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.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.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.Base Sequence: The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.DNA, Complementary: Single-stranded complementary DNA synthesized from an RNA template by the action of RNA-dependent DNA polymerase. cDNA (i.e., complementary DNA, not circular DNA, not C-DNA) is used in a variety of molecular cloning experiments as well as serving as a specific hybridization probe.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.Cloning, Organism: The formation of one or more genetically identical organisms derived by vegetative reproduction from a single cell. The source nuclear material can be embryo-derived, fetus-derived, or taken from an adult somatic cell.Sequence Alignment: The arrangement of two or more amino acid or base sequences from an organism or organisms in such a way as to align areas of the sequences sharing common properties. The degree of relatedness or homology between the sequences is predicted computationally or statistically based on weights assigned to the elements aligned between the sequences. This in turn can serve as a potential indicator of the genetic relatedness between the organisms.Restriction Mapping: Use of restriction endonucleases to analyze and generate a physical map of genomes, genes, or other segments of DNA.Genes, Bacterial: The functional hereditary units 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.Gene Library: A large collection of DNA fragments cloned (CLONING, MOLECULAR) from a given organism, tissue, organ, or cell type. It may contain complete genomic sequences (GENOMIC LIBRARY) or complementary DNA sequences, the latter being formed from messenger RNA and lacking intron sequences.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.Sequence Analysis, DNA: A multistage process that includes cloning, physical mapping, subcloning, determination of the DNA SEQUENCE, and information analysis.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).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.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.Recombinant Proteins: Proteins prepared by recombinant DNA technology.Blotting, Northern: Detection of RNA 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.Chromosome Mapping: Any method used for determining the location of and relative distances between genes on a chromosome.Gene Expression: The phenotypic manifestation of a gene or genes by the processes of GENETIC TRANSCRIPTION and GENETIC TRANSLATION.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.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.DNA, Bacterial: Deoxyribonucleic acid that makes up the genetic material of bacteria.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.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.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).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.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)Phylogeny: The relationships of groups of organisms as reflected by their genetic makeup.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.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.Tissue Distribution: Accumulation of a drug or chemical substance in various organs (including those not relevant to its pharmacologic or therapeutic action). This distribution depends on the blood flow or perfusion rate of the organ, the ability of the drug to penetrate organ membranes, tissue specificity, protein binding. The distribution is usually expressed as tissue to plasma ratios.Genomic Library: A form of GENE LIBRARY containing the complete DNA sequences present in the genome of a given organism. It contrasts with a cDNA library which contains only sequences utilized in protein coding (lacking introns).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.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.Multigene Family: A set of genes descended by duplication and variation from some ancestral gene. Such genes may be clustered together on the same chromosome or dispersed on different chromosomes. Examples of multigene families include those that encode the hemoglobins, immunoglobulins, histocompatibility antigens, actins, tubulins, keratins, collagens, heat shock proteins, salivary glue proteins, chorion proteins, cuticle proteins, yolk proteins, and phaseolins, as well as histones, ribosomal RNA, and transfer RNA genes. The latter three are examples of reiterated genes, where hundreds of identical genes are present in a tandem array. (King & Stanfield, A Dictionary of Genetics, 4th ed)Molecular Weight: The sum of the weight of all the atoms in a molecule.Bacterial Proteins: Proteins found in any species of bacterium.Cell Line: Established cell cultures that have the potential to propagate indefinitely.Organ Specificity: Characteristic restricted to a particular organ of the body, such as a cell type, metabolic response or expression of a particular protein or antigen.Sequence Analysis: A multistage process that includes the determination of a sequence (protein, carbohydrate, etc.), its fragmentation and analysis, and the interpretation of the resulting sequence information.Cosmids: Plasmids containing at least one cos (cohesive-end site) of PHAGE LAMBDA. They are used as cloning vehicles.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.Substrate Specificity: A characteristic feature of enzyme activity in relation to the kind of substrate on which the enzyme or catalytic molecule reacts.Carrier Proteins: Transport proteins that carry specific substances in the blood or across cell membranes.Oligonucleotide Probes: Synthetic or natural oligonucleotides used in hybridization studies in order to identify and study specific nucleic acid fragments, e.g., DNA segments near or within a specific gene locus or gene. The probe hybridizes with a specific mRNA, if present. Conventional techniques used for testing for the hybridization product include dot blot assays, Southern blot assays, and DNA:RNA hybrid-specific antibody tests. Conventional labels for the probe include the radioisotope labels 32P and 125I and the chemical label biotin.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.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.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.Genes, Fungal: The functional hereditary units of FUNGI.Sequence Homology: The degree of similarity between sequences. Studies of AMINO ACID SEQUENCE HOMOLOGY and NUCLEIC ACID SEQUENCE HOMOLOGY provide useful information about the genetic relatedness of genes, gene products, and species.In Situ Hybridization: A technique that localizes specific nucleic acid sequences within intact chromosomes, eukaryotic cells, or bacterial cells through the use of specific nucleic acid-labeled probes.Electrophoresis, Polyacrylamide Gel: Electrophoresis in which a polyacrylamide gel is used as the diffusion medium.Plant Proteins: Proteins found in plants (flowers, herbs, shrubs, trees, etc.). The concept does not include proteins found in vegetables for which VEGETABLE PROTEINS is available.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.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: A polynucleotide consisting essentially of chains with a repeating backbone of phosphate and ribose units to which nitrogenous bases are attached. RNA is unique among biological macromolecules in that it can encode genetic information, serve as an abundant structural component of cells, and also possesses catalytic activity. (Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed)Conserved Sequence: A sequence of amino acids in a polypeptide or of nucleotides in DNA or RNA that is similar across multiple species. A known set of conserved sequences is represented by a CONSENSUS SEQUENCE. AMINO ACID MOTIFS are often composed of conserved sequences.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.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.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.Kinetics: The rate dynamics in chemical or physical systems.Genes, Plant: The functional hereditary units of PLANTS.