Abstract
In vitro protein evolution is an efficient approach to study the structure and function of a protein, or to enhance its industrial utility (1). One round of evolution consists of random mutation of a protein-coding gene, expression the resulting library in a population of micro-organisms, and high-throughput screening or selection of clones that most strongly exhibit a desired phenotype (“winners”). After many rounds, mutations that confer the phenotype accumulate on a single allele, e.g., the authors have isolated an octuple mutant of the Escherichia coli β-glucuronidase with catalytic activity resistant to roughly 80-fold higher concentrations of glutaraldehyde than that of the wild-type enzyme (2). Here we describe a variation of the mutagenic polymerase chain reaction (PCR) (3,4) is described. The advantages of this method over other random mutagenesis techniques are explained.
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Matsumura, I., Ellington, A.D. (2002). Mutagenic Polymerase Chain Reaction of Protein-Coding Genes for In Vitro Evolution. In: Braman, J. (eds) In Vitro Mutagenesis Protocols. Methods in Molecular Biology™, vol 182. Humana Press, Totowa, NJ. https://doi.org/10.1385/1-59259-194-9:259
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DOI: https://doi.org/10.1385/1-59259-194-9:259
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-0-89603-910-0
Online ISBN: 978-1-59259-194-7
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