Abstract
AbstractClustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR‐associated systems (CRISPR/Cas) are revolutionary tools for predictable and repeatable genome editing. In this article, the CRISPR/Cas9 system will be used to engineer the genome of the yeast Saccharomyces cerevisiae. As a model organism utilized across biological disciplines, efficient and tailorable methods for engineering the yeast genome are indispensable for a variety of research, educational, and commercial applications. The protocols described here incorporate a simple restriction‐free cloning strategy and digestion‐based screening method that can be readily and easily modified to suit user‐designed experimental needs. © 2020 Wiley Periodicals LLC.Basic Protocol 1: Recombinant pCAS plasmid preparationBasic Protocol 2: Barcode/editing fragment assemblyBasic Protocol 3: Gene editing by yeast co‐transformationAlternate Protocol: Competent yeast preparation and transformation by a lithium acetate/single‐stranded carrier method
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