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.Cell Line: Established cell cultures that have the potential to propagate indefinitely.Base Sequence: The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.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.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.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.Polyethyleneimine: Strongly cationic polymer that binds to certain proteins; used as a marker in immunology, to precipitate and purify enzymes and lipids. Synonyms: aziridine polymer; Epamine; Epomine; ethylenimine polymer; Montrek; PEI; Polymin(e).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).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.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.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.Luciferases: Enzymes that oxidize certain LUMINESCENT AGENTS to emit light (PHYSICAL LUMINESCENCE). The luciferases from different organisms have evolved differently so have different structures and substrates.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.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.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.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.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.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.RNA, Small Interfering: Small double-stranded, non-protein coding RNAs (21-31 nucleotides) involved in GENE SILENCING functions, especially RNA INTERFERENCE (RNAi). Endogenously, siRNAs are generated from dsRNAs (RNA, DOUBLE-STRANDED) by the same ribonuclease, Dicer, that generates miRNAs (MICRORNAS). The perfect match of the siRNAs' antisense strand to their target RNAs mediates RNAi by siRNA-guided RNA cleavage. siRNAs fall into different classes including trans-acting siRNA (tasiRNA), repeat-associated RNA (rasiRNA), small-scan RNA (scnRNA), and Piwi protein-interacting RNA (piRNA) and have different specific gene silencing functions.Cell Line, Tumor: A cell line derived from cultured tumor cells.Genetic Therapy: Techniques and strategies which include the use of coding sequences and other conventional or radical means to transform or modify cells for the purpose of treating or reversing disease conditions.Liposomes: Artificial, single or multilaminar vesicles (made from lecithins or other lipids) that are used for the delivery of a variety of biological molecules or molecular complexes to cells, for example, drug delivery and gene transfer. They are also used to study membranes and membrane proteins.Gene Expression: The phenotypic manifestation of a gene or genes by the processes of GENETIC TRANSCRIPTION and GENETIC TRANSLATION.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.Transcription Factors: Endogenous substances, usually proteins, which are effective in the initiation, stimulation, or termination of the genetic transcription process.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.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.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.HeLa Cells: The first continuously cultured human malignant CELL LINE, derived from the cervical carcinoma of Henrietta Lacks. These cells are used for VIRUS CULTIVATION and antitumor drug screening assays.Green Fluorescent Proteins: Protein analogs and derivatives of the Aequorea victoria green fluorescent protein that emit light (FLUORESCENCE) when excited with ULTRAVIOLET RAYS. They are used in REPORTER GENES in doing GENETIC TECHNIQUES. Numerous mutants have been made to emit other colors or be sensitive to pH.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.Cations: Positively charged atoms, radicals or groups of atoms which travel to the cathode or negative pole during electrolysis.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).)Transcriptional Activation: Processes that stimulate the GENETIC TRANSCRIPTION of a gene or set of genes.Cricetinae: A subfamily in the family MURIDAE, comprising the hamsters. Four of the more common genera are Cricetus, CRICETULUS; MESOCRICETUS; and PHODOPUS.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.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.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.Recombinant Proteins: Proteins prepared by recombinant DNA technology.Signal Transduction: The intracellular transfer of information (biological activation/inhibition) through a signal pathway. In each signal transduction system, an activation/inhibition signal from a biologically active molecule (hormone, neurotransmitter) is mediated via the coupling of a receptor/enzyme to a second messenger system or to an ion channel. Signal transduction plays an important role in activating cellular functions, cell differentiation, and cell proliferation. Examples of signal transduction systems are the GAMMA-AMINOBUTYRIC ACID-postsynaptic receptor-calcium ion channel system, the receptor-mediated T-cell activation pathway, and the receptor-mediated activation of phospholipases. Those coupled to membrane depolarization or intracellular release of calcium include the receptor-mediated activation of cytotoxic functions in granulocytes and the synaptic potentiation of protein kinase activation. Some signal transduction pathways may be part of larger signal transduction pathways; for example, protein kinase activation is part of the platelet activation signal pathway.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.Cell Survival: The span of viability of a cell characterized by the capacity to perform certain functions such as metabolism, growth, reproduction, some form of responsiveness, and adaptability.Apoptosis: One of the mechanisms by which CELL DEATH occurs (compare with NECROSIS and AUTOPHAGOCYTOSIS). Apoptosis is the mechanism responsible for the physiological deletion of cells and appears to be intrinsically programmed. It is characterized by distinctive morphologic changes in the nucleus and cytoplasm, chromatin cleavage at regularly spaced sites, and the endonucleolytic cleavage of genomic DNA; (DNA FRAGMENTATION); at internucleosomal sites. This mode of cell death serves as a balance to mitosis in regulating the size of animal tissues and in mediating pathologic processes associated with tumor growth.Polylysine: A peptide which is a homopolymer of lysine.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.Cation Exchange Resins: High molecular weight insoluble polymers which contain functional anionic groups that are capable of undergoing exchange reactions with cations.Binding Sites: The parts of a macromolecule that directly participate in its specific combination with another molecule.Quaternary Ammonium Compounds: Derivatives of ammonium compounds, NH4+ Y-, in which all four of the hydrogens bonded to nitrogen have been replaced with hydrocarbyl groups. These are distinguished from IMINES which are RN=CR2.Cercopithecus aethiops: A species of CERCOPITHECUS containing three subspecies: C. tantalus, C. pygerythrus, and C. sabeus. They are found in the forests and savannah of Africa. The African green monkey (C. pygerythrus) is the natural host of SIMIAN IMMUNODEFICIENCY VIRUS and is used in AIDS research.Down-Regulation: A negative regulatory effect on physiological processes at the molecular, cellular, or systemic level. At the molecular level, the major regulatory sites include membrane receptors, genes (GENE EXPRESSION REGULATION), mRNAs (RNA, MESSENGER), and proteins.Nuclear Proteins: Proteins found in the nucleus of a cell. Do not confuse with NUCLEOPROTEINS which are proteins conjugated with nucleic acids, that are not necessarily present in the nucleus.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.DNA, Antisense: DNA that is complementary to the sense strand. (The sense strand has the same sequence as the mRNA transcript. The antisense strand is the template for mRNA synthesis.) Synthetic antisense DNAs are used to hybridize to complementary sequences in target RNAs or DNAs to effect the functioning of specific genes for investigative or therapeutic purposes.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.3T3 Cells: Cell lines whose original growing procedure consisted being transferred (T) every 3 days and plated at 300,000 cells per plate (J Cell Biol 17:299-313, 1963). Lines have been developed using several different strains of mice. Tissues are usually fibroblasts derived from mouse embryos but other types and sources have been developed as well. The 3T3 lines are valuable in vitro host systems for oncogenic virus transformation studies, since 3T3 cells possess a high sensitivity to CONTACT INHIBITION.DNA, Viral: Deoxyribonucleic acid that makes up the genetic material of viruses.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.Fibroblasts: Connective tissue cells which secrete an extracellular matrix rich in collagen and other macromolecules.Trans-Activators: Diffusible gene products that act on homologous or heterologous molecules of viral or cellular DNA to regulate the expression of proteins.Cell Division: The fission of a CELL. It includes CYTOKINESIS, when the CYTOPLASM of a cell is divided, and CELL NUCLEUS DIVISION.