Site-specific mutagenesis was accomplished using a solid support to generate single stranded vector and insert fragments which can be used to form gap-duplex plasmids through flanking, complementary double stranded regions. More than 80% mutants were obtained in both a single and a double primer approach. No special vectors or strains are needed and mismatch repair is avoided as the mutagenesis region is in a single stranded form when transformed into the Escherichia coil host cell. The fragments to be immobilized can be produced either by a polymerase chain reaction using general primers or by a site-specific restriction followed by a fill-in reaction. This novel method is rapid, simple and flexible and well suited for both manual and semi-automated in vitro mutagenesis protocols. © 1990 Oxford University Press.
CITATION STYLE
Hultman, T., Murby, M., Stähl, S., Hornes, E., & Uhlén, M. (1990). Solid phase in vitro mutagenesis using plasmid DNA template. Nucleic Acids Research, 18(17), 5107–5112. https://doi.org/10.1093/nar/18.17.5107
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