstream
Looking for online definition of chloramphenicol acetyl transferase in the Medical Dictionary? chloramphenicol acetyl transferase explanation free. What is chloramphenicol acetyl transferase? Meaning of chloramphenicol acetyl transferase medical term. What does chloramphenicol acetyl transferase mean?
Drug Resistance Plasmids[edit]. Research done on Lactobacillus fermentum strains has revealed the existence of tetracycline and ... "Drug Resistance Plasmids in Lactobacillus Fermentum." Journal of General and Applied Microbiology 26, no. 1 (1979): 71-74. ... erythromycin resistance plasmids.[15] Sensitivity to Antibiotics[edit]. While Lactobacillus fermentum has been found to have ...
Construction of expression plasmids[edit]. A number of engineered genetic sequences must be incorporated into the host cell to ... The reporter gene may be inserted into the E. coli genome by first inserting it into an episome, a type of plasmid with the ... Plasmids are engineered to produce a protein product in which the DNA-binding domain (BD) fragment is fused onto a protein ... This mutant yeast strain can be made to incorporate foreign DNA in the form of plasmids. In yeast two-hybrid screening, ...
For plasmid delivery[edit]. Individual genes can be inserted into specific sites on plasmids, and recombinant plasmids can be ... A method using macro-branched TAT has been proposed for plasmid DNA delivery into various cell lines and showed significant ... Nucleic acid-based macromolecules such as siRNA, antisense oligonucleotide, decoy DNA, and plasmid have been realized as ... "Oligomers of the Arginine-rich Motif of the HIV-1 TAT Protein Are Capable of Transferring Plasmid DNA into Cells". Journal of ...
SPINACH PLASMIDS. ...
Smaller independent pieces of DNA, called plasmids, are also found in archaea. Plasmids may be transferred between cells by ... Sota M; Top EM (2008). "Horizontal Gene Transfer Mediated by Plasmids". Plasmids: Current Research and Future Trends. Caister ... ISBN 978-1-904455-27-1. Lipps G (2008). "Archaeal Plasmids". Plasmids: Current Research and Future Trends. Caister Academic ... Schleper C; Holz I; Janekovic D; Murphy J; Zillig W (1 August 1995). "A multicopy plasmid of the extremely thermophilic ...
Plasmids containing the Factor IX gene, along with plasmids with a gene that codes for resistance to methotrexate, were ... The recombinant plasmids were inserted into Escherichia coli bacteria, which were "induced to produce 100,000 molecules of ... The gene that was inserted into the plasmid was created by reverse transcription of the mRNA found in pituitary glands to ... The artificial genes were "then inserted... into plasmids... among a group of genes that" are activated by lactose. Thus, the ...
It is used to increase the transfection efficiency of RNA (including mRNA and siRNA) or plasmid DNA into in vitro cell cultures ... DNA plasmids. Lipofectamine is a cationic liposome formulation, which complexes with negatively charged nucleic acid molecules ...
Cells harboring the plasmid will survive when exposed to the antibiotic, while those that have failed to take up plasmid ... "plasmid / plasmids , Learn Science at Scitable". www.nature.com. Retrieved 2017-12-06. Shizuya H, Birren B, Kim UJ, Mancino V, ... and a plasmid cloning vector. E. coli and plasmid vectors are in common use because they are technically sophisticated, ... That is, these plasmids could serve as cloning vectors to carry genes. Virtually any DNA sequence can be cloned and amplified, ...
Fertility plasmids, or f plasmids, allow for conjugation to occur whereas resistance plasmids, or r plasmids, contain genes ... Circular bacterial plasmids are classified according to the special functions that the genes encoded on the plasmid provide. ... Circular bacterial plasmids are also the basis for the production of DNA vaccines. Plasmid DNA vaccines are genetically ... The linear plasmids with a covalently attached protein may assist with bacterial conjugation and integration of the plasmids ...
Plasmids in Bacteria. Boston, MA: Springer US. ISBN 1-4613-2447-5. CS1 maint: Extra text: authors list (link) Roberts, edited ...
Plasmid. 61 (3): 171-175. doi:10.1016/j.plasmid.2008.12.002. Labes, G.; Bibb, M.; Wohlleben, W. (1 May 1997). "Isolation and ... Plasmid. 61 (3): 171-175. doi:10.1016/j.plasmid.2008.12.002. ATCC Type strain of Streptomyces ghanaensis at BacDive - the ... Muth, Gunter; Wohlleben, Wolfgang; Pohler, Alfred (March 1988). "The minimal replicon of the Streptomyces ghanaensis plasmid ... Plasmids in Bacteria. Boston, MA: Springer US. ISBN 1-4613-2447-5. CS1 maint: Extra text: authors list (link) ed.-in-chief, ...
As of 2014 Addgene's repository comprised 30,000 plasmids, deposited by 1,700 labs. Its plasmid collection contains plasmids ... Plasmid Cloning Guides Molecular Cloning Guides-References to help scientists design plasmid cloning experiments, including ... The plasmids are available to both academic and industry labs. Noteworthy depositors include:[citation needed] 12 Nobel Prize ... Addgene accepts plasmids from researchers for distribution and archival. The organization covers the operating costs of ...
