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Methods Mol Biol. 2010;634:3-14. doi: 10.1007/978-1-60761-652-8_1.

Mutagenesis protocols in Saccharomyces cerevisiae by in vivo overlap extension.

Author information

1
Instituto de Catálisis y Petroleoquímica (CSIC), Madrid, Spain. malcalde@icp.csic.es

Abstract

A high recombination frequency and its ease of manipulation has made Saccharomyces cerevisiae a unique model eukaryotic organism to study homologous recombination. Indeed, the well-developed recombination machinery in S. cerevisiae facilitates the construction of mutant libraries for directed evolution experiments. In this context, in vivo overlap extension (IVOE) is a particularly attractive protocol that takes advantage of the eukaryotic apparatus to carry out combinatorial saturation mutagenesis, site-directed recombination or site-directed mutagenesis, avoiding ligation steps and additional PCR reactions that are common to standard in vitro protocols.

PMID:
20676972
DOI:
10.1007/978-1-60761-652-8_1
[Indexed for MEDLINE]

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