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.Cell Nucleus: Within a eukaryotic cell, a membrane-limited body which contains chromosomes and one or more nucleoli (CELL NUCLEOLUS). The nuclear membrane consists of a double unit-type membrane which is perforated by a number of pores; the outermost membrane is continuous with the ENDOPLASMIC RETICULUM. A cell may contain more than one nucleus. (From Singleton & Sainsbury, Dictionary of Microbiology and Molecular Biology, 2d ed)Phosphatidylethanolamines: Derivatives of phosphatidic acids in which the phosphoric acid is bound in ester linkage to an ethanolamine moiety. Complete hydrolysis yields 1 mole of glycerol, phosphoric acid and ethanolamine and 2 moles of fatty acids.Cell Line, Transformed: Eukaryotic cell line obtained in a quiescent or stationary phase which undergoes conversion to a state of unregulated growth in culture, resembling an in vitro tumor. It occurs spontaneously or through interaction with viruses, oncogenes, radiation, or drugs/chemicals.Gene Expression Regulation, Neoplastic: Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control of gene action in neoplastic tissue.Sp1 Transcription Factor: Promoter-specific RNA polymerase II transcription factor that binds to the GC box, one of the upstream promoter elements, in mammalian cells. The binding of Sp1 is necessary for the initiation of transcription in the promoters of a variety of cellular and viral GENES.RNA Interference: A gene silencing phenomenon whereby specific dsRNAs (RNA, DOUBLE-STRANDED) trigger the degradation of homologous mRNA (RNA, MESSENGER). The specific dsRNAs are processed into SMALL INTERFERING RNA (siRNA) which serves as a guide for cleavage of the homologous mRNA in the RNA-INDUCED SILENCING COMPLEX. DNA METHYLATION may also be triggered during this process.Biolistics: Techniques where DNA is delivered directly into organelles at high speed using projectiles coated with nucleic acid, shot from a helium-powered gun (gene gun). One of these techniques involves immunization by DNA VACCINES, which delivers DNA-coated gold beads to the epidermis.Regulatory Sequences, Nucleic Acid: Nucleic acid sequences involved in regulating the expression of genes.Oligonucleotides, Antisense: Short fragments of DNA or RNA that are used to alter the function of target RNAs or DNAs to which they hybridize.Enhancer Elements, Genetic: Cis-acting DNA sequences which can increase transcription of genes. Enhancers can usually function in either orientation and at various distances from a promoter.Luminescent Proteins: Proteins which are involved in the phenomenon of light emission in living systems. Included are the "enzymatic" and "non-enzymatic" types of system with or without the presence of oxygen or co-factors.Phosphorylation: The introduction of a phosphoryl group into a compound through the formation of an ester bond between the compound and a phosphorus moiety.Cell Proliferation: All of the processes involved in increasing CELL NUMBER including CELL DIVISION.Up-Regulation: A positive regulatory effect on physiological processes at the molecular, cellular, or systemic level. At the molecular level, the major regulatory sites include membrane receptors, genes (GENE EXPRESSION REGULATION), mRNAs (RNA, MESSENGER), and proteins.Mutagenesis, Site-Directed: Genetically engineered MUTAGENESIS at a specific site in the DNA molecule that introduces a base substitution, or an insertion or deletion.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.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.Particle Size: Relating to the size of solids.Transgenes: Genes that are introduced into an organism using GENE TRANSFER TECHNIQUES.Flow Cytometry: Technique using an instrument system for making, processing, and displaying one or more measurements on individual cells obtained from a cell suspension. Cells are usually stained with one or more fluorescent dyes specific to cell components of interest, e.g., DNA, and fluorescence of each cell is measured as it rapidly transverses the excitation beam (laser or mercury arc lamp). Fluorescence provides a quantitative measure of various biochemical and biophysical properties of the cell, as well as a basis for cell sorting. Other measurable optical parameters include light absorption and light scattering, the latter being applicable to the measurement of cell size, shape, density, granularity, and stain uptake.Proto-Oncogene Proteins: Products of proto-oncogenes. Normally they do not have oncogenic or transforming properties, but are involved in the regulation or differentiation of cell growth. They often have protein kinase activity.Adenoviridae: A family of non-enveloped viruses infecting mammals (MASTADENOVIRUS) and birds (AVIADENOVIRUS) or both (ATADENOVIRUS). Infections may be asymptomatic or result in a variety of diseases.NF-kappa B: Ubiquitous, inducible, nuclear transcriptional activator that binds to enhancer elements in many different cell types and is activated by pathogenic stimuli. The NF-kappa B complex is a heterodimer composed of two DNA-binding subunits: NF-kappa B1 and relA.Nanoparticles: Nanometer-sized particles that are nanoscale in three dimensions. They include nanocrystaline materials; NANOCAPSULES; METAL NANOPARTICLES; DENDRIMERS, and QUANTUM DOTS. The uses of nanoparticles include DRUG DELIVERY SYSTEMS and cancer targeting and imaging.Response Elements: Nucleotide sequences, usually upstream, which are recognized by specific regulatory transcription factors, thereby causing gene response to various regulatory agents. These elements may be found in both promoter and enhancer regions.Fatty Acids, Monounsaturated: Fatty acids which are unsaturated in only one position.Mice, Nude: Mutant mice homozygous for the recessive gene "nude" which fail to develop a thymus. They are useful in tumor studies and studies on immune responses.Restriction Mapping: Use of restriction endonucleases to analyze and generate a physical map of genomes, genes, or other segments of DNA.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.Gene Silencing: Interruption or suppression of the expression of a gene at transcriptional or translational levels.Cell Transformation, Neoplastic: Cell changes manifested by escape from control mechanisms, increased growth potential, alterations in the cell surface, karyotypic abnormalities, morphological and biochemical deviations from the norm, and other attributes conferring the ability to invade, metastasize, and kill.Sequence Deletion: Deletion of sequences of nucleic acids from the genetic material of an individual.Drug Carriers: Forms to which substances are incorporated to improve the delivery and the effectiveness of drugs. Drug carriers are used in drug-delivery systems such as the controlled-release technology to prolong in vivo drug actions, decrease drug metabolism, and reduce drug toxicity. Carriers are also used in designs to increase the effectiveness of drug delivery to the target sites of pharmacological actions. Liposomes, albumin microspheres, soluble synthetic polymers, DNA complexes, protein-drug conjugates, and carrier erythrocytes among others have been employed as biodegradable drug carriers.Cell Differentiation: Progressive restriction of the developmental potential and increasing specialization of function that leads to the formation of specialized cells, tissues, and organs.Carrier Proteins: Transport proteins that carry specific substances in the blood or across cell membranes.Microscopy, Fluorescence: Microscopy of specimens stained with fluorescent dye (usually fluorescein isothiocyanate) or of naturally fluorescent materials, which emit light when exposed to ultraviolet or blue light. Immunofluorescence microscopy utilizes antibodies that are labeled with fluorescent dye.Electrophoretic Mobility Shift Assay: An electrophoretic technique for assaying the binding of one compound to another. Typically one compound is labeled to follow its mobility during electrophoresis. If the labeled compound is bound by the other compound, then the mobility of the labeled compound through the electrophoretic medium will be retarded.Time Factors: Elements of limited time intervals, contributing to particular results or situations.Genes, Viral: The functional hereditary units of VIRUSES.Polymers: Compounds formed by the joining of smaller, usually repeating, units linked by covalent bonds. These compounds often form large macromolecules (e.g., BIOPOLYMERS; PLASTICS).Gene Expression Regulation, Viral: Any of the processes by which cytoplasmic factors influence the differential control of gene action in viruses.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.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.Enzyme Activation: Conversion of an inactive form of an enzyme to one possessing metabolic activity. It includes 1, activation by ions (activators); 2, activation by cofactors (coenzymes); and 3, conversion of an enzyme precursor (proenzyme or zymogen) to an active enzyme.