... /Cas9 often employs a plasmid to transfect the target cells. The main components of this plasmid are displayed in the ... Each repetition is followed by short segments of spacer DNA from previous exposures to foreign DNA (e.g., a virus or plasmid). ... "CRISPR/Cas9 Plasmids". www.systembio.com. Retrieved 2015-12-17. "CRISPR Cas9 Genome Editing". www.origene.com. OriGene. ... A 2010 study showed that CRISPR-Cas cuts both strands of phage and plasmid DNA in S. thermophilus. Researchers studied a ...
Plasmids in Bacteria. Boston, MA: Springer US. ISBN 1-4613-2447-5. CS1 maint: Extra text: authors list (link) compiled, ...
The RNA world hypothesis is supported by RNA's ability to store, transmit, and duplicate genetic information, as DNA does. RNA can act as a ribozyme, a special type of enzyme. Because it can perform the tasks of both DNA and enzymes, RNA is believed to have once been capable of supporting independent life forms.[15] Some viruses use RNA as their genetic material, rather than DNA.[45] Further, while nucleotides were not found in experiments based on Miller-Urey experiment, their formation in prebiotically plausible conditions was reported in 2009;[22] the purine base known as adenine is merely a pentamer of hydrogen cyanide. Experiments with basic ribozymes, like Bacteriophage Qβ RNA, have shown that simple self-replicating RNA structures can withstand even strong selective pressures (e.g., opposite-chirality chain terminators).[46] Since there were no known chemical pathways for the abiogenic synthesis of nucleotides from pyrimidine nucleobases cytosine and uracil under prebiotic conditions, it ...
Plasmids. The use of plasmids has been validated in preclinical studies as a protective vaccine strategy for cancer and ... The overall efficacy of plasmid DNA immunization depends on increasing the plasmid's immunogenicity while also correcting for ... "Plasmids: Current Research and Future Trends. Caister Academic Press. ISBN 978-1-904455-35-6. .. ... "Plasmid DNA as Prophylactic and Therapeutic vaccines for Cancer and Infectious Diseases" ...
Kandavelou K, Chandrasegaran S (2008). "Plasmids for Gene Therapy". In Lipps, Georg. Plasmids: Current Research and Future ...
Kandavelou K, Chandrasegaran S (2008). "Plasmids for Gene Therapy". Plasmids: Current Research and Future Trends. Caister ... Since ZFN-encoding plasmids could be used to transiently express ZFNs to target a DSB to a specific gene locus in human cells, ... The ZFN-encoding plasmid-based approach has the potential to circumvent all the problems associated with the viral delivery of ... The construction of plasmid vectors is simple and straightforward. Custom-designed ZFNs that combine the non-specific cleavage ...
"Geraldine Seydoux Lab Plasmids". Addgene. Retrieved 2016-04-30. "CDB Symposium 2007 : Speaker Profiles". Cdb.riken.go.jp. ...
Maia M.; Sanchez J. M.; Vela G. R. (1988). "Plasmids of Azotobacter vinelandii". Journal of Bacteriology. 170 (4): 1984-1985. ... In addition to chromosomal DNA, Azotobacter can contain plasmids. Azotobacter species are ubiquitous in neutral and weakly ...
The P4 genome can also exist on its own within the host cell and can replicate as a free plasmid. Pruss, G; Goldstein, RN; ... Briani, Federica; Dehò, Gianni; Forti, Francesca; Ghisotti, Daniela (2002). "The Plasmid Status of Satellite Bacteriophage P4 ... ". Plasmid. 45 (1): 1-17. doi:10.1006/plas.2000.1497. PMID 11319927. Claverie J-M, Abergel C (2009). "Mimivirus and its ...
Construction of plasmid R6K derivatives in vitro: characterization of the R6K replication region. Plasmid. 1978 Sep;1(4):571-80 ... Plasmid R6K DNA replication. II. Direct nucleotide sequence repeats are required for an active gamma-origin. J Mol Biol. 1982 ... Trans-complementation-dependent replication of a low molecular weight origin fragment from plasmid R6K. Cell. 1978 Dec;15(4): ...
Cleavage of a plasmid in Halobacterium halobium.resulted in the loss of the ability to biosynthesize gas vesicles, indicating ... doi:10.1016/0147-619x(79)90021-0. DasSarma, S.; Damerval, T.; Jones, J. G.; Marsac, N. Tandeau de (1987-07-01). "A plasmid- ... Although there is evidence suggesting the early evolution of gas vesicles, plasmid transfer serves as an alternate explanation ... Weidinger, Gottfried; Klotz, Günther; Goebel, Werner (July 1979). "A large plasmid from Halobacterium halobium carrying genetic ...
... faecalis plasmid pCF10 conjugation". Plasmid. 64 (1): 26-35. doi:10.1016/j.plasmid.2010.03.002. PMC 2892192 . PMID 20332003. ... Anti-Q RNA (formerly Qa RNA) is a small ncRNA from the conjugal plasmid pCF10 of Enterococcus faecalis. It is coded in cis to ... Brantl S, Birch-Hirschfeld E, Behnke D (1993). "RepR protein expression on plasmid pIP501 is controlled by an antisense RNA- ... Novick RP, Iordanescu S, Projan SJ, Kornblum J, Edelman I (1989). "pT181 plasmid replication is regulated by a ...
Plasmid. 33 (3): 235-8. doi:10.1006/plas.1995.1026. PMID 7568472. Hsu MY, Inouye M, Inouye S (December 1990). "Retron for the ...
