Rearrangement Of Synthetic Chromosome V In Polyploid Yeast

Yeast polyploidization and aneuploidization belong to a wide range of genomic structural variations and are all important genetic variants of industrial yeast strains,of which yeast mating type switching is a very critical step.In this study,a yeast mating type switching method based on CRISPR/Cas9 was developed.Through one yeast transformation,mating type switching of haploid and diploid yeast cells can be completed,and the efficiency of mating type switching can reach more than 80%.Using a CRISPR/Cas9-mediated mating type switching method,a haploid yeast containing two synthetic chromosomes(synV and synX)was constructed and the construction of homologous wild-type triploid,tetraploid,and pentaploid yeast strains was achieved.In order to verify the effect of polyploid genome rearrangement,we integrated the zeaxanthin biosynthesis module on the wild type V chromosome to construct zeaxanthin-producing wild-type diploid,triploid and tetraploid yeasts.As well as hybrid diploid,triploid and tetraploid yeast strains containing synthetic chromosomes.In this study,rearrangement of synV chromosomes was induced in synthetic polyploid yeasts,and the yield of zeaxanthin was used to screen for the beneficial structural variation.We found that the number of strains with increased yield of zeaxanthin in the triploid yeast strain containing synV was the largest and the ratio could be up to 15%,and the maximum increase in zeaxanthin production reached 3.80 times.Analysis of the genotypes of synV rearranged yeast with elevated zeaxanthin yields revealed that a single chromosomal rearrangement would yield a yeast library of diverse genotypes,including aneuploid yeast.This study screened 3N-2 aneuploid yeast.An aneuploid yeast of 3N-2 was subjected to sporulation and tetrad dissection to obtain a 0.81-fold increase in zeaxanthin yield of 1.5 aneuploid yeast.This study can provide some theoretical guidance for the construction of excellent industrial strains.