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.Exons: The parts of a transcript of a split GENE remaining after the INTRONS are removed. They are spliced together to become a MESSENGER RNA or other functional RNA.Membrane Proteins: Proteins which are found in membranes including cellular and intracellular membranes. They consist of two types, peripheral and integral proteins. They include most membrane-associated enzymes, antigenic proteins, transport proteins, and drug, hormone, and lectin receptors.Protein Biosynthesis: The biosynthesis of PEPTIDES and PROTEINS on RIBOSOMES, directed by MESSENGER RNA, via TRANSFER RNA that is charged with standard proteinogenic AMINO ACIDS.Alternative Splicing: A process whereby multiple RNA transcripts are generated from a single gene. Alternative splicing involves the splicing together of other possible sets of EXONS during the processing of some, but not all, transcripts of the gene. Thus a particular exon may be connected to any one of several alternative exons to form a mature RNA. The alternative forms of mature MESSENGER RNA produce PROTEIN ISOFORMS in which one part of the isoforms is common while the other parts are different.Reverse Transcriptase Polymerase Chain Reaction: A variation of the PCR technique in which cDNA is made from RNA via reverse transcription. The resultant cDNA is then amplified using standard PCR protocols.Gene Expression Regulation, Enzymologic: Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control of gene action in enzyme synthesis.Introns: Sequences of DNA in the genes that are located between the EXONS. They are transcribed along with the exons but are removed from the primary gene transcript by RNA SPLICING to leave mature RNA. Some introns code for separate genes.Cattle: Domesticated bovine animals of the genus Bos, usually kept on a farm or ranch and used for the production of meat or dairy products or for heavy labor.DNA, Fungal: Deoxyribonucleic acid that makes up the genetic material of fungi.DNA Probes: Species- or subspecies-specific DNA (including COMPLEMENTARY DNA; conserved genes, whole chromosomes, or whole genomes) used in hybridization studies in order to identify microorganisms, to measure DNA-DNA homologies, to group subspecies, etc. The DNA probe hybridizes with a specific mRNA, if present. Conventional techniques used for testing for the hybridization product include dot blot assays, Southern blot assays, and DNA:RNA hybrid-specific antibody tests. Conventional labels for the DNA probe include the radioisotope labels 32P and 125I and the chemical label biotin. The use of DNA probes provides a specific, sensitive, rapid, and inexpensive replacement for cell culture techniques for diagnosing infections.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.Oligodeoxyribonucleotides: A group of deoxyribonucleotides (up to 12) in which the phosphate residues of each deoxyribonucleotide act as bridges in forming diester linkages between the deoxyribose moieties.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.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).COS Cells: CELL LINES derived from the CV-1 cell line by transformation with a replication origin defective mutant of SV40 VIRUS, which codes for wild type large T antigen (ANTIGENS, POLYOMAVIRUS TRANSFORMING). They are used for transfection and cloning. (The CV-1 cell line was derived from the kidney of an adult male African green monkey (CERCOPITHECUS AETHIOPS).)Cricetinae: A subfamily in the family MURIDAE, comprising the hamsters. Four of the more common genera are Cricetus, CRICETULUS; MESOCRICETUS; and PHODOPUS.Isoenzymes: Structurally related forms of an enzyme. Each isoenzyme has the same mechanism and classification, but differs in its chemical, physical, or immunological characteristics.Transcription Factors: Endogenous substances, usually proteins, which are effective in the initiation, stimulation, or termination of the genetic transcription process.Poly A: A group of adenine ribonucleotides in which the phosphate residues of each adenine ribonucleotide act as bridges in forming diester linkages between the ribose moieties.Nuclear Transfer Techniques: Methods of implanting a CELL NUCLEUS from a donor cell into an enucleated acceptor cell.Phenotype: The outward appearance of the individual. It is the product of interactions between genes, and between the GENOTYPE and the environment.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.DNA, Plant: Deoxyribonucleic acid that makes up the genetic material of plants.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.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.Consensus Sequence: A theoretical representative nucleotide or amino acid sequence in which each nucleotide or amino acid is the one which occurs most frequently at that site in the different sequences which occur in nature. The phrase also refers to an actual sequence which approximates the theoretical consensus. A known CONSERVED SEQUENCE set is represented by a consensus sequence. Commonly observed supersecondary protein structures (AMINO ACID MOTIFS) are often formed by conserved sequences.Physical Chromosome Mapping: Mapping of the linear order of genes on a chromosome with units indicating their distances by using methods other than genetic recombination. These methods include nucleotide sequencing, overlapping deletions in polytene chromosomes, and electron micrography of heteroduplex DNA. (From King & Stansfield, A Dictionary of Genetics, 5th ed)Gene Expression Regulation, Developmental: Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control of gene action during the developmental stages of an organism.Gene Expression Regulation, Bacterial: Any of the processes by which cytoplasmic or intercellular factors influence the differential control of gene action in bacteria.Liver: A large lobed glandular organ in the abdomen of vertebrates that is responsible for detoxification, metabolism, synthesis and storage of various substances.Bacillus: A genus of BACILLACEAE that are spore-forming, rod-shaped cells. Most species are saprophytic soil forms with only a few species being pathogenic.Chromosomes, Bacterial: Structures within the nucleus of bacterial cells consisting of or containing DNA, which carry genetic information essential to the cell.Enzyme Stability: The extent to which an enzyme retains its structural conformation or its activity when subjected to storage, isolation, and purification or various other physical or chemical manipulations, including proteolytic enzymes and heat.Proteins: Linear POLYPEPTIDES that are synthesized on RIBOSOMES and may be further modified, crosslinked, cleaved, or assembled into complex proteins with several subunits. The specific sequence of AMINO ACIDS determines the shape the polypeptide will take, during PROTEIN FOLDING, and the function of the protein.Xenopus laevis: The commonest and widest ranging species of the clawed "frog" (Xenopus) in Africa. This species is used extensively in research. There is now a significant population in California derived from escaped laboratory animals.Rabbits: The species Oryctolagus cuniculus, in the family Leporidae, order LAGOMORPHA. Rabbits are born in burrows, furless, and with eyes and ears closed. In contrast with HARES, rabbits have 22 chromosome pairs.Blotting, Western: Identification of proteins or peptides that have been electrophoretically separated by blot transferring from the electrophoresis gel to strips of nitrocellulose paper, followed by labeling with antibody probes.Binding Sites: The parts of a macromolecule that directly participate in its specific combination with another molecule.Oocytes: Female germ cells derived from OOGONIA and termed OOCYTES when they enter MEIOSIS. The primary oocytes begin meiosis but are arrested at the diplotene state until OVULATION at PUBERTY to give rise to haploid secondary oocytes or ova (OVUM).