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.MicroRNAs: Small double-stranded, non-protein coding RNAs, 21-25 nucleotides in length generated from single-stranded microRNA gene transcripts by the same RIBONUCLEASE III, Dicer, that produces small interfering RNAs (RNA, SMALL INTERFERING). They become part of the RNA-INDUCED SILENCING COMPLEX and repress the translation (TRANSLATION, GENETIC) of target RNA by binding to homologous 3'UTR region as an imperfect match. The small temporal RNAs (stRNAs), let-7 and lin-4, from C. elegans, are the first 2 miRNAs discovered, and are from a class of miRNAs involved in developmental timing.Oncogenes: Genes whose gain-of-function alterations lead to NEOPLASTIC CELL TRANSFORMATION. They include, for example, genes for activators or stimulators of CELL PROLIFERATION such as growth factors, growth factor receptors, protein kinases, signal transducers, nuclear phosphoproteins, and transcription factors. A prefix of "v-" before oncogene symbols indicates oncogenes captured and transmitted by RETROVIRUSES; the prefix "c-" before the gene symbol of an oncogene indicates it is the cellular homolog (PROTO-ONCOGENES) of a v-oncogene.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.NIH 3T3 Cells: A continuous cell line of high contact-inhibition established from NIH Swiss mouse embryo cultures. The cells are useful for DNA transfection and transformation studies. (From ATCC [Internet]. Virginia: American Type Culture Collection; c2002 [cited 2002 Sept 26]. Available from http://www.atcc.org/)Lipids: A generic term for fats and lipoids, the alcohol-ether-soluble constituents of protoplasm, which are insoluble in water. They comprise the fats, fatty oils, essential oils, waxes, phospholipids, glycolipids, sulfolipids, aminolipids, chromolipids (lipochromes), and fatty acids. (Grant & Hackh's Chemical Dictionary, 5th ed)Viral Proteins: Proteins found in any species of virus.Epithelial Cells: Cells that line the inner and outer surfaces of the body by forming cellular layers (EPITHELIUM) or masses. Epithelial cells lining the SKIN; the MOUTH; the NOSE; and the ANAL CANAL derive from ectoderm; those lining the RESPIRATORY SYSTEM and the DIGESTIVE SYSTEM derive from endoderm; others (CARDIOVASCULAR SYSTEM and LYMPHATIC SYSTEM) derive from mesoderm. Epithelial cells can be classified mainly by cell shape and function into squamous, glandular and transitional epithelial cells.Neoplasm Proteins: Proteins whose abnormal expression (gain or loss) are associated with the development, growth, or progression of NEOPLASMS. Some neoplasm proteins are tumor antigens (ANTIGENS, NEOPLASM), i.e. they induce an immune reaction to their tumor. Many neoplasm proteins have been characterized and are used as tumor markers (BIOMARKERS, TUMOR) when they are detectable in cells and body fluids as monitors for the presence or growth of tumors. Abnormal expression of ONCOGENE PROTEINS is involved in neoplastic transformation, whereas the loss of expression of TUMOR SUPPRESSOR PROTEINS is involved with the loss of growth control and progression of the neoplasm.Genes, ras: Family of retrovirus-associated DNA sequences (ras) originally isolated from Harvey (H-ras, Ha-ras, rasH) and Kirsten (K-ras, Ki-ras, rasK) murine sarcoma viruses. Ras genes are widely conserved among animal species and sequences corresponding to both H-ras and K-ras genes have been detected in human, avian, murine, and non-vertebrate genomes. The closely related N-ras gene has been detected in human neuroblastoma and sarcoma cell lines. All genes of the family have a similar exon-intron structure and each encodes a p21 protein.Cricetulus: A genus of the family Muridae consisting of eleven species. C. migratorius, the grey or Armenian hamster, and C. griseus, the Chinese hamster, are the two species used in biomedical research.Microscopy, Confocal: A light microscopic technique in which only a small spot is illuminated and observed at a time. An image is constructed through point-by-point scanning of the field in this manner. Light sources may be conventional or laser, and fluorescence or transmitted observations are possible.DEAE-Dextran: Used as a support for ion-exchange chromatography.Enzyme Inhibitors: Compounds or agents that combine with an enzyme in such a manner as to prevent the normal substrate-enzyme combination and the catalytic reaction.Carcinoma, Hepatocellular: A primary malignant neoplasm of epithelial liver cells. It ranges from a well-differentiated tumor with EPITHELIAL CELLS indistinguishable from normal HEPATOCYTES to a poorly differentiated neoplasm. The cells may be uniform or markedly pleomorphic, or form GIANT CELLS. Several classification schemes have been suggested.Repressor Proteins: Proteins which maintain the transcriptional quiescence of specific GENES or OPERONS. Classical repressor proteins are DNA-binding proteins that are normally bound to the OPERATOR REGION of an operon, or the ENHANCER SEQUENCES of a gene until a signal occurs that causes their release.Ultrasonics: A subfield of acoustics dealing in the radio frequency range higher than acoustic SOUND waves (approximately above 20 kilohertz). Ultrasonic radiation is used therapeutically (DIATHERMY and ULTRASONIC THERAPY) to generate HEAT and to selectively destroy tissues. It is also used in diagnostics, for example, ULTRASONOGRAPHY; ECHOENCEPHALOGRAPHY; and ECHOCARDIOGRAPHY, to visually display echoes received from irradiated tissues.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.HEK293 Cells: A cell line generated from human embryonic kidney cells that were transformed with human adenovirus type 5.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.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.Immunohistochemistry: Histochemical localization of immunoreactive substances using labeled antibodies as reagents.Cell Movement: The movement of cells from one location to another. Distinguish from CYTOKINESIS which is the process of dividing the CYTOPLASM of a cell.Phenotype: The outward appearance of the individual. It is the product of interactions between genes, and between the GENOTYPE and the environment.Kinetics: The rate dynamics in chemical or physical systems.Immediate-Early Proteins: Proteins that are coded by immediate-early genes, in the absence of de novo protein synthesis. The term was originally used exclusively for viral regulatory proteins that were synthesized just after viral integration into the host cell. It is also used to describe cellular proteins which are synthesized immediately after the resting cell is stimulated by extracellular signals.Protein-Serine-Threonine Kinases: A group of enzymes that catalyzes the phosphorylation of serine or threonine residues in proteins, with ATP or other nucleotides as phosphate donors.RNA, Viral: Ribonucleic acid that makes up the genetic material of viruses.Simian virus 40: A species of POLYOMAVIRUS originally isolated from Rhesus monkey kidney tissue. It produces malignancy in human and newborn hamster kidney cell cultures.Chitosan: Deacetylated CHITIN, a linear polysaccharide of deacetylated beta-1,4-D-glucosamine. It is used in HYDROGEL and to treat WOUNDS.Cell Adhesion: Adherence of cells to surfaces or to other cells.Rats, Sprague-Dawley: A strain of albino rat used widely for experimental purposes because of its calmness and ease of handling. It was developed by the Sprague-Dawley Animal Company.Transcription Factor AP-1: A multiprotein complex composed of the products of c-jun and c-fos proto-oncogenes. These proteins must dimerize in order to bind to the AP-1 recognition site, also known as the TPA-responsive element (TRE). AP-1 controls both basal and inducible transcription of several genes.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.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.Oligodeoxyribonucleotides, Antisense: Short fragments of DNA that are used to alter the function of target RNAs or DNAs to which they hybridize.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.Fluorescent Antibody Technique: Test for tissue antigen using either a direct method, by conjugation of antibody with fluorescent dye (FLUORESCENT ANTIBODY TECHNIQUE, DIRECT) or an indirect method, by formation of antigen-antibody complex which is then labeled with fluorescein-conjugated anti-immunoglobulin antibody (FLUORESCENT ANTIBODY TECHNIQUE, INDIRECT). The tissue is then examined by fluorescence microscopy.Mice, Inbred BALB CRNA: 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)L Cells (Cell Line): A cultured line of C3H mouse FIBROBLASTS that do not adhere to one another and do not express CADHERINS.Membrane Glycoproteins: Glycoproteins found on the membrane or surface of cells.Cell Transformation, Viral: An inheritable change in cells manifested by changes in cell division and growth and alterations in cell surface properties. It is induced by infection with a transforming virus.Cell Cycle: The complex series of phenomena, occurring between the end of one CELL DIVISION and the end of the next, by which cellular material is duplicated and then divided between two daughter cells. The cell cycle includes INTERPHASE, which includes G0 PHASE; G1 PHASE; S PHASE; and G2 PHASE, and CELL DIVISION PHASE.Deoxyribonuclease I: An enzyme capable of hydrolyzing highly polymerized DNA by splitting phosphodiester linkages, preferentially adjacent to a pyrimidine nucleotide. This catalyzes endonucleolytic cleavage of DNA yielding 5'-phosphodi- and oligonucleotide end-products. The enzyme has a preference for double-stranded DNA.Polyethylene Glycols: Polymers of ETHYLENE OXIDE and water, and their ethers. They vary in consistency from liquid to solid depending on the molecular weight indicated by a number following