Here we provide methods for setting up standard crosses with Schizosaccharomyces pombe strains. All strain genotypes and pedigrees should be recorded in a laboratory strain book. Matings between two haploid strains of interest are induced on solid medium poor in nitrogen. Usually, sporulation agar (SPA) plates are preferred, but for difficult matings it is advisable to try several mating media in parallel because one medium might allow for more efficient mating. Protoplast fusion can be used to produce zygotes from sterile mutants that fail to mate.