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Nucleic Acids Research, 1983, Vol. 11, No. 12 4229-4239
© 1983

MOLECULAR BIOLOGY

A general method to select for M13 clones carrying base pair substitution mutants constructed in vitro

Cinzia Traboni, Riccardo Cortese*, Gennaro Ciliberto and Gianni Cesareni

European Molecular Biology Laboratory Postfach 10.2209, 6900 Heidelberg, FRG

*to whom correspondence should be addressed.

Received March 28, 1983. Accepted May 10, 1983.

In this paper we describe a method to select base pair substitution mutants constructed in vitro. The mutagenesis is performed by forcing mistakes during in vitro synthesis from a primer annealed to a single stranded DNA template. The selection is based on the fact that, following transformation, the progeny of the strand elongated in vitro and the template strand have different phenotypes. The method is general and applicable to any DNA segment; the type of base pair substitution and its position can be chosen at will. The combined efficiency of mutagenesis and selection allows for 85% frequency of mutants in all analyzed clones.


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