the name. They are used as SURFACTANTS, dispersing agents, solvents, ointment and suppository bases, vehicles, and tablet excipients. Some specific groups are NONOXYNOLS, OCTOXYNOLS, and POLOXAMERS.Immunoblotting: Immunologic method used for detecting or quantifying immunoreactive substances. The substance is identified by first immobilizing it by blotting onto a membrane and then tagging it with labeled antibodies.Dose-Response Relationship, Drug: The relationship between the dose of an administered drug and the response of the organism to the drug.Microbubbles: Small encapsulated gas bubbles (diameters of micrometers) that can be used as CONTRAST MEDIA, and in other diagnostic and therapeutic applications. Upon exposure to sufficiently intense ultrasound, microbubbles will cavitate, rupture, disappear, release gas content. Such characteristics of the microbubbles can be used to enhance diagnostic tests, dissolve blood clots, and deliver drugs or genes for therapy.Liver: A large lobed glandular organ in the abdomen of vertebrates that is responsible for detoxification, metabolism, synthesis and storage of various substances.RNA, Antisense: RNA molecules which hybridize to complementary sequences in either RNA or DNA altering the function of the latter. Endogenous antisense RNAs function as regulators of gene expression by a variety of mechanisms. Synthetic antisense RNAs are used to effect the functioning of specific genes for investigative or therapeutic purposes.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.3' Untranslated Regions: The sequence at the 3' end of messenger RNA that does not code for product. This region contains transcription and translation regulating sequences.Precipitin Tests: Serologic tests in which a positive reaction manifested by visible CHEMICAL PRECIPITATION occurs when a soluble ANTIGEN reacts with its precipitins, i.e., ANTIBODIES that can form a precipitate.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.Cytoplasm: The part of a cell that contains the CYTOSOL and small structures excluding the CELL NUCLEUS; MITOCHONDRIA; and large VACUOLES. (Glick, Glossary of Biochemistry and Molecular Biology, 1990)Mice, Inbred C57BLBacteriophages: Viruses whose hosts are bacterial cells.Gene Knockdown Techniques: The artificial induction of GENE SILENCING by the use of RNA INTERFERENCE to reduce the expression of a specific gene. It includes the use of DOUBLE-STRANDED RNA, such as SMALL INTERFERING RNA and RNA containing HAIRPIN LOOP SEQUENCE, and ANTI-SENSE OLIGONUCLEOTIDES.Liver Neoplasms: Tumors or cancer of the LIVER.Mutagenesis: Process of generating a genetic MUTATION. It may occur spontaneously or be induced by MUTAGENS.Oligonucleotides: Polymers made up of a few (2-20) nucleotides. In molecular genetics, they refer to a short sequence synthesized to match a region where a mutation is known to occur, and then used as a probe (OLIGONUCLEOTIDE PROBES). (Dorland, 28th ed)PhosphoproteinsProtein Isoforms: Different forms of a protein that may be produced from different GENES, or from the same gene by ALTERNATIVE SPLICING.Mitogen-Activated Protein Kinases: A superfamily of PROTEIN-SERINE-THREONINE KINASES that are activated by diverse stimuli via protein kinase cascades. They are the final components of the cascades, activated by phosphorylation by MITOGEN-ACTIVATED PROTEIN KINASE KINASES, which in turn are activated by mitogen-activated protein kinase kinase kinases (MAP KINASE KINASE KINASES).Neoplasm Transplantation: Experimental transplantation of neoplasms in laboratory animals for research purposes.5' Flanking Region: The region of DNA which borders the 5' end of a transcription unit and where a variety of regulatory sequences are located.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.Kidney: Body organ that filters blood for the secretion of URINE and that regulates ion concentrations.Thymidine Kinase: An enzyme that catalyzes the conversion of ATP and thymidine to ADP and thymidine 5'-phosphate. Deoxyuridine can also act as an acceptor and dGTP as a donor. (From Enzyme Nomenclature, 1992) EC 2.7.1.21.Point Mutation: A mutation caused by the substitution of one nucleotide for another. This results in the DNA molecule having a change in a single base pair.Homeodomain Proteins: Proteins encoded by homeobox genes (GENES, HOMEOBOX) that exhibit structural similarity to certain prokaryotic and eukaryotic DNA-binding proteins. Homeodomain proteins are involved in the control of gene expression during morphogenesis and development (GENE EXPRESSION REGULATION, DEVELOPMENTAL).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.Proto-Oncogene Proteins c-akt: A protein-serine-threonine kinase that is activated by PHOSPHORYLATION in response to GROWTH FACTORS or INSULIN. It plays a major role in cell metabolism, growth, and survival as a core component of SIGNAL TRANSDUCTION. Three isoforms have been described in mammalian cells.PolyaminesTumor Suppressor Protein p53: Nuclear phosphoprotein encoded by the p53 gene (GENES, P53) whose normal function is to control CELL PROLIFERATION and APOPTOSIS. A mutant or absent p53 protein has been found in LEUKEMIA; OSTEOSARCOMA; LUNG CANCER; and COLORECTAL CANCER.DNA Footprinting: A method for determining the sequence specificity of DNA-binding proteins. DNA footprinting utilizes a DNA damaging agent (either a chemical reagent or a nuclease) which cleaves DNA at every base pair. DNA cleavage is inhibited where the ligand binds to DNA. (from Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed)Genome, Viral: The complete genetic complement contained in a DNA or RNA molecule in a virus.Jurkat Cells: A CELL LINE derived from human T-CELL LEUKEMIA and used to determine the mechanism of differential susceptibility to anti-cancer drugs and radiation.Isoenzymes: Structurally related forms of an enzyme. Each isoenzyme has the same mechanism and classification, but differs in its chemical, physical, or immunological characteristics.Protein Transport: The process of moving proteins from one cellular compartment (including extracellular) to another by various sorting and transport mechanisms such as gated transport, protein translocation, and vesicular transport.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.Genes, Dominant: Genes that influence the PHENOTYPE both in the homozygous and the heterozygous state.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.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).Protamines: A group of simple proteins that yield basic amino acids on hydrolysis and that occur combined with nucleic acid in the sperm of fish. Protamines contain very few kinds of amino acids. Protamine sulfate combines with heparin to form a stable inactive complex; it is used to neutralize the anticoagulant action of heparin in the treatment of heparin overdose. (From Merck Index, 11th ed; Martindale, The Extra Pharmacopoeia, 30th ed, p692)Cell Membrane: The lipid- and protein-containing, selectively permeable membrane that surrounds the cytoplasm in prokaryotic and eukaryotic cells.Retroviridae: Family of RNA viruses that infects birds and mammals and encodes the enzyme reverse transcriptase. The family contains seven genera: DELTARETROVIRUS; LENTIVIRUS; RETROVIRUSES TYPE B, MAMMALIAN; ALPHARETROVIRUS; GAMMARETROVIRUS; RETROVIRUSES TYPE D; and SPUMAVIRUS. A key feature of retrovirus biology is the synthesis of a DNA copy of the genome which is integrated into cellular DNA. After integration it is sometimes not expressed but maintained in a latent state (PROVIRUSES).Receptors, Cytoplasmic and Nuclear: Intracellular receptors that can be found in the cytoplasm or in the nucleus. They bind to extracellular signaling molecules that migrate through or are transported across the CELL MEMBRANE. Many members of this class of receptors occur in the cytoplasm and are transported to the CELL NUCLEUS upon ligand-binding where they signal via DNA-binding and transcription regulation. Also included in this category are receptors found on INTRACELLULAR MEMBRANES that act via mechanisms similar to CELL SURFACE RECEPTORS.Luminescent Agents: Compound such as LUMINESCENT PROTEINS that cause or emit light (PHYSICAL LUMINESCENCE).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.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.Proto-Oncogene Proteins c-jun: Cellular DNA-binding proteins encoded by the c-jun genes (GENES, JUN). They are involved in growth-related transcriptional control. There appear to be three distinct functions: dimerization (with c-fos), DNA-binding, and transcriptional activation. Oncogenic transformation can take place by constitutive expression of c-jun.Hep G2 Cells: A human liver tumor cell line used to study a variety of liver-specific metabolic functions.CCAAT-Enhancer-Binding Proteins: A class of proteins that were originally identified by their ability to bind the DNA sequence CCAAT. The typical CCAAT-enhancer binding protein forms dimers and consists of an activation domain, a DNA-binding basic region, and a leucine-rich dimerization domain (LEUCINE ZIPPERS). CCAAT-BINDING FACTOR is structurally distinct type of CCAAT-enhancer binding protein consisting of a trimer of three different subunits.DNA Mutational Analysis: Biochemical identification of mutational changes in a nucleotide sequence.