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.Chromosome Walking: A technique with which an unknown region of a chromosome can be explored. It is generally used to isolate a locus of interest for which no probe is available but that is known to be linked to a gene which has been identified and cloned. A fragment containing a known gene is selected and used as a probe to identify other overlapping fragments which contain the same gene. The nucleotide sequences of these fragments can then be characterized. This process continues for the length of the chromosome.Codon: A set of three nucleotides in a protein coding sequence that specifies individual amino acids or a termination signal (CODON, TERMINATOR). Most codons are universal, but some organisms do not produce the transfer RNAs (RNA, TRANSFER) complementary to all codons. These codons are referred to as unassigned codons (CODONS, NONSENSE).Chromosomes, Artificial, Bacterial: DNA constructs that are composed of, at least, a REPLICATION ORIGIN, for successful replication, propagation to and maintenance as an extra chromosome in bacteria. In addition, they can carry large amounts (about 200 kilobases) of other sequence for a variety of bioengineering purposes.Chickens: Common name for the species Gallus gallus, the domestic fowl, in the family Phasianidae, order GALLIFORMES. It is descended from the red jungle fowl of SOUTHEAST ASIA.Clone Cells: A group of genetically identical cells all descended from a single common ancestral cell by mitosis in eukaryotes or by binary fission in prokaryotes. Clone cells also include populations of recombinant DNA molecules all carrying the same inserted sequence. (From King & Stansfield, Dictionary of Genetics, 4th ed)Pseudomonas: A genus of gram-negative, aerobic, rod-shaped bacteria widely distributed in nature. Some species are pathogenic for humans, animals, and plants.Gene Expression Regulation, Plant: Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control of gene action in plants.Bacillus subtilis: A species of gram-positive bacteria that is a common soil and water saprophyte.Fungal Proteins: Proteins found in any species of fungus.Tumor Cells, Cultured: Cells grown in vitro from neoplastic tissue. If they can be established as a TUMOR CELL LINE, they can be propagated in cell culture indefinitely.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.-.Protein Sorting Signals: Amino acid sequences found in transported proteins that selectively guide the distribution of the proteins to specific cellular compartments.Oxidoreductases: The class of all enzymes catalyzing oxidoreduction reactions. The substrate that is oxidized is regarded as a hydrogen donor. The systematic name is based on donor:acceptor oxidoreductase. The recommended name will be dehydrogenase, wherever this is possible; as an alternative, reductase can be used. Oxidase is only used in cases where O2 is the acceptor. (Enzyme Nomenclature, 1992, p9)Brain: The part of CENTRAL NERVOUS SYSTEM that is contained within the skull (CRANIUM). Arising from the NEURAL TUBE, the embryonic brain is comprised of three major parts including PROSENCEPHALON (the forebrain); MESENCEPHALON (the midbrain); and RHOMBENCEPHALON (the hindbrain). The developed brain consists of CEREBRUM; CEREBELLUM; and other structures in the BRAIN STEM.Xenopus: An aquatic genus of the family, Pipidae, occurring in Africa and distinguished by having black horny claws on three inner hind toes.Insect Proteins: Proteins found in any species of insect.Genes, Viral: The functional hereditary units of VIRUSES.CHO Cells: CELL LINE derived from the ovary of the Chinese hamster, Cricetulus griseus (CRICETULUS). The species is a favorite for cytogenetic studies because of its small chromosome number. The cell line has provided model systems for the study of genetic alterations in cultured mammalian cells.Arabidopsis: A plant genus of the family BRASSICACEAE that contains ARABIDOPSIS PROTEINS and MADS DOMAIN PROTEINS. The species A. thaliana is used for experiments in classical plant genetics as well as molecular genetic studies in plant physiology, biochemistry, and development.Hydrogen-Ion Concentration: The normality of a solution with respect to HYDROGEN ions; H+. It is related to acidity measurements in most cases by pH = log 1/2[1/(H+)], where (H+) is the hydrogen ion concentration in gram equivalents per liter of solution. (McGraw-Hill Dictionary of Scientific and Technical Terms, 6th ed)Protein Conformation: The characteristic 3-dimensional shape of a protein, including the secondary, supersecondary (motifs), tertiary (domains) and quaternary structure of the peptide chain. PROTEIN STRUCTURE, QUATERNARY describes the conformation assumed by multimeric proteins (aggregates of more than one polypeptide chain).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.-.Cells, Cultured: Cells propagated in vitro in special media conducive to their growth. Cultured cells are used to study developmental, morphologic, metabolic, physiologic, and genetic processes, among others.Chromosomes, Artificial, Yeast: Chromosomes in which fragments of exogenous DNA ranging in length up to several hundred kilobase pairs have been cloned into yeast through ligation to vector sequences. These artificial chromosomes are used extensively in molecular biology for the construction of comprehensive genomic libraries of higher organisms.DNA, Viral: Deoxyribonucleic acid that makes up the genetic material of viruses.Pichia: Yeast-like ascomycetous fungi of the family Saccharomycetaceae, order SACCHAROMYCETALES isolated from exuded tree sap.Plants: Multicellular, eukaryotic life forms of kingdom Plantae (sensu lato), comprising the VIRIDIPLANTAE; RHODOPHYTA; and GLAUCOPHYTA; all of which acquired chloroplasts by direct endosymbiosis of CYANOBACTERIA. They are characterized by a mainly photosynthetic mode of nutrition; essentially unlimited growth at localized regions of cell divisions (MERISTEMS); cellulose within cells providing rigidity; the absence of organs of locomotion; absence of nervous and sensory systems; and an alternation of haploid and diploid generations.Swine: Any of various animals that constitute the family Suidae and comprise stout-bodied, short-legged omnivorous mammals with thick skin, usually covered with coarse bristles, a rather long mobile snout, and small tail. Included are the genera Babyrousa, Phacochoerus (wart hogs), and Sus, the latter containing the domestic pig (see SUS SCROFA).Genetic Markers: A phenotypically recognizable genetic trait which can be used to identify a genetic locus, a linkage group, or a recombination event.Testis: The male gonad containing two functional parts: the SEMINIFEROUS TUBULES for the production and transport of male germ cells (SPERMATOGENESIS) and the interstitial compartment containing LEYDIG CELLS that produce ANDROGENS.Temperature: The property of objects that determines the direction of heat flow when they are placed in direct thermal contact. The temperature is the energy of microscopic motions (vibrational and translational) of the particles of atoms.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.Alcohol Oxidoreductases: A subclass of enzymes which includes all dehydrogenases acting on primary and secondary alcohols as well as hemiacetals. They are further classified according to the acceptor which can be NAD+ or NADP+ (subclass 1.1.1), cytochrome (1.1.2), oxygen (1.1.3), quinone (1.1.5), or another acceptor (1.1.99).Kidney: Body organ that filters blood for the secretion of URINE and that regulates ion concentrations.Oryza sativa: Annual cereal grass of the family POACEAE and its edible starchy grain, rice, which is the staple food of roughly one-half of the world's population.Protein Binding: The process in which substances, either endogenous or exogenous, bind to proteins, peptides, enzymes, protein precursors, or allied compounds. Specific protein-binding measures are often used as assays in diagnostic assessments.Protein Structure, Tertiary: The level of protein structure in which combinations of secondary protein structures (alpha helices, beta sheets, loop regions, and motifs) pack together to form folded shapes called domains. Disulfide bridges between cysteines in two different parts of the polypeptide chain along with other interactions between the chains play a role in the formation and stabilization of tertiary structure. Small proteins usually consist of only one domain but larger proteins may contain a number of domains connected by segments of polypeptide chain which lack regular secondary structure.Mutagenesis: Process of generating a genetic MUTATION. It may occur spontaneously or be induced by MUTAGENS.Expressed Sequence Tags: Partial cDNA (DNA, COMPLEMENTARY) sequences that are unique to the cDNAs from which they were derived.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.Membrane Glycoproteins: Glycoproteins found on the membrane or surface of cells.Alleles: Variant forms of the same gene, occupying the same locus on homologous CHROMOSOMES, and governing the variants in production of the same gene product.Macromolecular Substances: Compounds and molecular complexes that consist of very large numbers of atoms and are generally over 500 kDa in size. In biological systems macromolecular substances usually can be visualized using ELECTRON MICROSCOPY and are distinguished from ORGANELLES by the lack of a membrane structure.Hybrid Cells: Any cell, other than a ZYGOTE, that contains elements (such as NUCLEI and CYTOPLASM) from two or more different cells, usually produced by artificial CELL FUSION.

