Efficient method for site-directed mutagenesis of plasmids and cloning of single-stranded DNA fragments

V. L. Drutsa, V. R. Kaberdin, O. N. Koroleva, I. A. Shilov

A. N. Belozersky Laboratory of Molecular Biology and Bioorganic Chemistry and Chemical Department, M. V. Lomonosov Moscow State University, Moscow

Abstract: A three primer variant of the earlier devised oligonucleotide-directed mutagenesis in plasmids is described, useful also for the fast cloning of single-stranded DNA products of the asymmetric polymerase chain reaction (PCR). Using this method for plasmid pHD-001-14-11, a 59 b. p. deletion and a 7 b. p. insertion were simultaneously introduced at 81% frequency, and the PCR-copied phage fd transcription terminator (26 b. p.) was inserted with the yield of 67%.

Russian Journal of Bioorganic Chemistry 1991, 17 (11):1487-1493

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