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To date, all my transfections were unsuccessfull. Does , anybody have any ideas why the transfections won't work? , , Thanks , ... Transfection problems. John Ladasky jladasky at pmgm.Stanford.EDU Thu Jan 29 18:50:09 EST 1998 *Previous message: Transfection ... construct using the calsium phosphate method of transfection. The DNA I , used for this transfection was isolated with ... Rentia I have used Promega's Wizard minipreps and maxipreps to prepare plasmid DNA that was suitable for transfection. The cell ...

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*  Transfections: post #1

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*  RV- transfection-transduction problem: post #1

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*  transfection of suspension line cells - Molecular Biology

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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.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.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.GC 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.Coles PhillipsPolyethylenimineDNA 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.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.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.Spatiotemporal gene expressionGene electrotransfer: Gene electrotransfer is a versatile biotechnology technique that enables the transfer of genetic material into prokaryotic or eukaryotic cells. It is based on a physical method named electroporation, where a transient increase in the permeability of cell membrane is achieved when submitted to short and intense electric pulses, thus enabling the transport of large molecules (naked plasmid DNA, antisense oligonucleotides, siRNA) into cells that otherwise cannot permeate through the cell membrane.LiposomeDNA-binding proteinPituitary-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.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.Synapto-pHluorin: Synapto-pHluorin is a genetically encoded optical indicator of vesicle release and recycling. It is used in neuroscience to study transmitter release.Protein 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.Spinel group: The spinels are any of a class of minerals of general formulation A2+B3+2O2−4 which crystallise in the cubic (isometric) crystal system, with the oxide anions arranged in a cubic close-packed lattice and the cations A and B occupying some or all of the octahedral and tetrahedral sites in the lattice. Although the charges of A and B in the prototypical spinel structure are +2 and +3, respectively, other combinations incorporating divalent, trivalent, or tetravalent cations, including magnesium, zinc, iron, manganese, aluminium, chromium, titanium, and silicon, are also possible.Baby hamster kidney cell: Baby Hamster Kidney fibroblasts (aka BHK cells) are an adherent cell line used in molecular biology.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.PolylysineLipofectamine: Lipofectamine or Lipofectamine 2000 is a common transfection reagent, produced and sold by Invitrogen, used in molecular and cellular biology. Invitrogen (2012).DNA binding site: DNA binding sites are a type of binding site found in DNA where other molecules may bind. DNA binding sites are distinct from other binding sites in that (1) they are part of a DNA sequence (e.Tetramethylammonium chlorideProximity ligation assay: Proximity ligation assay (in situ PLA) is a technology that extends the capabilities of traditional immunoassays to include direct detection of proteins, protein interactions and modifications with high specificity and sensitivity. Protein targets can be readily detected and localized with single molecule resolution and objectively quantified in unmodified cells and tissues.Dermal fibroblast: Dermal fibroblasts are cells within the dermis layer of skin which are responsible for generating connective tissue and allowing the skin to recover from injury. Using organelles (particularly the rough endoplasmic reticulum), dermal fibroblasts generate and maintain the connective tissue which unites separate cell layers.MinC: The MinC protein is one of three proteins encoded by the minB operon and which is required to generate pole to pole oscillations prior to bacterial cell division as a means of specifying the midzone of the cell. This function is achieved by preventing the formation of the divisome Z-ring around the poles.Senescence-associated beta-galactosidase: Senescence-associated beta-galactosidase (SA-β-gal or SABG) is a hypothetical hydrolase enzyme that catalyzes the hydrolysis of β-galactosides into monosaccharides only in senescent cells.RNAi-Based Identification System and interference of Specific Cancer Cells: A “classifier” was created to classify cells by identifying specific characteristics of Cervical Cancer. These characteristics were consistent with HeLa cells, which served as the target cell line for cell death.Gene gunAntisense therapy: Antisense therapy is a form of treatment for genetic disorders or infections. When the genetic sequence of a particular gene is known to be causative of a particular disease, it is possible to synthesize a strand of nucleic acid (DNA, RNA or a chemical analogue) that will bind to the messenger RNA (mRNA) produced by that gene and inactivate it, effectively turning that gene "off".Enhancer (genetics)Citrine (colour): Citrine is a colour], the most common reference for which is certain coloured varieties of [[quartz which are a medium deep shade of golden yellow. Citrine has been summarized at various times as yellow, greenish-yellow, brownish yellow or orange.Hyperphosphorylation: Hyperphosphorylation occurs when a biochemical with multiple phosphorylation sites is fully saturated. Hyperphosphorylation is one of the signalling mechanisms used by the cell to regulate mitosis.Coulter counter: 150px|thumb|right|The tip of the Coulter counter in a buffer solution, counting cells in solution.Flow cytometry: In biotechnology, flow cytometry is a laser-based, biophysical technology employed in cell counting, cell sorting, biomarker detection and protein engineering, by suspending cells in a stream of fluid and passing them by an electronic detection apparatus. It allows simultaneous multiparametric analysis of the physical and chemical characteristics of up to thousands of particles per second.Nanoparticle: Nanoparticles are particles between 1 and 100 nanometers in size. In nanotechnology, a particle is defined as a small object that behaves as a whole unit with respect to its transport and properties.Lipokine: A lipokine is a lipid-controlling hormone. The term "lipokine" was first used by Haiming Cao in 2008 to classify fatty acids which modulate lipid metabolism by what he called a "chaperone effect".Nude mouseThermal cyclerCharged Aerosol Release Experiment: The Charged Aerosol Release Experiment also known as CARE, is a project run by NASA which will use a rocket to release dust in the upper atmosphere to form a dusty plasma in space. NASA plans to trigger cloud formation around the rocket's exhaust particles.Total internal reflection fluorescence microscope: A total internal reflection fluorescence microscope (TIRFM) is a type of microscope with which a thin region of a specimen, usually less than 200 nm can be observed.Temporal analysis of products: Temporal Analysis of Products (TAP), (TAP-2), (TAP-3) is an experimental technique for studying

(1/63574) 11q23.1 and 11q25-qter YACs suppress tumour growth in vivo.

Frequent allelic deletion at chromosome 11q22-q23.1 has been described in breast cancer and a number of other malignancies, suggesting putative tumour suppressor gene(s) within the approximately 8 Mb deleted region. In addition, we recently described another locus, at the 11q25-qter region, frequently deleted in breast cancer, suggesting additional tumour suppressor gene(s) in this approximately 2 Mb deleted region. An 11q YAC contig was accessed and three YACs, one containing the candidate gene ATM at 11q23.1, and two contiguous YACs (overlapping for approximately 400-600 kb) overlying most of the 11q25 deleted region, were retrofitted with a G418 resistance marker and transfected into murine A9 fibrosarcoma cells. Selected A9 transfectant clones (and control untransfected and 'irrelevant' alphoid YAC transfectant A9 clones) were assayed for in vivo tumorigenicity in athymic female Balb c-nu/nu mice. All the 11q YAC transfectant clones demonstrated significant tumour suppression compared to the control untransfected and 'irrelevant' YAC transfected A9 cells. These results define two discrete tumour suppressor loci on chromosome 11q by functional complementation, one to a approximately 1.2 Mb region on 11q23.1 (containing the ATM locus) and another to a approximately 400-600 kb subterminal region on 11q25-qter.  (+info)

(2/63574) 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)

(3/63574) Expression of the naturally occurring truncated trkB neurotrophin receptor induces outgrowth of filopodia and processes in neuroblastoma cells.

We have investigated the effects of the truncated trkB receptor isoform T1 (trkB.T1) by transient transfection into mouse N2a neuroblastoma cells. We observed that expression of trkB.T1 leads to a striking change in cell morphology characterized by outgrowth of filopodia and processes. A similar morphological response was also observed in SH-SY5Y human neuroblastoma cells and NIH3T3 fibroblasts transfected with trkB.T1. N2a cells lack endogenous expression of trkB isoforms, but express barely detectable amounts of its ligands, brain-derived neurotrophic factor (BDNF) and neurotrophin-4 (NT-4). The morphological change was ligand-independent, since addition of exogenous BDNF or NT-4 or blockade of endogenous trkB ligands did not influence this response. Filopodia and process outgrowth was significantly suppressed when full-length trkB.TK+ was cotransfected together with trkB.T1 and this inhibitory effect was blocked by tyrosine kinase inhibitor K252a. Transfection of trkB.T1 deletion mutants showed that the morphological response is dependent on the extracellular, but not the intracellular domain of the receptor. Our results suggest a novel ligand-independent role for truncated trkB in the regulation of cellular morphology.  (+info)

(4/63574) B-MYB transactivates its own promoter through SP1-binding sites.

