Solid phasein vitromutagenesis using plasmid DNA template
Open Access
- 1 January 1990
- journal article
- research article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 18 (17) , 5107-5112
- https://doi.org/10.1093/nar/18.17.5107
Abstract
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.This publication has 22 references indexed in Scilit:
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