Mutagenesis Protocols in Saccharomyces cerevisiae by In Vivo Overlap Extension
Mutagenesis Protocols in Saccharomyces cerevisiae by In Vivo Overlap Extension
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DOI：
10.1007/978-1-60761-652-8_1
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发表时间：
2010-01-01
期刊：
影响因子：
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通讯作者：
Alcalde, Miguel
中科院分区：
文献类型：
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作者：
Alcalde, Miguel
高重组频率和易操纵性使酿酒酵母成为研究同源重组的唯一模式真核生物。事实上，酿酒酵母中发达的重组机制有助于构建用于定向进化实验的突变文库。在这种背景下，体内重叠延伸(IVOE)是一种特别有吸引力的协议，它利用真核细胞进行组合饱和突变、定点重组或定点突变，避免了标准体外协议常见的连接步骤和额外的PCR反应。
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.