Nabeshima, S; Hotta, Y; Okanishi, M (September 1984). "Construction of plasmid vectors from Streptomyces kasugaensis plasmids, ... Toyama, Hiromi; Okanishi, Masanori; Umezawa, Hamao (May 1981). "Physical characterization of plasmids from Streptomyces ... kasugaensis MB273". Plasmid. 5 (3): 306-312. doi:10.1016/0147-619X(81)90007-X. Chae, Won-Bok; Kim, Young-Bum; Choi, Sung-Won; ...
Terminal Inverted Repeat Linear Plasmid Plasmid Transfer Mycobacterium Smegmatis Conjugative Plasmid These keywords were added ... Holloway, B. W., 1979, Plasmids that mobilize bacterial chromosome, Plasmid 2: 1-19.PubMedCrossRefGoogle Scholar ... Hopwood, D. A., Lydiate, D. J., Malpartida, F., and Wright, H. M., 1985, Conjugative plasmids in Streptomyces, in: Plasmids in ... a conjugative plasmid will mobilize nontransferable plasmids in soil, Curr. Microbiol. 19: 115-121.Google Scholar ...
Plasmid-mediated resistance is the transfer of antibiotic resistance genes which are carried on plasmids. The plasmids can be ... Other smaller plasmids (usually < 10 kb in size) can be mobilized by a conjugative plasmid (usually > 30 kb) in order to be ... are common for resistance plasmids in Enterobacteriaceae. Often multiple beta-lactamase genes are found on the same plasmid ... It is very common for the resistance genes or entire resistance cassettes to be re-arranged on the same plasmid or be moved to ...
... everything you need for studying or teaching Plasmid. ... Immediately download the Plasmid summary, chapter-by-chapter ... Plasmid Plasmids are naturally occurring, stable genetic elements found in bacteria, fungi, and even in the mitochondria of ... Plasmids Plasmids are extra-chromosomal, covalently closed circular (CCC) molecules of double stranded (ds) DNA that are ... Plasmids Plasmids are extra-chromosomal, covalently closed circular (CCC) molecules of double stranded (ds) DNA that are ...
... Kevin ODonnell odonnell at sasa.gov.uk Wed Dec 13 12:22:28 EST 1995 *Previous message: TOL plasmids ... the presence of such a plasmid. Does anybody have a protocol that will ,let me visualize such a plasmid on a gel? ( ... In article ,4ai9os$up at news2.ucsd.edu,, rcaspi at sdcc3.ucsd.edu (Ron Caspi) says: , ,Hi, is there a TOL plasmid expert out ... unfortunately, pulse field ,is not available). , ,Many thanks in advance , I did my PhD on a TOL plasmid from Pseudomonas ...
... the End jgraham at bronze.ucs.indiana.edu Fri Dec 11 10:10:11 EST 1992 *Previous message: LARGE PLASMIDS ... Ian, Ive had much luck sith a 11.5 Kb low copy (pSC101) plasmid. I grow in TB terrific broth as described in the new ...
Plasmids are circular deoxyribonucleic acid (DNA) molecules that replicate independently of the bacterial chromosome. They are ... Plasmid, in microbiology, an extrachromosomal genetic element that occurs in many bacterial strains. ... Plasmid, in microbiology, an extrachromosomal genetic element that occurs in many bacterial strains. Plasmids are circular ... small extra DNA molecules called plasmids. Bacterial plasmids are circular DNA molecules; some carry genes for resistance to ...
Plasmids + copies of the DNA fragment produce quantities of recombinant DNA.. This recombinant DNA stew is allowed to transform ... The plasmid is opened up and the gene is freed from its parent DNA strand. They have complementary sticky ends. The opened ... In DNA cloning, a DNA fragment that contains a gene of interest is inserted into a cloning vector or plasmid. ... The plasmid carrying genes for antibiotic resistance, and a DNA strand, which contains the gene of interest, are both cut with ...
... and design complex plasmids for molecular biologists. ... Planning Plasmids Message Subject. (Your Name) has forwarded a ...
The result is plasmids consisting of genes that have each been adapted to different bacterial species. This facilitates further ... Antibiotic resistance-carrying plasmids from different bacteria can meet and exchange genetic material. ... Antibiotic resistance-carrying plasmids from different bacteria can meet and exchange genetic material. The result is plasmids ...
Plasmids Blast Server. Find out more about wu-blast Retrieve result for BLAST job id:. ...
Comprehensive and highly practical, E. coli Plasmid Vectors offers those new to the field a basic guide to the use of plasmid ... The authors describe readily reproducible methods for cloning DNA into plasmid vectors, transforming plasmids into E. coli, and ... In E. coli Plasmid Vectors, experienced bench researchers describe their proven techniques for the manipulation of recombinant ...
Scientists have succeeded in creating the first organism with alien DNA. In normal DNA, which can be found within the genes of every organism , the twin strands of the double helix are bonded together with four bases, known as T, G, A, and C. In this new organism, the researchers added two new bases, X and Y, creating a new form of DNA that has never occurred in billions of years of evolution on Earth or elsewhere in the universe. Remarkably, the semi-synthetic alien organism continued to reproduce normally, preserving the new alien DNA during reproduction. In the future, this breakthrough should allow for the creation of highly customized organisms - bacteria, animals, humans - that behave in weird and wonderful ways that mundane four-base DNA would never allow.. ...
