Bacterial Artificial Chromosome Mutagenesis Using Recombineering
Figure 1
General applications of recombineering in BAC modifications. Recombineering is applicable in various mutagenesis strategies, depending on the design and nature of the targeting substrate and target site; see text for more details. Stippled boxes denote homologous sequences for recombination. (a) Gene replacement. Recombineering can be employed to replace a target site with any sequence of interest. (b) Insertion. DNA can also be introduced by recombineering without removing any of the existing sequence. (c) Selection/counterselection. Recombineering can mediate subtle modifications such as nucleotide substitutions via two rounds of recombinations, by first introducing a selectable cassette followed by replacement of the cassette with the modified version of the target site. (d) Gap repair cloning. A target site of interest can be cloned from a fragment or plasmid into a linearised vector in vivo by recombineering, through recombination between the ends of the cloning vector and target site.
