Gene Transfer and Expression Protocols

Preparation of Recombinant Plasmid DNA for DNA-Mediated Gene Transfer.- Calcium Phosphate Mediated Gene Transfer into Established Cell Lines.- Transfection of the ChloramphenicolAcetyltransferase Gene into Eukaryotic Cells Using Diethyl-Aminoethyl (DEAE)-Dextran.- Polybrene/DMSOAssisted Gene Transfer.- Electroporation Technique of DNA Transfection.- Irradiation and Fusion Gene Transfer.- Direct Use of ? Phage Particles for DNA Transfection.- Cationic Liposome-Mediated Transfection with Lipofectin(TM) Reagent.- Preparation of High-Molecular-Weight DNA for Use in DNA Transfection.- Chromosome-Mediated Gene Thnsfer.- Manipulation of Adenovirus Vectors.- Manipulation of Vaccinia Virus Vectors.- Manipulation of Baculovirus Vectors.- Manipulation of SV40 Vectors.- Choice and Manipulation of Retroviral Vectors.- Use of Tissue-Plasminogen Activator as a Reporter Gene.- Use of Escherichiu coli (E. coli) lacZ (?-Galactosidase) as a Reporter Gene.- Application of the Firefly Luciferase Reporter Gene.- Use of Vectors to Confer Resistance to Antibiotics G418 and Hygromycin in Stably Transfected Cell Lines.- Selection of Cells Defective in Pyrimidine (TK?) and Purine (APRT? and HPRT? Salvage.- Quantitative and Qualitative Analysis of Exogenous Gene Expression by the S1 Nuclease ProtectionAssay.- The RNase Protection Assay.- Primer Extension Analysis of mRNA Isolated from Transfected Cell Lines.- Northern Blot Analysis of Gene Expression.- Analysis of Transcriptional Initiation in Isolated Nuclei.- Immunoperoxidase Staining of Gene Products in Cultured Cells Using Monoclonal Antibodies.- The Use of Flow Cytometry to Detect Transfected Gene Products.- Determination of Foreign Gene Copy Number in Stably Transfected Cell Lines by Southern Transfer Analysis.- Evaluation of Extrachromosomal Gene Copy Number of Transiently Transfected Cell Lines.- Use of Dpn I Restriction Enzyme to Assess Newly Replicated Gene Copies in Amplifiable Vector Systems.- Use of Polymerase Chain Reaction (PCR) to Detect Homologous Recombination in Transfected Cell Lines.- Induction of Erythroid-Specific Expression in Murine Erythroleukemia (MEL) Cell Lines.