My plasmids have only the 2u replication origin though, also pretty common. I transformed them with the LEU plasmid first and ... 2-micron plasmids. I was always under the impression that yeast could maintain ,only 1 2-micron plasmid - that if you ... My leu2, ura3 yeast with one LEU2 plasmid and one URA3 plasmid, both 2u in origin, are just fine, and I know other people have ... Maybe you run into problems with one plasmid curing the cell of the other if you have plasmids based on complete 2u sequence. ...
Aside from conjugative plasmids, the other main types of plasmids are. * Resistance plasmids: Carry genes for resistance to ... theyre called conjugative plasmids. Conjugative plasmids can facilitate the transfer of themselves, other plasmids, and even ... Some plasmids exist in many copies inside one cell and are called high-copy-number plasmids, whereas others are less numerous ... Although the enzymes involved in DNA replication of plasmids are the same as those used for the chromosome, some plasmids are ...
KorSA (plasmid) [Streptomyces ambofaciens] KorSA (plasmid) [Streptomyces ambofaciens]. gi,298043,emb,CAA79637.1, ...
Cloning Plasmid/DNA NEB # Features Concentration MW/Size Size Price pBR322 Vector N3033S/L Commonly used cloning vectors Tet ... Home Tools & Resources Selection Charts Cloning Plasmids and DNAs Cloning Plasmids and DNAs. Cloning Plasmid/DNA. NEB #. ... Kits are available for total RNA purification, plasmid miniprep, gel extraction, and DNA & RNA cleanup. For maximum convenience ... Kits are available for total RNA purification, plasmid miniprep, gel extraction, and DNA & RNA cleanup. For maximum convenience ...
... G.C.Tomlin mjfidgct at fs1.ce.umist.ac.uk Fri Jan 23 17:00:52 EST 1998 *Previous message: Postdoctoral ... Could someone please send me plasmids pRS413 and pRS414? Many thanks, Greg Tomlin ********************** Dept. of Biochemistry ...
Transformation efficiency declines linearly with increasing plasmid size. Relaxed and supercoiled plasmids transform with ... Studies on transformation of Escherichia coli with plasmids.. Hanahan D.. Abstract. Factors that affect the probability of ... A set of conditions is described under which about one in every 400 plasmid molecules produces a transformed cell. These ... Competition with both transforming and non-transforming plasmids indicates that each cell is capable of taking up many DNA ...
They conducted whole-genome analyses on the samples to study the bacterial plasmids, or rings of DNA, that can confer ... Understanding the plasmid exchange and when the plasmids get into the pathogens that infect our patients, she says, could help ... Antibiotic-resistant plasmids flourish in hospital plumbing. American Society for Microbiology. Journal. mBio. Keywords. * ... "How much should we care that there are a bunch of plasmids down in the wastewater system if theyre not infecting our patients ...
Biological resource centers are bigger and better than ever before, storing and distributing shared reagents, plasmids, and ...
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For ultrafast purification ... Principle ...,HiSpeed,Plasmid,Kits,biological,advanced biology technology,biology laboratory ... Basic Plasmid DNA Isolation Protocol. 9. RNA-free Plasmid DNA Isolation Protocol. 10. Basic Plasmid DNA Isolation Protocol. 11 ... Up to 200 g (Midi) or 750 g (Maxi) yield of high-copy plasmid DNA * Actual yields depend on plasmid copy number, size of insert ... New Reporter Plasmids for Studying Interferon-Stimulated Signal Transduction Pathways. 6. New Fusion Trans-Activator Plasmids ...
Plasmid publishes original research on genetic elements in all kingdoms of life with emphasis on maintenance, transmission and ... Plasmids promote the immigration of other plasmids but repress co-colonizing plasmids. ... Plasmids promote the immigration of other plasmids but repress co-colonizing plasmids. ... Recently published articles from Plasmid * Evolution in situ of ARI-A in pB2-1, a type 1 IncC plasmid recovered from Klebsiella ...
The particles are typically 65-85 nm, fully encapsulate the plasmid and are serum-stable. ... Plasmid-lipid particles which are useful for transfection of cells in vitro or in vivo are described. The particles can be ... No plasmid was recovered in the absence of DODAC while, at a DODAC:plasmid charge ratio of 2:1, 90% of the plasmid was ... 10, the free plasmid eluted in the void volume of the column while, in FIG. 11, the plasmid incubated in serum eluted in the ...
A new study led by scientists at the University of Oxford has found that small DNA molecules known as plasmids are one of the ... This occurs because bacteria usually carry more than one copy of a plasmid, which allows resistance genes carried by plasmids ... Our research shows a new role for plasmids in antibiotic resistance by demonstrating that plasmids drive the evolution of novel ... A new study led by scientists at the University of Oxford has found that small DNA molecules known as plasmids are one of the ...
