Massively Parallel Single Amino Acid Mutagenesis

Kitzman, Jacob O.; Starita, Lea M.; Lo, Russell S.; Fields, Stanley; Shendure, Jay

doi:10.1038/nmeth.3223

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Massively Parallel Single Amino Acid Mutagenesis

3 2015
By Kitzman, Jacob O. ; Starita, Lea M. ; Lo, Russell S. ; ...
http://dx.doi.org/10.1038/nmeth.3223
Source: Nat Methods. 12(3):203-206

[PDF-299.10 KB]

English

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Details:

Alternative Title:

Nat Methods
Personal Author:

Kitzman, Jacob O. ; Starita, Lea M. ; Lo, Russell S. ; Fields, Stanley ; Shendure, Jay

Kitzman, Jacob O. ; Starita, Lea M. ; Lo, Russell S. ; Fields, Stanley ; Shendure, Jay Less -
Description:

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.
Keywords:

[+]

Amino Acids DNA-Binding Proteins Gene Library Haplotypes Humans Mutagenesis, Site-Directed Mutation, Missense Oligonucleotide Array Sequence Analysis Saccharomyces Cerevisiae Proteins Transcription Factors Tumor Suppressor Protein P53 Uracil-DNA Glycosidase
Source:

Nat Methods. 12(3):203-206
Pubmed ID:

25559584
Pubmed Central ID:

PMC4344410
Document Type:

Journal Article
Funding:

P41 GM103533/GM/NIGMS NIH HHSUnited States/ ; DP1HG007811/DP/NCCDPHP CDC HHSUnited States/ ; P41GM103533/GM/NIGMS NIH HHSUnited States/ ; Howard Hughes Medical InstituteUnited States/ ; DP1 HG007811/HG/NHGRI NIH HHSUnited States/

P41 GM103533/GM/NIGMS NIH HHSUnited States/ ; DP1HG007811/DP/NCCDPHP CDC HHSUnited States/ ; P41GM103533/GM/NIGMS NIH HHSUnited States/ ; Howard Hughes Medical InstituteUnited States/ ; DP1 HG007811/HG/NHGRI NIH HHSUnited States/ Less -
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CDC Public Access
Main Document Checksum:

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urn:sha256:e670878282a4d0a64da5a868adcba15e2662681790fc895fd33ac9cd1370ad8e
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https://stacks.cdc.gov/view/cdc/30311/cdc_30311_DS1.pdf
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