[Protocol] Molecular genetic tools and techniques in fission yeast

The molecular genetic tools used in fission yeast have generally been adapted from methods and approaches developed for use in the budding yeast, Saccharomyces cerevisiae. Initially, the molecular genetics of Schizosaccharomyces pombe was developed to aid gene identification, but it is now applied extensively to the analysis of gene function and the manipulation of noncoding sequences that affect chromosome dynamics. Much current research using fission yeast thus relies on the basic processes of introducing DNA into the organism and the extraction of DNA for subsequent analysis. Targeted integration into specific genomic loci is often used to create site-specific mutants or changes to noncoding regulatory elements for subsequent phenotypic analysis. It is also regularly used to introduce additional sequences that generate tagged proteins or to create strains in which the levels of wild-type protein can be manipulated through transcriptional regulation and/or protein degradation. Here, we draw together a collection of core molecular genetic techniques that underpin much of modern research using S. pombe. We summarize the most useful methods that are routinely used and provide guidance, learned from experience, for the successful application of these methods.