  • Plasmids are extremely valuable tools in the fields of molecular biology and genetics, specifically in the area of genetic engineering ( q.v. ). They play a critical role in such procedures as gene cloning, recombinant protein production ( e.g., of human insulin), and gene therapy research. (britannica.com)
  • A foreign DNA element (such as the gene for insulin) is then spliced into the plasmid. (britannica.com)
  • In DNA cloning, a DNA fragment that contains a gene of interest is inserted into a cloning vector or plasmid . (accessexcellence.org)
  • The plasmid carrying genes for antibiotic resistance, and a DNA strand, which contains the gene of interest, are both cut with the same restriction endonuclease . (accessexcellence.org)
  • The plasmid is opened up and the gene is freed from its parent DNA strand. (accessexcellence.org)
  • The opened plasmid and the freed gene are mixed with DNA ligase, which reforms the two pieces as recombinant DNA. (accessexcellence.org)
  • I think Rose & Broach (Gene Expression Technology pp 234-279, ed Goeddel 1990) is a good starting point for the 2u plasmid stability literature. (bio.net)
  • Plasmid welcomes topics such as horizontal gene transfer, including antibiotic resistance transfer, and molecular aspects of microbial ecology. (elsevier.com)
  • The pGLO plasmid is a small circular piece of DNA that contains the gene to produce green fluorescent protein in the model organism. (reference.com)
  • Within the pGLO plasmid, the gene that codes for GFP is typically linked to a gene promoter of interest. (reference.com)
  • This plasmid carries the E. coli malE gene, encoding the maltose binding protein (MBP)(3), fused in-frame to the coding region of the Sce VMA intein-chitin binding domain (55 kDa)(1,4). (neb.com)
  • This term is no longer commonly used for plasmids, since it is now clear that a transposon (jumping gene or mobile genetic unit) makes a plasmid into an episome. (newworldencyclopedia.org)
  • We found that larger plasmids frequently have NAP gene homologs. (hindawi.com)
  • Plasmids with H-NS gene homologs had less G+C content. (hindawi.com)
  • It should be noted that plasmids with the NAP gene homolog also carried the relaxase gene involved in the conjugative transfer of plasmids more frequently than did those without the NAP gene homolog, implying that plasmid-encoded NAP homologs positively contribute to transmissible plasmids. (hindawi.com)
  • With respect to therapeutic compositions for gene therapy, the DNA is provided typically in the form of a plasmid for complexing with the cationic amphiphile. (google.com)
  • Novel and highly effective plasmid constructs are also disclosed, including those that are particularly effective at providing gene therapy for clinical conditions complicated by inflammation. (google.com)
  • The PathDetect Control Reporter Plasmid allows you to determine the direct or indirect involvement of new gene products, growth factors, and drug candidates in specific pathways. (agilent.com)
  • These plasmids carried conserved IncHI5 backbones composed of repHI5B and a repFIB -like gene (replication), parABC (partition), and tra1 (conjugal transfer). (frontiersin.org)
  • However, the design and construction of plasmid donors for each gene can be laborious and prone to troubleshooting. (genetics.org)
  • However, ssDNA donors are limited to small insertions, and, like plasmid donors, must be designed and generated for each gene that is targeted by HDR. (genetics.org)
  • The complete nucleotide sequence and gene organization of the three virulence plasmids from Yersinia pestis KIM5 were determined. (asm.org)
  • This US-enhanced expression of plasmid DNA will be discussed to emphasize the technical feasibility of US in gene therapy and biotechnology. (ingentaconnect.com)
  • Plasmids can be considered to be part of the mobilome , since they are often associated with conjugation , a mechanism of horizontal gene transfer . (thefullwiki.org)
  • Rather, plasmids provide a mechanism for horizontal gene transfer within a population of microbes and typically provide a selective advantage under a given environmental state. (thefullwiki.org)
  • The gene to be replicated is inserted into copies of a plasmid containing genes that make cells resistant to particular antibiotics and a multiple cloning site (MCS, or polylinker), which is a short region containing several commonly used restriction sites allowing the easy insertion of DNA fragments at this location. (thefullwiki.org)
  • The GeneArt™ gene synthesis platform can design plasmid vectors from modular, easily exchangeable fragments encoding regulatory and functional elements or from custom-made elements. (thermofisher.com)
  • 2 Gene Transfer with Plasmid Biopharmaceuticals. (wiley.com)
  • The sequence was a 93,629- bp plasmid encoding a single antimicrobial drug resistance gene, bla CTX-M-14. (cdc.gov)
  • Bacterial plasmids are key vectors of horizontal gene transfer, mediating the mobilization of genetic material from 1 bacterium to another. (cdc.gov)
  • In addition, plasmid replication and gene expression may interfere with bacterial growth in ways independent of any resistance functions carried by that plasmid. (genetics.org)
  • These results blur the current categorization of mobilizable and nonmobilizable plasmids and indicate that conjugative elements play a role in horizontal gene transfer even more significant than previously recognized. (mit.edu)
  • Tha AvaII-AvaII fragment that spanned the PstI site in the bla gene of this interim vector was replaced with the corresponding AvaII fragment from pUC8, eliminating this PstI site, making the PstI site in the MCS unique, to form plasmid pATH11. (stanford.edu)
  • This plasmid contains the gene for ampicillin resistance as well as the entire ß-galactosidase gene. (carolina.com)
