Abstract
Random mutagenesis methods only partially cover the mutational space, and are constrained by DNA synthesis length limitations. Here, we demonstrate 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.
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