Site-directed mutagenesis of whole viral genomes.
This chapter introduces an efficient and accurate site-directed mutagenesis protocol, which allows the color selection of mutants through the simultaneous activation or deactivation of the alpha-peptide of beta-galactosidase. It uses double-stranded plasmid DNA as the mutational template. This protocol can efficiently create mutations of large inserts at multiple sites simultaneously and can be used to perform multiple rounds of mutation on the same construct. Thus, constructs containing whole open-reading frames and whole viral genomes can be subjected to site-directed mutagenesis and used for subsequent functional studies.
Liu
Li
L
Centre for Infectious Disease, Institute of Cell and Molecular Science, Barts and The London, Queen Mary's School of Medicine and Dentistry, The Blizard Building, 4 Newark Street, Whitechapel, London E1 2AT, UK.
Lomonossoff
George P
GP
D001483:Base Sequence; D017931:DNA Primers; D004279:DNA, Viral / Q000235:genetics; D016679:Genome, Viral; D016297:Mutagenesis, Site-Directed / Q000379:methods*; D009691:Nucleic Acid Denaturation; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D010766:Phosphorylation; D015183:Restriction Mapping / Q000379:methods; D013698:Templates, Genetic
D017931:DNA Primers; D004279:DNA, Viral; D009838:Oligodeoxyribonucleotides