*  Molecular Cloning |authorSTREAM

An introductory lecture on molecular cloning as part of a genetics class. Covers amplification of genes in E. Coli and PCR.- ... Does anyone know how molecular cloning works? DNA Cloning : 2 methods : DNA Cloning : 2 methods Molecular cloning using a live ... Molecular cloning with live cell intermediate: Summary : Molecular cloning with live cell intermediate: Summary First you need ... Molecular cloning with live cell intermediate: Summary : Molecular cloning with live cell intermediate: Summary First you need ...
authorstream.com/Presentation/Robin80H-101553-molecular-cloning-pcr-genetics-science-technology-ppt-powerpoint/

*  Starting cells for making one's own competent cells - Molecular Cloning - BioForum

... posted in Molecular Cloning: When making chemically or electrically-competent cells, is it usual to start with non-competent ...
protocol-online.org/forums/topic/18969-starting-cells-for-making-ones-own-competent-cells/

*  Help! Plasmid disappearing after mini-prep. - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. Help! Plasmid disappearing after mini-prep. - A260 shows good quality ...
protocol-online.org/biology-forums-2/posts/10729.html

*  plasmid, BAC, or cosmid? - Molecular Cloning - BioForum

... i'm looking into cloning of a big chunk of genomic sequence of about 25kb to put into a vector and transfect it into some cell ... is the cloning strategy for such large insert the same as typical cloning strategies? I frequently clone insert up to around ... Hi guys, i'm looking into cloning of a big chunk of genomic sequence of about 25kb to put into a vector and transfect it into ... Some of these contain a high copy origin intended to be removed as part of the cloning process. You might also consider the ...
protocol-online.org/forums/topic/10293-plasmid-bac-or-cosmid/

*  Inserts too small after transformation - Molecular Cloning

Molecular Cloning. Inserts too small after transformation - transformation (Mar/13/2009 ). Hi all!. I have cloned a 5kb insert ... I cannot clone a shorter fragment due to the fact that we want to have a full genome viral clone.. I have never heard of ... Using cleaned up PCR framents for cloning I always get a lot more white cfu's than blue ones using my own protocol! ... I tried the same cloning and transformation strategy in another vector: pUC18 but no luck either. Both vectors used are high ...
protocol-online.org/biology-forums-2/posts/6876.html

*  Cloning advice - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. Cloning advice - (May/21/2013 ). I clone a 915bp genes into PCDNA3. ... sometimes you lose things during cloning. typically there is something unanticipated like a recombination event or poor primer ... do you have other colonies you picked from the cloning to sequence? ...
protocol-online.org/biology-forums-2/posts/29332.html

*  12.4kb vector 6.7kb insert - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. 12.4kb vector 6.7kb insert - (May/12/2012 ). Hi all, I am having ... the cloning will not work without one of the ori's becoming inoperable (i.e. mutant). ...
protocol-online.org/biology-forums-2/posts/25515.html

*  Stuck from last august! need help urgently - cloning and pcr ! - Molecular Cloning - BioForum

... posted in Molecular Cloning: Hi I have been trying to clone a pcr product of 3065bp in pEF6V5 His TOPO vector. I have tried ... Is it okay if i use this topo vector for restriction based cloning instead of TA cloning.. i am worried if topoisomerase wud ... But there are many other approaches -- you are not forced to use TA cloning. TOPO blunt kits, for example, would directly clone ... to your primers to introduce restriction sites and clone using conventional restriction enzyme cloning. ...
protocol-online.org/forums/topic/28389-stuck-from-last-august-need-help-urgently-cloning-and-pcr/

*  control DNA transformation- using pGEM kit - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. control DNA transformation- using pGEM kit - transformation (Feb/02/2011 ... This was another of those times! I've just read in our Molecular Cloning book (Sambrook and Russel), page 1.125, "Colonies that ... I am very new to this forum and cloning. I am working on a school project on cloning. I am only seeing white colonies on the ... I am very new to this forum and cloning. I am working on a school project on cloning. I am only seeing white colonies on the ...
protocol-online.org/biology-forums-2/posts/19360.html

*  Expansion of commercial competent bacterial cell lines? - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. Expansion of commercial competent bacterial cell lines? - (Nov/14/2013 ) ...
protocol-online.org/biology-forums-2/posts/30999.html

*  Competent DH5 alpha - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. Competent DH5 alpha - (Mar/16/2009 ). Hi, I'm searching a good protocol ...
protocol-online.org/biology-forums-2/posts/6924.html

*  tryptone soy broth - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. tryptone soy broth - (Mar/25/2009 ). we are running out of LB and the ... we use DH5 alpha as cloning strain ...
protocol-online.org/biology-forums-2/posts/7143.html

*  Mutagenesis PCR - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. Mutagenesis PCR - (Aug/09/2014 ). Hi, I have two questions about ... I'd be showing you the door because molecular cloning work like this is about as easy as it gets in comparison to some of the ... If I am ever fortunate enough to have a lab of my own and my student refused to do some kind of cloning because it was "too ... And if you don't have primers that flank the ends of your insert where your cloning restriction sites are, get two of those. ...
protocol-online.org/biology-forums-2/posts/32911.html

*  Help with cloning!. - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. Help with cloning!. - Cloning problems (May/10/2009 ). Pages: 1 2 3 Next ... I have failed each time with the cloning, due to a number of reasons, since how ever much I try it seems that I can t get a ... I have failed each time with the cloning, due to a number of reasons, since how ever much I try it seems that I can t get a ... For the past three months I have been trying to clone a 2KB fragment ( cut out from a 7kb plasmid) to a 12kb back bone. ...
protocol-online.org/biology-forums-2/posts/7984.html

*  pCS2 - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. pCS2 - (Mar/07/2009 ). Hi,. What kind of vector is pCS2-flag vector. Can ...
protocol-online.org/biology-forums-2/posts/6768.html