B-MYB is an ubiquitous protein required for mammalian cell growth. In this report we show that B-MYB transactivates its own promoter through a 120 bp segment proximal to the transcription start site. The B-MYB-responsive element does not contain myb-binding sites and gel-shift analysis shows that SP1, but not B-MYB, protein contained in SAOS2 cell extracts binds to the 120 bp B-myb promoter fragment. B-MYB-dependent transactivation is cooperatively increased in the presence of SP1, but not SP3 overexpression. When the SP1 elements of the B-myb promoter are transferred in front of a heterologous promoter, an increased response to B-MYB results. In contrast, c-MYB, the prototype member of the Myb family, is not able to activate the luciferase construct containing the SP1 elements. With the use of an SP1-GAL4 fusion protein, we have determined that the cooperative activation occurs through the domain A of SP1. These observations suggest that B-MYB functions as a coactivator of SP1, and that diverse combinations of myb and SP1 sites may dictate the responsiveness of myb-target genes to the various members of the myb family.  (+info)

(5/63574) Arrestin function in G protein-coupled receptor endocytosis requires phosphoinositide binding.

Internalization of agonist-activated G protein-coupled receptors is mediated by non-visual arrestins, which also bind to clathrin and are therefore thought to act as adaptors in the endocytosis process. Phosphoinositides have been implicated in the regulation of intracellular receptor trafficking, and are known to bind to other coat components including AP-2, AP180 and COPI coatomer. Given these observations, we explored the possibility that phosphoinositides play a role in arrestin's function as an adaptor. High-affinity binding sites for phosphoinositides in beta-arrestin (arrestin2) and arrestin3 (beta-arrestin2) were identified, and dissimilar effects of phosphoinositide and inositol phosphate on arrestin interactions with clathrin and receptor were characterized. Alteration of three basic residues in arrestin3 abolished phosphoinositide binding with complete retention of clathrin and receptor binding. Unlike native protein, upon agonist activation, this mutant arrestin3 expressed in COS1 cells neither supported beta2-adrenergic receptor internalization nor did it concentrate in coated pits, although it was recruited to the plasma membrane. These findings indicate that phosphoinositide binding plays a critical regulatory role in delivery of the receptor-arrestin complex to coated pits, perhaps by providing, with activated receptor, a multi-point attachment of arrestin to the plasma membrane.  (+info)

(6/63574) The MAP kinase ERK2 inhibits the cyclic AMP-specific phosphodiesterase HSPDE4D3 by phosphorylating it at Ser579.

The extracellular receptor stimulated kinase ERK2 (p42(MAPK))-phosphorylated human cAMP-specific phosphodiesterase PDE4D3 at Ser579 and profoundly reduced ( approximately 75%) its activity. These effects could be reversed by the action of protein phosphatase PP1. The inhibitory state of PDE4D3, engendered by ERK2 phosphorylation, was mimicked by the Ser579-->Asp mutant form of PDE4D3. In COS1 cells transfected to express PDE4D3, challenge with epidermal growth factor (EGF) caused the phosphorylation and inhibition of PDE4D3. This effect was blocked by the MEK inhibitor PD98059 and was not apparent using the Ser579-->Ala mutant form of PDE4D3. Challenge of HEK293 and F442A cells with EGF led to the PD98059-ablatable inhibition of endogenous PDE4D3 and PDE4D5 activities. EGF challenge of COS1 cells transfected to express PDE4D3 increased cAMP levels through a process ablated by PD98059. The activity of the Ser579-->Asp mutant form of PDE4D3 was increased by PKA phosphorylation. The transient form of the EGF-induced inhibition of PDE4D3 is thus suggested to be due to feedback regulation by PKA causing the ablation of the ERK2-induced inhibition of PDE4D3. We identify a novel means of cross-talk between the cAMP and ERK signalling pathways whereby cell stimuli that lead to ERK2 activation may modulate cAMP signalling.  (+info)

(7/63574) Coupling of the cell cycle and myogenesis through the cyclin D1-dependent interaction of MyoD with cdk4.

Proliferating myoblasts express the muscle determination factor, MyoD, throughout the cell cycle in the absence of differentiation. Here we show that a mitogen-sensitive mechanism, involving the direct interaction between MyoD and cdk4, restricts myoblast differentiation to cells that have entered into the G0 phase of the cell cycle under mitogen withdrawal. Interaction between MyoD and cdk4 disrupts MyoD DNA-binding, muscle-specific gene activation and myogenic conversion of 10T1/2 cells independently of cyclin D1 and the CAK activation of cdk4. Forced induction of cyclin D1 in myotubes results in the cytoplasmic to nuclear translocation of cdk4. The specific MyoD-cdk4 interaction in dividing myoblasts, coupled with the cyclin D1-dependent nuclear targeting of cdk4, suggests a mitogen-sensitive mechanism whereby cyclin D1 can regulate MyoD function and the onset of myogenesis by controlling the cellular location of cdk4 rather than the phosphorylation status of MyoD.  (+info)

(8/63574) Assembly requirements of PU.1-Pip (IRF-4) activator complexes: inhibiting function in vivo using fused dimers.

Gene expression in higher eukaryotes appears to be regulated by specific combinations of transcription factors binding to regulatory sequences. The Ets factor PU.1 and the IRF protein Pip (IRF-4) represent a pair of interacting transcription factors implicated in regulating B cell-specific gene expression. Pip is recruited to its binding site on DNA by phosphorylated PU.1. PU.1-Pip interaction is shown to be template directed and involves two distinct protein-protein interaction surfaces: (i) the ets and IRF DNA-binding domains; and (ii) the phosphorylated PEST region of PU.1 and a lysine-requiring putative alpha-helix in Pip. Thus, a coordinated set of protein-protein and protein-DNA contacts are essential for PU.1-Pip ternary complex assembly. To analyze the function of these factors in vivo, we engineered chimeric repressors containing the ets and IRF DNA-binding domains connected by a flexible POU domain linker. When stably expressed, the wild-type fused dimer strongly repressed the expression of a rearranged immunoglobulin lambda gene, thereby establishing the functional importance of PU.1-Pip complexes in B cell gene expression. Comparative analysis of the wild-type dimer with a series of mutant dimers distinguished a gene regulated by PU.1 and Pip from one regulated by PU.1 alone. This strategy should prove generally useful in analyzing the function of interacting transcription factors in vivo, and for identifying novel genes regulated by such complexes.  (+info)



Reagents

  • Transfection reagents introduce nucleic acid constructs into eukaryotic cells for protein synthesis and the overexpression of mutant DNA constructs. (atcc.org)
  • ATCC ® Transfection reagents may be used for the transfer of nucleic acids for a variety of applications, including protein expression and the overexpression of mutant genes. (atcc.org)
  • due to their positive charge, these transfection reagents interact with the negatively charged backbone of nucleic acids and cell membranes. (atcc.org)
  • These exceptional transfection reagents, combined with ATCC's technical know-how, will enable your transcriptional and translational gene studies. (atcc.org)
  • ATCC offers a line of transfection reagents tailored for effective nucleic acid transfer into a wide variety of cells for a multitude of applications. (atcc.org)
  • siRNAs can be transiently transfected using commonly available transfection reagents. (bio-medicine.org)
  • The siPORT Transfection II Kit contains samples of both of these reagents as well as positive and negative control siRNAs for protocol optimization. (bio-medicine.org)
  • All commercial transfection reagents are proprietary formulations that are competing for market share by claiming certain advantages (ie, broad cell type compatibility, low cytotoxicity, high efficiency). (openwetware.org)
  • Cationic lipid transfection reagents are suitable for transfecting into a wide variety of dividing cell cultures. (openwetware.org)
  • A major advantage of the method is the mild treatment of the cells in comparison to liposome-based transfection reagents (lipofection) and electroporation, which may result in the death of 20-50% of cells. (wikipedia.org)
  • Altogen Biosystems is a life science company that specializes in manufacturing transfection reagents. (wikipedia.org)
  • Altogen Biosystems has developed an array of optimized transfection reagents, and offers custom life science research services at its GLP-compliant laboratory. (wikipedia.org)
  • Mirus Bio (formerly Mirus Bio Corporation), develops and manufactures transfection reagents and related products for life science research. (wikipedia.org)