  • Plasmid-mediated colistin resistance (mcr-1 gene): three months later, the story unfolds. (eurosurveillance.org)
  • Detection of the plasmid-mediated colistin-resistance gene mcr-1 in faecal metagenomes of Dutch travellers. (eurosurveillance.org)
  • Litrup E , Kiil K , Hammerum AM , Roer L , Nielsen EM , Torpdahl M . Plasmid-borne colistin resistance gene mcr-3 in Salmonella isolates from human infections, Denmark, 2009-17. (eurosurveillance.org)
  • Santa Cruz Biotechnology, Inc. offers a broad range of gene editing products including CRISPR/Cas9 Knockout and CRISPR Double Nickase plasmids for gene silencing. (scbt.com)
  • Myopodin gene silencers are available as Myopodin CRISPR/Cas9 Knockout plasmids and Myopodin Double Nickase Plasmids. (scbt.com)
  • Myopodin CRISPR/dCas9 Activation Plasmids and CRISPR Lenti Activation Systems for gene activation are also available. (scbt.com)
  • A major difference between the high-copy-number cloning vector type of plasmids used in the experiments mentioned above and many of the conjugative low-copy-number plasmids is, in addition to their potential for horizontal dissemination within and between bacterial species, the presence of stability systems in the latter. (genetics.org)
  • We have developed an eight-plasmid DNA transfection system for the rescue of infectious influenza A virus from cloned cDNA. (pnas.org)
  • In this plasmid-based expression system, viral cDNA is inserted between the RNA polymerase I (pol I) promoter and terminator sequences. (pnas.org)
  • The novel receptor is encoded for by cDNA carried on plasmid pGEM-hRARn, which has been desposited with the American Type Culture Collection for patent purposes. (reverso.net)
  • Plasmids responsible for "fertility" (or "chromosome mobilizing ability" [Cma]) (24) were also identified genetically in some other strains, including Streptomyces rimosus (18), Streptomyces lividans (29), Streptomyces erythreus (now called Saccharopolyspora erythrea ) (15), Streptomyces venezuelae (17), and Streptomyces ambofaciens (66). (springer.com)
  • In addition, since the plasmids that carry ESBL genes also commonly encode resistance determinants for many other antibiotics, ESBL strains are often resistant to many non-beta-lactam antibiotics as well, leaving very few options for the treatment. (wikipedia.org)
  • These plasmids are interesting because of their size, they range between 100 and 1800 Kb, and in some strains they represent about 50% of the bacterial genome. (elsevier.com)
  • Our system uses gRNA-based identification markers that allow messages to be addressed to specific strains via Cas9-mediated cleavage of messages sent to the wrong recipient, which we show reduces plasmid transfer by four orders of magnitude. (caltech.edu)
  • Malgorzata Adamczyk and Grazyna Jagura-Burdzy: "Spread and survival of promiscuous IncP-1 plasmids", Acta Biochimica Polonica, Vol 50, no. 2/2003, p. 425-453 LEWIS C. INGRAM, M. H. RICHMOND, AND R. B. SYKES: "Molecular Characterization of the R Factors Implicated in the Carbenicillin Resistance of a Sequence of Pseudomonas aeruginosa Strains Isolated from Burns", ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Feb. 1973, p. 279-288 Thomas CM et al. (wikipedia.org)
  • The enterobacterial strains produced a beta-lactamase with the same isoelectric point, immunological reactions, and substrate profile as those of the prototype PSE-1 enzyme determined by Pseudomonas plasmid RPL11. (asm.org)
  • Although some strains exhibited different antibiotic resistance patterns, some of their plasmids had similar migration patterns on agarose gel electrophoresis. (thefreelibrary.com)
  • Pesticinogenic and Ca2+-dependent strains of Yersinia pestis harbored plasmids of about 6 and 45 megadaltons, respectively. (asm.org)
  • We analyzed the differences in information capacity between prokaryotic chromosomes, genomic islands (GI), phages, and plasmids. (wur.nl)
  • Plasmid pPCP1 (9,610 bp) has a GC content of 45.3% and encodes two previously known virulence factors, an associated protein, and a single copy of IS 100 . (asm.org)
  • The present invention relates to Mycobacterium ulceran virulence plasmid , pMUM001 and particularly to a cluster of genes carried by this plasmid that encode polyketide synthases (PKSs) and polyketide-modifying enzymes necessary and sufficient for mycolactone biosynthesis. (reverso.net)
  • La présente invention se rapporte à un plasmide de virulence de Mycobacterium ulcerans, pMUM001, et notamment à une famille de gènes portés par ce plasmid et codant les polycétide-synthases (PKS) et les enzymes modificatrices de polycétides nécessaires et suffisantes pour la synthèse de la mycolactone. (reverso.net)
  • Recent studies demonstrated that the plasmid-driven expression of all eight vRNAs from a pol I promoter and the coexpression of the polymerase complex proteins result in the formation of infectious influenza A virus ( 16 , 17 ). (pnas.org)
  • Using this method in cultured S2R+ cells, we efficiently tagged four endogenous proteins with the bright fluorescent protein mNeonGreen, thereby demonstrating that an existing collection of CRISPaint universal donor plasmids is compatible with insect cells. (genetics.org)
  • Plasmids may carry genes that provide resistance to naturally occurring antibiotics in a competitive environmental niche, or alternatively the proteins produced may act as toxins under similar circumstances. (thefullwiki.org)
  • Another major use of plasmids is to make large amounts of proteins. (thefullwiki.org)
  • We also discuss the current understanding of control mechanisms affecting the plasmid copy number and how host chaperone proteins and proteases can affect plasmid maintenance in bacterial cells. (asmscience.org)
