Plasmid-based one-pot saturation mutagenesis.

Wrenbeck, Emily E; Klesmith, Justin R; Stapleton, James A; Adeniran, Adebola; Tyo, Keith E J; Whitehead, Timothy A · Nat Methods · 2016

basic_science · Level V

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Abstract

Deep mutational scanning is a foundational tool for addressing the functional consequences of large numbers of mutants, but a more efficient and accessible method for construction of user-defined mutagenesis libraries is needed. Here we present nicking mutagenesis, a robust, single-day, one-pot saturation mutagenesis method performed on routinely prepped plasmid dsDNA. The method can be used to produce comprehensive or single- or multi-site saturation mutagenesis libraries.

Medical subject headings