Reagent

  • Select the transfection reagent for your model system below. (atcc.org)
  • GeneX Plus Transfection Reagent is designed to efficiently transfect a broad spectrum of cell types. (atcc.org)
  • TransfeX™ Transfection Reagent has been optimized for use on a wide range of cell types, including cells that are generally difficult to transfect, such as hTERT immortalized cell lines, primary cells, and stem cells. (atcc.org)
  • This reagent affords high levels of gene expression in a variety of cell types and is suitable for both transient and stable transfection. (atcc.org)
  • Since the reagent is composed of animal-origin free components and is serum compatible, there is no need for any culture medium change after transfection. (atcc.org)
  • GeneX Plus Transfection Reagent has been optimized for use with HEK293T/17 SF suspension cells (ATCC ACS-4500) and HEK Plus SFM (ATCC ACS-4002) to reproducibly express a wide range of proteins at high levels. (atcc.org)
  • GeneX Plus Transfection Reagent is a high-performance, animal-origin free, broad spectrum reagent that provides exceptional transfection of plasmid DNA into mammalian cells. (atcc.org)
  • Sigma's Universal Transfection Reagent is a unique formulation of a proprietary polymer blend. (sigmaaldrich.com)
  • Universal Transfection Reagent is used for transient and stable transfection of nucleic acids into various eukaryotic cell lines and hard-to-transfect primary cells. (sigmaaldrich.com)
  • The Universal Transfection Reagent is provided as 1mL liquid in a single vial. (sigmaaldrich.com)
  • Universal Transfection Reagent can be stored at 2-8 °C for up to three months. (sigmaaldrich.com)
  • Optimal amount of Universal Transfection Reagent used for hard-to-transfect cells is generally 4 µL per 3 µg of plasmid DNA. (sigmaaldrich.com)
  • Tube B: to prepare transfection reagent, in a separate tube, add 60 µL Universal Transfection Reagent to 500 µL DMEM (high glucose, without serum). (sigmaaldrich.com)
  • and (ii) removing the non-complexed transfection reagent. (google.com)
  • Alternatively consider Xfect Single Shots transfection reagent , a high performing transfection reagent provided in super-convenient, individual lyophilized, aliquots, just add plasmid and apply to your cells. (clontech.com)
  • Have you heard about Xfect Single Shots Transfection Reagent ? (clontech.com)
  • I use the lipofectamine reagent for transfection (have also tried FuGENE 6). (protocol-online.org)
  • Fourth, the printed slide treated with transfection reagent is placed into a square dish with the printed side facing up. (wikipedia.org)
  • Effectene Reagent is used in conjunction with the enhancer and the DNA condensation buffer (Buffer EC) to achieve high transfection efficiency. (wikipedia.org)
  • Lipofectamine or Lipofectamine 2000 is a common transfection reagent, produced and sold by Invitrogen, used in molecular and cellular biology. (wikipedia.org)
  • Lipofectamine reagent contains lipid subunits that can form liposomes in an aqueous environment, which entrap the transfection payload, i.e. (wikipedia.org)

plasmid

  • Rentia I have used Promega's Wizard minipreps and maxipreps to prepare plasmid DNA that was suitable for transfection. (bio.net)
  • The procedure stated below is designed for the transfection of 293T cells with 1 µg/µl DNA plasmid (diluted in sterile molecular biology water). (sigmaaldrich.com)
  • Calcium phosphate transfection is one of the most commonly used methods to deliver plasmid DNA into mammalian cells. (clontech.com)
  • It is used to increase the transfection efficiency of RNA (including mRNA and siRNA) or plasmid DNA into in vitro cell cultures by lipofection. (wikipedia.org)

NeoFX Transfection Agent

  • siPORT NeoFX Transfection Agent refines siRNA transfection protocols resulting in less optimization. (bio-medicine.org)

vectors

  • Viral delivery of RNAi vectors is a powerful alternative to transfection for these cell types as well as for in vivo applications. (openwetware.org)
  • Non-viral methods include physical methods such as electroporation, microinjection, gene gun, impalefection, hydrostatic pressure, continuous infusion, and sonication and chemical, such as lipofection, which is a lipid-mediated DNA-transfection process utilizing liposome vectors. (wikipedia.org)
  • While DNA-based vectors (viruses, plasmids) that encode a short RNA molecule can also be used, short-RNA transfection does not risk modification of the cell's DNA, a characteristic that has led to the development of short RNA as a new class of macromolecular drugs. (wikipedia.org)
  • Besides gelatin, atelocollagen and fibronetin are also successful transfection vectors for introducing foreign DNA into the cell nucleus. (wikipedia.org)
  • This method of transfection was invented by Dr. Yongliang Chu at Life Technologies, Inc. Lipofection Transfection Vectors in gene therapy Invitrogen (2012). (wikipedia.org)

efficiency of transfection

  • 5%) and antibiotics does not inhibit transfection and in some cases increases the efficiency of transfection. (sigmaaldrich.com)
  • But also in non-dividing cells, research has shown that Lipofectamine improves the efficiency of transfection, which suggests that it additionally helps the transfected genetic material penetrate the intact nuclear envelope. (wikipedia.org)

eukaryotic

  • Transfection is the process of deliberately introducing naked or purified nucleic acids into eukaryotic cells. (wikipedia.org)
  • The process of introducing foreign DNA into eukaryotic cells is known as transfection. (wikipedia.org)

term transfection

  • Because the term transformation had another sense in animal cell biology (a genetic change allowing long-term propagation in culture, or acquisition of properties typical of cancer cells), the term transfection acquired, for animal cells, its present meaning of a change in cell properties caused by introduction of DNA. (wikipedia.org)
  • The meaning of the term transfection has evolved. (wikipedia.org)
  • Because the term transformation had another sense in animal cell biology (a genetic change allowing long-term propagation in culture, or acquisition of properties typical of cancer cells), the term transfection acquired, for animal cells, its present meaning of a change in cell properties caused by introduction of DNA (or other nucleic acid species such as RNA or SiRNA). (wikipedia.org)

siRNA transfection

  • Optimizing siRNA Transfection for RNAi ( Low transfection. (bio-medicine.org)
  • This article summarizes the use of reverse transfection to maximize performance of siRNA in cultured cells and offers suggestions on how to optimize siRNA transfection parameters. (bio-medicine.org)

reverse transfection

  • Both siPORT NeoFX and siPORT Amine Transfection Agents can be used for reverse transfection or standard transfection of siRNAs into a wide variety of cell types. (bio-medicine.org)
  • Here, we show evidence that an alternative transfection procedure, termed reverse transfection or neofection , offers several key benefits over the traditional pre-plating method. (bio-medicine.org)
  • Reverse transfection involves simultaneously transfecting and plating cells, much like procedures used for transfecting suspension cells. (bio-medicine.org)
  • Reverse transfection is the invention and development of a microarray-driven gene expression system by Junald Ziauddin and David M. Sabatini in 2001. (wikipedia.org)
  • The advantages of reverse transfection (over conventional transfection) are: The addition and attachment of target cells to the DNA-loaded surface can lead to a higher probability of cell-DNA contact, potentially leading to higher transfection efficiency. (wikipedia.org)

transfer of nucleic acids

  • Optical transfection (or any derivations of laser transfection, optotransfection, phototransfection): A specific type of optical transfection - the transfer of nucleic acids into a cell using light for the purposes of eliciting protein translation from those acids. (wikipedia.org)

calcium phosphate

  • Due to its highly consistent pH and salt concentration, our CalPhos calcium phosphate transfection kit gives you the consistent transfection efficiency that you cannot always achieve with home brew versions. (clontech.com)
  • For this reason CalPhos has remained one of the most popular commercially available calcium phosphate transfection kits for over 15 years. (clontech.com)
  • Transfection can be carried out using calcium phosphate (i.e. tricalcium phosphate), by electroporation, by cell squeezing or by mixing a cationic lipid with the material to produce liposomes which fuse with the cell membrane and deposit their cargo inside. (wikipedia.org)

gene

  • Low transfection efficiency and low cell viability are the most frequent causes of unsuccessful gene silencing experiments. (bio-medicine.org)
  • The goal of transfection optimization is to determine the conditions that will provide maximum gene knockdown while maintaining an acceptable level of viability for the particular cell type (see sidebar, Two-Step Optimization Protocol ). (bio-medicine.org)
  • Gene knockdown by transfection of exogenous siRNA is often unsatisfactory because the effect is only transient, especially in rapidly dividing cells. (wikipedia.org)

adherent

  • We would be very interested to hear from anyone who has optimized this RNA transfection protocol, or used any other protocols for non-adherent cells. (bio.net)
  • I know tranfection of adherent lines has been done, but I haven't found too much material on suspension line transfection. (protocol-online.org)
  • As DNA is printed on a glass slide for the transfection process (the deliberate introduction of nucleic acids into cells) to occur before the addition of adherent cells, the order of addition of DNA and adherent cells is reverse that of conventional transfection. (wikipedia.org)
  • In particular, adherent mammalian cell lines and primary cell cultures show very high transfection rates. (wikipedia.org)
  • Adherent cells can also be transfected using electroporation, providing researchers with an alternative to trypsinizing their cells prior to transfection. (wikipedia.org)

electroporation

  • As with DNA, RNA can be delivered to cells by a variety of means including microinjection, electroporation, and lipid-mediated transfection. (wikipedia.org)
  • Electroporation has proven efficient for use on tissues in vivo, for in utero applications as well as in ovo transfection. (wikipedia.org)

introduction of nucleic

  • Because of this strict definition of transfection, optical transfection also refers only to the introduction of nucleic acid species. (wikipedia.org)