  • Plasmids were first clearly implicated in this conjugative process in the most studied strain, Streptomyces coelicolor A3(2), when certain derivatives of the wild-type isolate were found to differ in their "fertility" properties-that is, in the frequency with which they generated chromosomal recombinants when mated with various other derivatives-and this ability was inherited "infectiously" (2, 28, 72). (springer.com)
  • I used sucrose gradients to isolate my plasmid preps - got good enough DNA to produce good restriction patterns, clone specific fragments etc. (bio.net)
  • described an ESBL-producing isolate from calves with diarrhea that carried bla CTX-M-14 on an IncK plasmid, denoted pCT ( 15 , 16 ). (cdc.gov)
  • Q9: Can the ZymoPURE kits be used to isolate large plasmid constructs (BAC/PAC)? (zymoresearch.com)
  • Functional vRNP molecules could be generated either by infection with helper virus or by cotransfection of protein expression plasmids encoding PB1, PB2, PA, or nucleoproteins ( 11 , 13 - 15 ). (pnas.org)
  • In addition to 5 previously described genes, such as murine toxin, capsule antigen, capsule anchoring protein, etc., 30 homologues to genes of several bacterial species were found in this plasmid, and another 44 open reading frames without homology to any known or hypothetical protein in the databases were predicted. (asm.org)
  • Plasmids that can be mobilized by conjugative elements are generally thought to contain an origin of transfer (oriT), from which mobilization initiates, and to encode a mobilization protein (Mob, a relaxase) that nicks a site in oriT and covalently attaches to the DNA to be transferred. (mit.edu)
  • taxA coded for a protein of 181 amino acids that showed similarity to TraY of F-like plasmids and to the Arc-repressor superfamily. (unboundmedicine.com)
  • The wash regimen has been optimized to ensure the plasmid DNA is free of endotoxins, salt, and protein. (zymoresearch.com)
  • Different types of plasmids have been reported and it is possible for plasmids of different varieties to coexist in a single cell. (newworldencyclopedia.org)
  • c) removing said detergent from said solution of step (b) to provide a solution of serum-stable plasmid-lipid particles, wherein said plasmid is encapsulated in a lipid bilayer and said particles are resistant to degradation in serum, and wherein the particles have a diameter ranging from about 50 to about 150 nm. (google.com)
  • 11. A method in accordance with claim 10, wherein said plasmid-lipid particle comprises a plasmid, DODAC, POPC and a PEG-Ceramide selected from the group consisting of PEG-Cer-C 20 and PEG-Cer-C 14 . (google.com)
  • 4. A composition according to claim 3 wherein said plasmid further facilitates interaction of said polymerase with itself. (google.com)
  • Understanding the plasmid exchange and when the plasmids get into the pathogens that infect our patients, she says, could help hospitals improve their monitoring of resistance-conferring genes: "In the big picture, the concern is the spread of these resistant organisms worldwide and some regions of the world are not tracking the spread of the hospital isolates. (eurekalert.org)
  • 1997) on isolates from fish revealed a similar size range of R plasmids (3 to 63. (thefreedictionary.com)
  • The plasmid spread to unrelated E. coli isolates within an index cattle farm and persisted within the environment. (cdc.gov)
  • In this study, we report the full sequence and analysis of pCT and demonstrate the spread of pCT-like plasmids in animal and human E. coli isolates from the United Kingdom, Europe, Australia, and Asia. (cdc.gov)
  • In addition, most isolates examined possessed a cryptic 65-megadalton plasmid. (asm.org)
  • A set of conditions is described under which about one in every 400 plasmid molecules produces a transformed cell. (nih.gov)
  • Competition with both transforming and non-transforming plasmids indicates that each cell is capable of taking up many DNA molecules, and that the establishment of a transformation event is neither helped nor hindered significantly by the presence of multiple plasmids. (nih.gov)
  • A new study led by scientists at the University of Oxford has found that small DNA molecules known as plasmids are one of the key culprits in spreading the major global health threat of antibiotic resistance. (news-medical.net)
  • Currently, however, the term plasmid is restricted only to those accessory DNA molecules that are found in addition to the main chromosomes. (newworldencyclopedia.org)
  • David Wessner reviews Vector NTI Suite, an INternet integrated software tool that can catalog, analyze, and design complex plasmids for molecular biologists. (sciencemag.org)
  • The term plasmid was first introduced by the American molecular biologist Joshua Lederberg in 1952 to describe any extrachromosomal hereditary determinant. (newworldencyclopedia.org)
  • The presence of these non-antimicrobial-drug resistance R plasmid DNA sequences in SGI1 constitutes a molecular signature that firmly establishes the aquaculture origin of the florfenicol resistance and the tetR/ tetG genes in the S. (thefreedictionary.com)
  • The term plasmid was first introduced by the American molecular biologist Joshua Lederberg in 1952. (thefullwiki.org)
  • Plasmids of IncH2 and IncFIme were shown to contain 8 x 10(6)-molecular-weight transposons Tn1401 and Tn1402 that encoded PSE-1 beta-lactamase production, resistance to streptomycin and spectinomycin via AAD(3"), and resistance to sulfonamide. (asm.org)
  • Emergence of plasmid-mediated colistin resistance mechanism MCR-1 in animals and human beings in China: a microbiological and molecular biological study. (eurosurveillance.org)
