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Nature Research, Nature Methods, 3(12), p. 203-206, 2015

DOI: 10.1038/nmeth.3223

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Massively Parallel Single Amino Acid Mutagenesis

Journal article published in 2015 by Jacob O. Kitzman, Lea M. Starita, Russell S. Lo, Stanley Fields, Jay Shendure ORCID
This paper is made freely available by the publisher.
This paper is made freely available by the publisher.

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Abstract

Random mutagenesis methods only partially cover the mutational space and are constrained by DNA synthesis length limitations. Here we demonstrate programmed allelic series (PALS), a single-volume, site-directed mutagenesis approach using microarray-programmed oligonucleotides. We created libraries including nearly every missense mutation as singleton events for the yeast transcription factor Gal4 (99.9% coverage) and human tumor suppressor p53 (93.5%). PALS-based comprehensive missense mutational scans may aid structure-function studies, protein engineering, and the interpretation of variants identified by clinical sequencing.