Protocol provides a quick way to assay for RNA-dependent RNA polymerase (RdRP) activity in crude protein samples. RdRP transcription products remain at the origin during paper chromatography. - [Read An Assay for RNA-Dependent RNA Polymerase Activity Protocol]
Cultivating animal cells in the laboratory is an indispensable technique for cell biologists. However, most normal primary cell lines, while faithfully reproducing the phenotype of their tissue of origin, do not grow indefinitely in culture. After a series of population doublings (the number of which varies by species, cell type, and culture conditions) primary cells enter a state where they no longer divide. - [Read Immortalization of Cells in Culture]
Transient transfection into 293T cells is a convenient way to overexpress and obtain both cellular and extracellular (secreted or membrane) proteins. 293 is a human renal epithelial cell line which is transformed by adenovirus E1A gene product. 293T is a derivative which also express SV40 large T antigen, allowing episomal replication of plasmids containing the SV40 origin and early promoter region. They (both) have the unusual property of being highly transfectable. - [Read Transient Transfection Into 293T Cells Protocol]
This protocol describes the electroporation of the BMH 81-17 mut S strain that is recommended for tranformation of the site directed mutagenesis of dsDNA (See Protocol on Site-Directed Mutagenesis on Double Stranded DNA). BMH 81-17 mut S are a mismatch repair defective (mut S) Escherichia coli strain. The probability that the two mutations will cosegregate during the first round of DNA replication is increased in this strain.