Whole-Genome Profiling of a Novel Mutagenesis Technique Using Proofreading-Deficient DNA Polymerase ?.

ABSTRACT: A novel mutagenesis technique using error-prone DNA polymerase ? (pol?), the disparity mutagenesis model of evolution, has been successfully employed to generate novel microorganism strains with desired traits. However, little else is known about the spectra of mutagenic effects caused by disparity mutagenesis. We evaluated and compared the performance of the pol?MKII mutator, which expresses the proofreading-deficient and low-fidelity pol?, in Saccharomyces cerevisiae haploid strain with that of the commonly used chemical mutagen ethyl methanesulfonate (EMS). This mutator strain possesses exogenous mutant pol? supplied from a plasmid, tthereby leaving the genomic one intact. We measured the mutation rate achieved by each mutagen and performed high-throughput next generation sequencing to analyze the genome-wide mutation spectra produced by the 2 mutagenesis methods. The mutation frequency of the mutator was approximately 7 times higher than that of EMS. Our analysis confirmed the strong G/C to A/T transition bias of EMS, whereas we found that the mutator mainly produces transversions, giving rise to more diverse amino acid substitution patterns. Our present study demonstrated that the pol?MKII mutator is a useful and efficient method for rapid strain improvement based on in vivo mutagenesis.

SUBMITTER: Shiwa Y 

PROVIDER: S-EPMC3364565 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature