Whole-Genome Sequencing of Yeast Cells.

Abstract

The budding yeast, Saccharomyces cerevisiae, has been widely used for genetic studies of fundamental cellular functions. The isolation and analysis of yeast mutants is a commonly used and powerful technique to identify the genes that are involved in a process of interest. Furthermore, natural genetic variation among wild yeast strains has been studied for analysis of polygenic traits by quantitative trait loci mapping. Whole-genome sequencing, often combined with bulk segregant analysis, is a powerful technique that helps determine the identity of mutations causing a phenotype. Here, we describe protocols for the construction of libraries for S. cerevisiae whole-genome sequencing. We also present a bioinformatic pipeline to determine the genetic variants in a yeast strain using whole-genome sequencing data. This pipeline can also be used for analyzing Schizosaccharomyces pombe mutants. © 2019 by John Wiley & Sons, Inc. Basic Protocol 1: Generation of haploid spores for bulk segregant analysis Basic Protocol 2: Extraction of genomic DNA from yeast cells Basic Protocol 3: Shearing of genomic DNA for library preparation Basic Protocol 4: Construction and amplification of DNA libraries Support Protocol 1: Annealing oligonucleotides for forming Y-adapters Support Protocol 2: Size selection and cleanup using SPRI beads Basic Protocol 5: Identification of genomic variants from sequencing data.