*  Molecular Cloning - BioForum - Page 70

Any techniques related to molecular cloning including restriction digestion, ligation, transformation, plasmid... ... Molecular Cloning. Any techniques related to molecular cloning including restriction digestion, ligation, transformation, ... InsTA Clone t/a cloning blues( actually whites) Started by rabos, 25 Jul 2009 * 2 replies ... Cloning a whole gene in pBAD (any comments pls) Started by arvinsign, 09 Aug 2009 * 4 replies ...
protocol-online.org/forums/forum/42-molecular-cloning/page-70?prune_day=100&sort_by=Z-A&sort_key=last_post&topicfilter=all

*  A Modified MultiSite Gateway Cloning Strategy for Consolidation of Genes in Plants | SpringerLink

Molecular Biotechnology. February 2013. , Volume 53, Issue 2, pp 129-138 , Cite as ... 2000). GATEWAY recombinational cloning: application to the cloning of large numbers of open reading frames or ORFeomes. Methods ... In this study, we modified gateway MultiSite cloning vectors and developed a hybrid cloning strategy which combines advantages ... Gene cloning vectors that facilitate the fusion, overexpression or down regulation of genes in plant cells are being used with ...
https://link.springer.com/article/10.1007/s12033-012-9499-6

*  Molecular Cloning and Primary Structure of the Chicken Transforming Growth Factor-β2 Gene - UQ eSpace

Molecular Cloning and Primary Structure of the Chicken Transforming Growth Factor-β2 Gene. Burt, DW and Paton, IR (1991) ... Molecular Cloning and Primary Structure of the Chicken Transforming Growth Factor-β2 Gene ... Molecular Cloning and Primary Structure of the Chicken Transforming Growth Factor-β2 Gene. Dna and Cell Biology, 10 10: 723-734 ...
https://espace.library.uq.edu.au/view/UQ:683677

*  Self-Ligation of digested vector - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. Self-Ligation of digested vector - (Jun/20/2009 ). Hello/Nimen Hao. I ... What I want to do now is to self-ligate the vector to clone a gene using different enzyme sites. What should I do, I know! but ...
protocol-online.org/biology-forums-2/posts/8717.html

*  competent cells stock - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. competent cells stock - (Jul/11/2011 ). we're run out out of our ...
protocol-online.org/biology-forums-2/posts/22052.html

*  Subcloning into pMIR-REPORT-luc - Molecular Cloning - BioForum

I'm trying to use Ambion's pMIR-REPORT-luc reporter plasmid to clone in a 40bp insert in the 3' UTR of the luciferase gene. The ... posted in Molecular Cloning: Hey everyone, I'm about to pull my hair out over this and am hoping to find some help here. ... i've been trying to use the very same vector and my problem is that even if I'm getting cellswith the clone, all of the have ... I'm trying to use Ambion's pMIR-REPORT-luc reporter plasmid to clone in a 40bp insert in the 3' UTR of the luciferase gene. The ...
protocol-online.org/forums/topic/10406-subcloning-into-pmir-report-luc/

*  detection of restriction? - Molecular Cloning

Top : New Forum Archives (2009-): : Molecular Cloning. detection of restriction? - (Sep/26/2009 ). i have a 387bp template. its ...
protocol-online.org/biology-forums-2/posts/10509.html

*  Molecular-Cloning

Sequencing of the cloned plasmid - (reply: 5) 341. 4 way cloning into TA Vector - (reply: 10) 342. I get band double the size ... Neeed Help , cloning Confirmation ?! - (reply: 5) 345. Puzzling Cloning Problem With a Specific Vector - (reply: 1) 346. Can I ... Very difficult cloning project - how to clone something that is not compatible w - (reply: 4) ... TA Cloning of ds cDNA - (reply: 6) 357. Promoter distance from ATG - (reply: 3) 358. Measuring Fluorescene of RFP - (reply: 7) ...
protocol-online.org/biology-forums-2/Molecular-Cloning/more11.html

*  HNRPM Antibody (monoclonal) (M01) - Mouse monoclonal antibody raised against a partial recombinant HNRPM. WB, IF, E - Buy Now!...

Researchers worldwide depend on Abgent FLcDNA clones to accelerate their research. Expression-ready clones are convenient ... With state-of-the art molecular biology and protein biochemistry labs, we work with our clients to rapidly evaluate in parallel ... PMID 18790094.Cloning and characterization of a novel deletion mutant of heterogeneous nuclear ribonucleoprotein M4 from human ... HNRPM monoclonal antibody (M01), clone 2B6. Western Blot analysis of HNRPM expression in HepG2 ( (Cat # AT2398a ) ...
abgent.com/products/AT2398a-HNRPM-Antibody-monoclonal-M01

*  Anti-Rat Nestin Antibody, clone Rat-401 (4D4) - Mouse Anti-Rat Monoclonal Antibody WB, IHC-P, IHC-F, IF - Buy Now! |Abgent