  • Recent studies have shown that some plasmids carry genes encoding NAP homologs, which play important roles in transcriptional regulation networks between plasmids and host chromosomes. (hindawi.com)
  • Plasmids are important as they can replicate and express genes despite being outside the organism's chromosomes. (sciencephoto.com)
  • There was an association between relative entropy and AT content in chromosomes, phages, plasmids and GIs with the strongest association being in phages. (wur.nl)
  • Conclusions: We argue that relative entropy differences reflect how plasmids, phages and GIs interact with microbial host chromosomes and that all these biological entities are, or have been, subjected to different selective pressures. (wur.nl)
  • A plasmid is an extra chromosomal DNA molecule separate from the chromosomal DNA which is capable of replicating independently from the chromosomal DNA. (thefullwiki.org)
  • We provide evidence that HR plasmids may encode machinery for their horizontal transfer. (apsnet.org)
  • Maybe you run into problems with one plasmid curing the cell of the other if you have plasmids based on complete 2u sequence. (bio.net)
  • Addgene will store your samples in triplicate (including one at an offsite backup facility), sequence key regions of your plasmid for validation, and handle the appropriate Material Transfer Agreements (MTAs) with your institution. (addgene.org)
  • 600 bases of sequence from pUC-TMV using GenElute Plasmid Miniprep Kit. (sigmaaldrich.com)
  • In addition, GeneOptimizer® sequence optimization further refines plasmid design, helping to accommodate your design features and achieve maximum expression in your host. (thermofisher.com)
  • The BglII-HindIII fragment (nt 1392 of trpE to the end of the trpD sequence present in pKRS101) was replaced with a BamHI-EcoRI fragment and an EcoRI-HindIII fragment, both from the MCS of M13mp12 to make plasmid pATH1 (see GenBank M32985). (stanford.edu)
  • In such procedures, a plasmid is cut at a specific site (or sites) using enzymes called restriction endonucleases. (britannica.com)
  • Plasmid DNA is ready for immediate use in restriction enzyme digestion (Figure 1) cloning, PCR, transcription, conventional and automated sequencing (Figure 2) . (sigmaaldrich.com)
  • Plasmid DNA purified with GenElute Plasmid Miniprep Kit is suitable for restriction enzyme digestions. (sigmaaldrich.com)
  • Restriction digestions of pUC19 purified from JM109 using the GenElute Plasmid Miniprep Kit. (sigmaaldrich.com)
  • GenElute™ Plasmid Miniprep Kit offers a simple, rapid, and cost-effective method for isolating up to 20 µg plasmid DNA from E. coli cultures. (sigmaaldrich.com)
  • As an added convenience, the ZymoPURE™ Plasmid Miniprep Kit contains colored buffers that permit error-free visualization and identification of complete bacterial cell lysis and neutralization. (zymoresearch.com)
  • Comprehensive and highly practical, E. coli Plasmid Vectors offers those new to the field a basic guide to the use of plasmid vectors in the cloning host E. coli, and those more experienced researchers a broad-ranging, proven array of successful techniques. (springer.com)
  • pMYB5 is a control plasmid for the IMPACT™ Kit (NEB #E6901) (1,2). (neb.com)
  • Using complex cloning strategies when needed, we can help you develop completely customized vectors, and you retain any intellectual property associated with the resulting expression plasmid. (thermofisher.com)
  • Additionally, plasmids automatically amplify the number of copies of these new and improved resistance genes. (news-medical.net)
  • Depending on the size of the plasmids, the number of copies of the same plasmid in a single cell varies from one to several hundreds. (newworldencyclopedia.org)
  • In E. coli, multiple plasmid copies appear to cluster together, creating a few multiplasmid clusters in each cell. (wikipedia.org)
  • Plasmids serve as important tools in genetics and biotechnology labs, where they are commonly used to multiply (make many copies of) or express particular genes. (thefullwiki.org)
  • When introduced into a host organism via transformation, a plasmid will be replicated, creating numerous copies of the DNA fragment under study. (jove.com)
  • Activity 2 will help students conceptualize the mechanics involved in cutting and ligating DNAs into a plasmid vector with "sticky ends" of complementary DNA base pairs. (accessexcellence.org)
  • The new findings add to a growing body of evidence suggesting that the conduits of hospital wastewater serve as a vast and resilient reservoir for plasmids that can confer the genes responsible for antibiotic resistance. (eurekalert.org)
  • ZymoPURE™ technology uses a modified alkaline lysis method and features novel binding chemistry that yields highly concentrated plasmid DNA (up to 3 µg/µl). (zymoresearch.com)
  • Plus, earn free Addgene plasmids . (addgene.org)
  • There is no cost to deposit your plasmids to Addgene. (addgene.org)
  • Additionally, Addgene will create a webpage for your lab where you can direct scientists to request your plasmids. (addgene.org)
  • The Huda Zoghbi Lab has deposited plasmids at Addgene for distribution to the research community. (addgene.org)
  • Transformation efficiency declines linearly with increasing plasmid size. (nih.gov)
  • AFAIK would be better to redesign the plasmid, unless the GFP is on the same plasmid, it would be useless for checking transformation efficiency if it has much higher transformation efficiency than your target. (protocol-online.org)
  • [ 1 ] Microbial transformation with plasmid DNA is neither parasitic nor symbiotic in nature, since each implies the presence of an independent species living in a commensal or detrimental state with the host organism. (thefullwiki.org)
  • Transformation can also be carried out by introducing a plasmid into a bacterial cell. (ukessays.com)