Molecular Biology

  • This procedure is highly suited for transient transfection used for various molecular biology studies. (wikipedia.org)

siRNAs

  • To achieve maximum effectiveness of exogenously introduced siRNAs, transfection optimization experiments are required. (bio-medicine.org)
  • siRNAs can also be introduced into cells by transfection. (wikipedia.org)

viral

  • Between 24-48 hours post-transfection, efficiency and viral titer can be determined, or cells can be collected for other applications. (sigmaaldrich.com)
  • Chemical-based transfection can be divided into several kinds: cyclodextrin, polymers, liposomes, or nanoparticles (with or without chemical or viral functionalization. (wikipedia.org)
  • Altogen Biosystems began as a company focusing on non-viral methods of transfection. (wikipedia.org)
  • The term can also be understood as DNA transfection using a viral vector. (wikipedia.org)

molecules

  • Long-RNA transfection is the process of deliberately introducing RNA molecules longer than about 25nt into living cells. (wikipedia.org)
  • A distinction is made between short- and long-RNA transfection because exogenous long RNA molecules elicit an innate immune response in cells that can cause a variety of nonspecific effects including translation block, cell-cycle arrest, and apoptosis. (wikipedia.org)

nucleic acids into cells

  • Optical transfection is the process of introducing nucleic acids into cells using light. (wikipedia.org)

stable

  • While transient transfection is advantageous for fast analysis of shRNA mediated effects, stable transfection ensures long-term, reproducible as well as defined shRNA effects. (openwetware.org)
  • Stable expression can be influenced by two factors: The transfection method used and the vector containing the shRNA of interest. (openwetware.org)
  • The transfection method determines which cell type can be targeted for stable integration through antibiotic selection . (openwetware.org)
  • who used a frequency tripled Nd:YAG to generate stable and transient transfection of normal rat kidney cells. (wikipedia.org)

protocols

plasmids

  • However, with the invention of different types of microarray printing systems, hundreds of transfection mixes (containing different DNA of interest) may be printed on the same slide for cell uptake of plasmids. (wikipedia.org)

Transfecting

  • As a result, while a cell can be repeatedly transfected with short RNA with few non-specific effects, repeatedly transfecting cells with even a small amount of long RNA can cause cell death unless measures are taken to suppress or evade the innate immune system (see "Long-RNA transfection" below). (wikipedia.org)

Invitrogen

  • You can check the transfection manual from Invitrogen /Life technologies website. (protocol-online.org)

Lipofection

  • Lipofection (or liposome transfection) is a technique used to inject genetic material into a cell by means of liposomes, which are vesicles that can easily merge with the cell membrane since they are both made of a phospholipid bilayer. (wikipedia.org)
  • In contrast, methods like lipofection offer only statistical hits between cargo and cells, because of the three-dimensional motion of cells and transfection aggregates in a liquid suspension. (wikipedia.org)
  • Additionally, synergistic effects in transfection efficiency can arise from the possible combination of lipofection and magnet-assisted transfection. (wikipedia.org)

cells

  • GeneX Plus displays high transfection efficiency with low cytotoxicity in suspension cells such as HEK-293T/SF, THP-1, as well as difficult-to-transfect cell lines such as RAW 264.7 and SH-SY5Y. (atcc.org)
  • These transfection parameters include culture conditions, choice and amount of transfection agent, exposure time of transfection agent to cells, and siRNA quantity and quality. (bio-medicine.org)
  • There is no single transfection parameter that by itself ensures efficient siRNA uptake by cells in culture. (bio-medicine.org)
  • ated transfection procedure involves pre-plating cells, meaning the cells are allowed to attach, recover, and grow for 24 hours prior to transfection. (bio-medicine.org)
  • For maximal cell viability during transfection, cells must be healthy at the beginning of the experiment--healthy cells are easier to transfect than poorly maintained cells. (bio-medicine.org)
  • Suitable for transfection of Bone Marrow MSCs (PCS-500-012), Dermal Microvascular Endothelial cells (PCS-110-010), THP-1 cells (TIB-202), Raw 264.7 cells (TIB-71), and SH-SY5Y cells (CRL-2266). (atcc.org)
  • The apparatus may be a catheter arrangement with various embodiments for applying heat to a patient's cells in vivo in order to improve transfection efficiency or application efficiency. (google.com)
  • Plate the cells 18-24 hours before transfection. (sigmaaldrich.com)
  • To prepare cells for transfection, remove the medium in each dish and replace with 5 mL fresh, complete culture medium. (sigmaaldrich.com)
  • cells are seeded 24hrs prior to transfection at a density of approx 1*10^4 cells/ well. (protocol-online.org)
  • For many disease models, the most desirable cell types such as immune system or primary cells are not amenable to transfection. (openwetware.org)
  • In animal cells, transfection is the preferred term as transformation is also used to refer to progression to a cancerous state (carcinogenesis) in these cells. (wikipedia.org)
  • Transfection of animal cells typically involves opening transient pores or "holes" in the cell membrane to allow the uptake of material. (wikipedia.org)
  • Transfection can result in unexpected morphologies and abnormalities in target cells. (wikipedia.org)
  • The original meaning of transfection was "infection by transformation", i.e., introduction of genetic material, DNA or RNA, from a prokaryote-infecting virus or bacteriophage into cells, resulting in an infection. (wikipedia.org)
  • Magnet-assisted transfection is a transfection method which uses magnetic interactions to deliver DNA into target cells. (wikipedia.org)

simplify

  • Ambion provides a series of tools to simplify RNAi experiments including two transfection agents designed specifically for siRNA delivery. (bio-medicine.org)
  • Simplify your transfections. (clontech.com)

presence of serum

  • Magnet-assisted transfection can also be performed in the presence of serum, which is a further benefit. (wikipedia.org)

ensures

  • This feature ensures reproducibility of transfection-complex formation. (wikipedia.org)

formulations

  • These innovations also served as the basis for the company's transfection formulations and nucleic acid labeling and conjugation chemistries. (wikipedia.org)

RNAi

  • Failure to optimize critical transfection parameters can render RNAi effects undetectable in cell culture. (bio-medicine.org)
  • Short-RNA transfection is routinely used in biological research to knock down the expression of a protein of interest (using siRNA) or to express or block the activity of a miRNA (using short RNA that acts independently of the cell's RNAi machinery, and therefore is not referred to as siRNA). (wikipedia.org)

infection

  • The word transfection is a blend of trans- and infection. (wikipedia.org)

method

  • construct using the calsium phosphate method of transfection. (bio.net)
  • Magnet-assisted transfection is a relatively new and time-saving method to introduce nucleic acids into a target cell with increased efficiency. (wikipedia.org)

protein

  • Inhibiting only three proteins, interferon-β, STAT2, and EIF2AK2 is sufficient to rescue human fibroblasts from the cell death caused by frequent transfection with long, protein-encoding RNA. (wikipedia.org)

Mechanism

  • H. diminuta has an effective mechanism for interspecies transfection. (wikipedia.org)

dish

  • Transfect up to 700 wells of a 6 well plate (or 100 transfections on a 10 cm dish). (clontech.com)

cell types

  • Ambion's siRNA Delivery Resource has further details on transfection optimization, recommendations for transfection agent, and conditions to use with specific cell types. (bio-medicine.org)
  • Since this time, the optical transfection of a host of mammalian cell types has been demonstrated using a variety of laser sources, including the 405 nm continuous wave (cw), 488 nm cw, or pulsed sources such as the 800 nm femtosecond pulsed Ti:Sapphire or 1064 nm nanosecod pulsed Nd:YAG. (wikipedia.org)

density

  • An appropriate seeding density should be used so the cell culture plate is 90-95% confluent at the time of transfection. (sigmaaldrich.com)

biological

  • To be in line with the current definition of transfection in the biological community, non-nucleic acids (such as fluorophores) cannot, by definition, be optically transfected (only optically injected). (wikipedia.org)