... clone Rat-401 (4D4) , Mouse Anti-Rat Monoclonal Antibody validated in WB, IHC-P, IHC-F, IF (ABD10658), Abgent ... With state-of-the art molecular biology and protein biochemistry labs, we work with our clients to rapidly evaluate in parallel ... Researchers worldwide depend on Abgent FLcDNA clones to accelerate their research. Expression-ready clones are convenient ... Anti-Rat Nestin Antibody, clone Rat-401 (4D4) is for research use only and not for use in diagnostic or therapeutic procedures. ...
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Ligation-independent cloning: Ligation-independent cloning (LIC) is a form of molecular cloning that is able to be performed without the use of restriction endonucleases or DNA ligase. This allows genes that have restriction sites to be cloned without worry of chopping up the insert.Coles PhillipsProtein primary structure: The primary structure of a peptide or protein is the linear sequence of its amino acid structural units, and partly comprises its overall biomolecular structure. By convention, the primary structure of a protein is reported starting from the amino-terminal (N) end to the carboxyl-terminal (C) end.Symmetry element: A symmetry element is a point of reference about which symmetry operations can take place. In particular, symmetry elements can be centers of inversion, axes of rotation and mirror planes.CS-BLASTList of strains of Escherichia coli: Escherichia coli is a well studied bacterium that was first identified by Theodor Escherich, after whom it was later named.Library (biology): In molecular biology, a library is a collection of DNA fragments that is stored and propagated in a population of micro-organisms through the process of molecular cloning. There are different types of DNA libraries, including cDNA libraries (formed from reverse-transcribed RNA), genomic libraries (formed from genomic DNA) and randomized mutant libraries (formed by de novo gene synthesis where alternative nucleotides or codons are incorporated).DNA sequencer: A DNA sequencer is a scientific instrument used to automate the DNA sequencing process. Given a sample of DNA, a DNA sequencer is used to determine the order of the four bases: G (guanine), C (cytosine), A (adenine) and T (thymine).DNA condensation: DNA condensation refers to the process of compacting DNA molecules in vitro or in vivo. Mechanistic details of DNA packing are essential for its functioning in the process of gene regulation in living systems.Mature messenger RNA: Mature messenger RNA, often abbreviated as mature mRNA is a eukaryotic RNA transcript that has been spliced and processed and is ready for translation in the course of protein synthesis. Unlike the eukaryotic RNA immediately after transcription known as precursor messenger RNA, it consists exclusively of exons, with all introns removed.Triparental mating: Triparental mating is a form of Bacterial conjugation where a conjugative plasmid present in one bacterial strain assists the transfer of a mobilizable plasmid present in a second bacterial strain into a third bacterial strain. Plasmids are introduced into bacteria for such purposes as transformation, cloning, or transposon mutagenesis.Chromosome regionsThermal cyclerOpen reading frame: In molecular genetics, an open reading frame (ORF) is the part of a reading frame that has the potential to code for a protein or peptide. An ORF is a continuous stretch of codons that do not contain a stop codon (usually UAA, UAG or UGA).Restriction fragment: A restriction fragment is a DNA fragment resulting from the cutting of a DNA strand by a restriction enzyme (restriction endonucleases), a process called restriction. Each restriction enzyme is highly specific, recognising a particular short DNA sequence, or restriction site, and cutting both DNA strands at specific points within this site.Branching order of bacterial phyla (Gupta, 2001): There are several models of the Branching order of bacterial phyla, one of these was proposed in 2001 by Gupta based on conserved indels or protein, termed "protein signatures", an alternative approach to molecular phylogeny. Some problematic exceptions and conflicts are present to these conserved indels, however, they are in agreement with several groupings of classes and phyla.Multiple cloning site: A multiple cloning site (MCS), also called a polylinker, is a short segment of DNA which contains many (up to ~20) restriction sites - a standard feature of engineered plasmids. Restriction sites within an MCS are typically unique, occurring only once within a given plasmid.Eukaryotic transcription: Eukaryotic transcription is the elaborate process that eukaryotic cells use to copy genetic information stored in DNA into units of RNA replica. Gene transcription occurs in both eukaryotic and prokaryotic cells.ParaHox: The ParaHox gene cluster is an array of homeobox genes (involved in morphogenesis, the regulation of patterns of anatomical development) from the Gsx, Xlox (Pdx) and Cdx gene families.Molar mass distribution: In linear polymers the individual polymer chains rarely have exactly the same degree of polymerization and molar mass, and there is always a distribution around an average value. The molar mass distribution (or molecular weight distribution) in a polymer describes the relationship between the number of moles of each polymer species (Ni) and the molar mass (Mi) of that species.Ferric uptake regulator family: In molecular biology, the ferric uptake regulator (FUR) family of proteins includes metal ion uptake regulator proteins. These are responsible for controlling the intracellular concentration of iron in many bacteria.Silent mutation: Silent mutations are mutations in DNA that do not significantly alter the phenotype of the organism in which they occur. Silent mutations can occur in non-coding regions (outside of genes or within introns), or they may occur within exons.Specificity constant: In the field of biochemistry, the specificity constant (also called kinetic efficiency or k_{cat}/K_{M}), is a measure of how efficiently an enzyme converts substrates into products. A comparison of specificity constants can also be used as a measure of the preference of an enzyme for different substrates (i.Allele-specific oligonucleotide: An allele-specific oligonucleotide (ASO) is a short piece of synthetic DNA complementary to the sequence of a variable target DNA. It acts as a probe for the presence of the target in a Southern blot assay or, more commonly, in the simpler Dot blot assay.RNA transfection: RNA transfection is the process of deliberately introducing RNA into a living cell. RNA can be purified from cells after lysis or synthesized from free nucleotides either chemically, or enzymatically using an RNA polymerase to transcribe a DNA template.Avery–MacLeod–McCarty experimentMargaret Jope: Margaret Jope (1913–2004) was a Scottish biochemist, born as Henrietta Margaret Halliday in Peterhead, Scotland.YjdF RNA motifStreptomyces peucetius: Streptomyces peucetius ATCC 27952GC box: In molecular biology, a GC box is a distinct pattern of nucleotides found in the promoter region of some eukaryotic genes upstream of the TATA box and approximately 110 bases upstream from the transcription initiation site. It has a consensus sequence GGGCGG which is position dependent and orientation independent.Burst kinetics: Burst kinetics is a form of enzyme kinetics that refers to an initial high velocity of enzymatic turnover when adding enzyme to substrate. This initial period of high velocity product formation is referred to as the "Burst Phase".Zuotin: Z-DNA binding protein 1, also known as Zuotin, is a Saccharomyces cerevisiae yeast gene.Alternative splicing: Alternative splicing is a regulated process during gene expression that results in a single gene coding for multiple proteins. In this process, particular exons of a gene may be included within or excluded from the final, processed messenger RNA (mRNA) produced from that gene.Membrane protein: Membrane proteins are proteins that interact with biological membranes. They are one of the common types of protein along with soluble globular proteins, fibrous proteins, and disordered proteins.Translational regulation: Translational regulation refers to the control of the levels of protein synthesized from its mRNA. The corresponding mechanisms are primarily targeted on the control of ribosome recruitment on the initiation codon, but can also involve modulation of the elongation or termination of protein synthesis.Intron: right|thumbnail|270px|Representation of intron and [[exons within a simple gene containing a single intron.]]Beef cattle: Beef cattle are cattle raised for meat production (as distinguished from dairy cattle, used for milk production). The meat of adult cattle is known as beef.Composite transposon: A composite transposon is similar in function to simple transposons and Insertion Sequence (IS) elements in that it has protein coding DNA segments flanked by inverted, repeated sequences that can be recognized by transposase enzymes. A composite transposon, however, is flanked by two separate IS elements which may or may not be exact replicas.CpG OligodeoxynucleotideOperon: In genetics, an operon is a functioning unit of genomic DNA containing a cluster of genes under the control of a single promoter. The genes are transcribed together into an mRNA strand and either translated together in the cytoplasm, or undergo trans-splicing to create monocistronic mRNAs that are translated separately, i.Direct repeat: Direct repeats are a type of genetic sequence that consists of two or more repeats of a specific sequence.Baby hamster kidney cell: Baby Hamster Kidney fibroblasts (aka BHK cells) are an adherent cell line used in molecular biology.Isozyme: Isozymes (also known as isoenzymes or more generally as Multiple forms of enzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes usually display different kinetic parameters (e.Pituitary-specific positive transcription factor 1: POU domain, class 1, transcription factor 1 (Pit1, growth hormone factor 1), also known as POU1F1, is a transcription factor for growth hormone.Extension Poly(A) Test: The extension Poly(A) Test (ePAT) describes a method to determine the poly(A) tail lengths of mRNA molecules. It was developed and described by A.Phenotype microarray: The phenotype microarray approach is a technology for high-throughput phenotyping of cells.Signature-tagged mutagenesis: Signature-tagged mutagenesis (STM) is a genetic technique used to study gene function. Recent advances in genome sequencing have allowed us to catalogue a large variety of organisms' genomes, but the function of the genes they contain is still largely unknown.DNA-binding proteinAntitermination: Antitermination is the prokaryotic cell's aid to fix premature termination of RNA synthesis during the transcription of RNA. It occurs when the RNA polymerase ignores the termination signal, and it provides a mechanism whereby one or more genes at the end of an operon can be switched either on or off, depending on the polymerase either recognizing or not recognizing the termination signal.Liver sinusoid: A liver sinusoid is a type of sinusoidal blood vessel (with fenestrated, discontinuous endothelium) that serves as a location for the oxygen-rich blood from the hepatic artery and the nutrient-rich blood from the portal vein.SIU SOM Histology GIBacillus alcalophilus: Bacillus alcalophilus is a Gram-positive, rod-shaped species of bacteria. Likely strains of this species have been isolated from highly alkaline waste water.Circular bacterial chromosome: A circular bacterial chromosome is a bacterial chromosome in the form of a molecule of circular DNA. Unlike the linear DNA of most eukaryotes, typical bacterial chromosomes are circular.Lattice protein: Lattice proteins are highly simplified computer models of proteins which are used to investigate protein folding.

