Multiple rounds of mutagenesis are possible without intermediate subcloning steps. Introduction Site-directed mutagenesis is a key technology for studying enzyme activity and molecular recognition. The availability and ease of synthesis of oligonucleotides permits widespread and intensive use of mutagenesis techniques for studying a variety of problems. Promega's Altered Sites II Mutagenesis Systems provide rapid, efficient, and flexible alternatives for site-directed mutagenesis and protein expression. Initial oligonucleotide hybridization techniques used single-stranded template DNA, usually derived from M13 phagemid vectors, which was hybridized to the mutagenic oligonucleotide.
New protocol allows use of double-stranded DNA template.
Selection for the mutant strand allows multiple oligonucleotides to be incorporated in a single round of mutagenesis. Four independent mutations have been incorporated in a single round of mutagenesis. The mutagenesis procedure from double-stranded DNA through mutant selection can be performed in a single day. Ex 2, for coexpression with other plasmids. Each vector contains two selectable antibiotic resistances which can be used alternatively to provide mutant strand selection, allowing multiple rounds of mutagenesis to be performed without subcloning of the insert.