Massively parallel single-nucleotide mutagenesis using reversibly terminated inosine

作者：Haller Gabe; Alvarado David; McCall Kevin; Mitra Robi D; Dobbs Matthew B; Gurnett Christina A^*

来源：Nature Methods, 2016, 13(11): 923-+.

DOI：10.1038/NMTH.4015

Large-scale mutagenesis of target DNA sequences allows researchers to comprehensively assess the effects of single-nucleotide changes. Here we demonstrate the construction of a systematic allelic series (SAS) using massively parallel single-nucleotide mutagenesis with reversibly terminated deoxyinosine triphosphates (rtITP). We created a mutational library containing every possible single-nucleotide mutation surrounding the active site of the TEM-1 beta-lactamase gene. When combined with high-throughput functional assays, SAS mutational libraries can expedite the functional assessment of genetic variation.

• 出版日期2016-11