Scramble of heterozygous diploid yeast

Abstract

Provided are compositions and methods for making a diploid yeast having heterozygosity for at least one chromosome. The method includes mating haploid yeast having at least a first modified chromosome comprising a synthetic chromosome suitable for recombination-site-mediated evolution (as a SCRaMbLE-ready modification). The SCRaMbLE-ready modification includes introduced site-specific recombinase recognition sites that can be recognized by a recombinase. Yeast that have at least one SCRaMbLE-ready modification of a chromosome are mated with a haploid yeast devoid of the SCRaMbLE-ready modification to obtain diploid SCRaMbLE-ready yeast. Subsequent to mating the haploid SCRaMbLE-ready yeast to yeast devoid of the SCRaMbLE-ready modification the method includes using the recombinase to recombine the first modified chromosome to obtain heterozygous diploid yeast comprising at least one recombined (SCRaMbLEd) chromosome and a homologous non-SCRaMbLEd chromosome. The method further includes identifying heterozygous diploid yeast that comprise the at least one SCRaMbLEd chromosome that have a changed phenotype that is different from the phenotype of the diploid SCRaMbLE-ready yeast. Yeast made by the methods, compositions of matter made by the yeast, spores made by the yeast, and kits containing the yeast and/or their spores are included.