(1/69323) Novel regulation of the homeotic gene Scr associated with a crustacean leg-to-maxilliped appendage transformation.

Homeotic genes are known to be involved in patterning morphological structures along the antero-posterior axis of insects and vertebrates. Because of their important roles in development, changes in the function and expression patterns of homeotic genes may have played a major role in the evolution of different body plans. For example, it has been proposed that during the evolution of several crustacean lineages, changes in the expression patterns of the homeotic genes Ultrabithorax and abdominal-A have played a role in transformation of the anterior thoracic appendages into mouthparts termed maxillipeds. This homeotic-like transformation is recapitulated at the late stages of the direct embryonic development of the crustacean Porcellio scaber (Oniscidea, Isopoda). Interestingly, this morphological change is associated with apparent novelties both in the transcriptional and post-transcriptional regulation of the Porcellio scaber ortholog of the Drosophila homeotic gene, Sex combs reduced (Scr). Specifically, we find that Scr mRNA is present in the second maxillary segment and the first pair of thoracic legs (T1) in early embryos, whereas protein accumulates only in the second maxillae. In later stages, however, high levels of SCR appear in the T1 legs, which correlates temporally with the transformation of these appendages into maxillipeds. Our observations provide further insight into the process of the homeotic leg-to-maxilliped transformation in the evolution of crustaceans and suggest a novel regulatory mechanism for this process in this group of arthropods.  (+info)

(2/69323) Mrj encodes a DnaJ-related co-chaperone that is essential for murine placental development.

We have identified a novel gene in a gene trap screen that encodes a protein related to the DnaJ co-chaperone in E. coli. The gene, named Mrj (mammalian relative of DnaJ) was expressed throughout development in both the embryo and placenta. Within the placenta, expression was particularly high in trophoblast giant cells but moderate levels were also observed in trophoblast cells of the chorion at embryonic day 8.5, and later in the labyrinth which arises from the attachment of the chorion to the allantois (a process called chorioallantoic fusion). Insertion of the ROSAbetageo gene trap vector into the Mrj gene created a null allele. Homozygous Mrj mutants died at mid-gestation due to a failure of chorioallantoic fusion at embryonic day 8.5, which precluded formation of the mature placenta. At embryonic day 8.5, the chorion in mutants was morphologically normal and expressed the cell adhesion molecule beta4 integrin that is known to be required for chorioallantoic fusion. However, expression of the chorionic trophoblast-specific transcription factor genes Err2 and Gcm1 was significantly reduced. The mutants showed no abnormal phenotypes in other trophoblast cell types or in the embryo proper. This study indicates a previously unsuspected role for chaperone proteins in placental development and represents the first genetic analysis of DnaJ-related protein function in higher eukaryotes. Based on a survey of EST databases representing different mouse tissues and embryonic stages, there are 40 or more DnaJ-related genes in mammals. In addition to Mrj, at least two of these genes are also expressed in the developing mouse placenta. The specificity of the developmental defect in Mrj mutants suggests that each of these genes may have unique tissue and cellular activities.  (+info)

(3/69323) Requirement of a novel gene, Xin, in cardiac morphogenesis.

A novel gene, Xin, from chick (cXin) and mouse (mXin) embryonic hearts, may be required for cardiac morphogenesis and looping. Both cloned cDNAs have a single open reading frame, encoding proteins with 2,562 and 1,677 amino acids for cXin and mXin, respectively. The derived amino acid sequences share 46% similarity. The overall domain structures of the predicted cXin and mXin proteins, including proline-rich regions, 16 amino acid repeats, DNA-binding domains, SH3-binding motifs and nuclear localization signals, are highly conserved. Northern blot analyses detect a single message of 8.9 and 5.8 kilo base (kb) from both cardiac and skeletal muscle of chick and mouse, respectively. In situ hybridization reveals that the cXin gene is specifically expressed in cardiac progenitor cells of chick embryos as early as stage 8, prior to heart tube formation. cXin continues to be expressed in the myocardium of developing hearts. By stage 15, cXin expression is also detected in the myotomes of developing somites. Immunofluorescence microscopy reveals that the mXin protein is colocalized with N-cadherin and connexin-43 in the intercalated discs of adult mouse hearts. Incubation of stage 6 chick embryos with cXin antisense oligonucleotides results in abnormal cardiac morphogenesis and an alteration of cardiac looping. The myocardium of the affected hearts becomes thickened and tends to form multiple invaginations into the heart cavity. This abnormal cellular process may account in part for the abnormal looping. cXin expression can be induced by bone morphogenetic protein (BMP) in explants of anterior medial mesoendoderm from stage 6 chick embryos, a tissue that is normally non-cardiogenic. This induction occurs following the BMP-mediated induction of two cardiac-restricted transcription factors, Nkx2.5 and MEF2C. Furthermore, either MEF2C or Nkx2.5 can transactivate a luciferase reporter driven by the mXin promoter in mouse fibroblasts. These results suggest that Xin may participate in a BMP-Nkx2.5-MEF2C pathway to control cardiac morphogenesis and looping.  (+info)

(4/69323) Mechanisms of GDF-5 action during skeletal development.

Mutations in GDF-5, a member of the TGF-beta superfamily, result in the autosomal recessive syndromes brachypod (bp) in mice and Hunter-Thompson and Grebe-type chondrodysplasias in humans. These syndromes are all characterised by the shortening of the appendicular skeleton and loss or abnormal development of some joints. To investigate how GDF-5 controls skeletogenesis, we overexpressed GDF-5 during chick limb development using the retrovirus, RCASBP. This resulted in up to a 37.5% increase in length of the skeletal elements, which was predominantly due to an increase in the number of chondrocytes. By injecting virus at different stages of development, we show that GDF-5 can increase both the size of the early cartilage condensation and the later developing skeletal element. Using in vitro micromass cultures as a model system to study the early steps of chondrogenesis, we show that GDF-5 increases chondrogenesis in a dose-dependent manner. We did not detect changes in proliferation. However, cell suspension cultures showed that GDF-5 might act at these stages by increasing cell adhesion, a critical determinant of early chondrogenesis. In contrast, pulse labelling experiments of GDF-5-infected limbs showed that at later stages of skeletal development GDF-5 can increase proliferation of chondrocytes. Thus, here we show two mechanisms of how GDF-5 may control different stages of skeletogenesis. Finally, our data show that levels of GDF-5 expression/activity are important in controlling the size of skeletal elements and provides a possible explanation for the variation in the severity of skeletal defects resulting from mutations in GDF-5.  (+info)

(5/69323) Molecular cloning and epitope analysis of the peanut allergen Ara h 3.

Peanut allergy is a significant IgE-mediated health problem because of the increased prevalence, potential severity, and chronicity of the reaction. Following our characterization of the two peanut allergens Ara h 1 and Ara h 2, we have isolated a cDNA clone encoding a third peanut allergen, Ara h 3. The deduced amino acid sequence of Ara h 3 shows homology to 11S seed-storage proteins. The recombinant form of this protein was expressed in a bacterial system and was recognized by serum IgE from approximately 45% of our peanut-allergic patient population. Serum IgE from these patients and overlapping, synthetic peptides were used to map the linear, IgE-binding epitopes of Ara h 3. Four epitopes, between 10 and 15 amino acids in length, were found within the primary sequence, with no obvious sequence motif shared by the peptides. One epitope is recognized by all Ara h 3-allergic patients. Mutational analysis of the epitopes revealed that single amino acid changes within these peptides could lead to a reduction or loss of IgE binding. By determining which amino acids are critical for IgE binding, it might be possible to alter the Ara h 3 cDNA to encode a protein with a reduced IgE-binding capacity. These results will enable the design of improved diagnostic and therapeutic approaches for food-hypersensitivity reactions.  (+info)

(6/69323) TIF1gamma, a novel member of the transcriptional intermediary factor 1 family.

We report the cloning and characterization of a novel member of the Transcriptional Intermediary Factor 1 (TIF1) gene family, human TIF1gamma. Similar to TIF1alpha and TIF1beta, the structure of TIF1beta is characterized by multiple domains: RING finger, B boxes, Coiled coil, PHD/TTC, and bromodomain. Although structurally related to TIF1alpha and TIF1beta, TIF1gamma presents several functional differences. In contrast to TIF1alpha, but like TIF1beta, TIF1 does not interact with nuclear receptors in yeast two-hybrid or GST pull-down assays and does not interfere with retinoic acid response in transfected mammalian cells. Whereas TIF1alpha and TIF1beta were previously found to interact with the KRAB silencing domain of KOX1 and with the HP1alpha, MODI (HP1beta) and MOD2 (HP1gamma) heterochromatinic proteins, suggesting that they may participate in a complex involved in heterochromatin-induced gene repression, TIF1gamma does not interact with either the KRAB domain of KOX1 or the HP1 proteins. Nevertheless, TIF1gamma, like TIF1alpha and TIF1beta, exhibits a strong silencing activity when tethered to a promoter. Since deletion of a novel motif unique to the three TIF1 proteins, called TIF1 signature sequence (TSS), abrogates transcriptional repression by TIF1gamma, this motif likely participates in TIF1 dependent repression.  (+info)

(7/69323) Anopheles gambiae Ag-STAT, a new insect member of the STAT family, is activated in response to bacterial infection.

A new insect member of the STAT family of transcription factors (Ag-STAT) has been cloned from the human malaria vector Anopheles gambiae. The domain involved in DNA interaction and the SH2 domain are well conserved. Ag-STAT is most similar to Drosophila D-STAT and to vertebrate STATs 5 and 6, constituting a proposed ancient class A of the STAT family. The mRNA is expressed at all developmental stages, and the protein is present in hemocytes, pericardial cells, midgut, skeletal muscle and fat body cells. There is no evidence of transcriptional activation following bacterial challenge. However, bacterial challenge results in nuclear translocation of Ag-STAT protein in fat body cells and induction of DNA-binding activity that recognizes a STAT target site. In vitro treatment with pervanadate (vanadate and H2O2) translocates Ag-STAT to the nucleus in midgut epithelial cells. This is the first evidence of direct participation of the STAT pathway in immune responses in insects.  (+info)

(8/69323) In vivo expression of the nucleolar group I intron-encoded I-dirI homing endonuclease involves the removal of a spliceosomal intron.

The Didymium iridis DiSSU1 intron is located in the nuclear SSU rDNA and has an unusual twin-ribozyme organization. One of the ribozymes (DiGIR2) catalyses intron excision and exon ligation. The other ribozyme (DiGIR1), which along with the endonuclease-encoding I-DirI open reading frame (ORF) is inserted in DiGIR2, carries out hydrolysis at internal processing sites (IPS1 and IPS2) located at its 3' end. Examination of the in vivo expression of DiSSU1 shows that after excision, DiSSU1 is matured further into the I-DirI mRNA by internal DiGIR1-catalysed cleavage upstream of the ORF 5' end, as well as truncation and polyadenylation downstream of the ORF 3' end. A spliceosomal intron, the first to be reported within a group I intron and the rDNA, is removed before the I-DirI mRNA associates with the polysomes. Taken together, our results imply that DiSSU1 uses a unique combination of intron-supplied ribozyme activity and adaptation to the general RNA polymerase II pathway of mRNA expression to allow a protein to be produced from the RNA polymerase I-transcribed rDNA.  (+info)



Somatic


  • Somatic cell nuclear transfer (SCNT) is much more widely and efficiently practiced in cattle than in any other species, making this arguably the most important mammal cloned to date. (springer.com)
  • We emphasize the impact of epigenetic and genetic differences between embryonic, germ, and somatic donor cell types on cloning efficiency. (springer.com)
  • Somatic cell cloning without micromanipulators. (springer.com)
  • Somatic cell nuclear transfer ("therapeutic cloning"), which is one way to derive stem cells, shows potential in generating functional replacement cells such as insulin-producing cells associated with diabetes. (innovations-report.com)

Stem Cells


  • Nuclear cloning, stem cells, and genomic reprogramming. (springer.com)

adult


  • Lastly, we discuss adult phenotypes and fitness of cloned cattle and their offspring and illustrate some of the more imminent commercial cattle cloning applications. (springer.com)
  • Bioengineered tissues were created from heart, skeletal muscle and kidney cells cloned from adult cow skin cells. (innovations-report.com)

Biological


  • In this paper, we review current cattle cloning methodologies and their potential technical or biological pitfalls at any step of the procedure. (springer.com)