Large-scale genome reorganization in Saccharomyces cerevisiae through combinatorial loss of mini-chromosomes

• Published in: Journal of bioscience and bioengineering Vol. 113; no. 6; pp. 675 - 682
• Main Authors: Ueda, Youji, Ikushima, Shigehito, Sugiyama, Minetaka, Matoba, Ryo, Kaneko, Yoshinobu, Matsubara, Kenichi, Harashima, Satoshi
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.06.2012; Elsevier
• Subjects: Bioengineering; Biological and medical sciences; Biotechnology; Cell culture; Chromosome Deletion; chromosomes; Chromosomes, Fungal - genetics; DNA microarray; essential genes; Fundamental and applied biological sciences. Psychology; gene dosage; Genome, Fungal; Haploidy; Large scale genome reorganization; Mini chromosome; mitochondrial genome; PCR-mediated chromosome splitting; Polymerase Chain Reaction - methods; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - ultrastructure; yeasts; DNA microarray; PCR-mediated chromosome splitting; Saccharomyces cerevisiae; Large scale genome reorganization; Mini chromosome; Ascomycota; Yeast; DNA chip; Chromosome; Fungi; Polymerase chain reaction; Large scale; Genome
• ISSN: 1389-1723; 1347-4421
• DOI: 10.1016/j.jbiosc.2012.01.013

A highly efficient technique, termed PCR-mediated chromosome splitting (PCS), was used to create cells containing a variety of genomic constitutions in a haploid strain of Saccharomyces cerevisiae. Using PCS, we constructed two haploid strains, ZN92 and SH6484, that carry multiple mini-chromosomes. In strain ZN92, chromosomes IV and XI were split into 16 derivative chromosomes, seven of which had no known essential genes. Strain SH6484 was constructed to have 14 mini-chromosomes carrying only non-essential genes by splitting chromosomes I, II, III, VIII, XI, XIII, XIV, XV, and XVI. Both strains were cultured under defined nutrient conditions and analyzed for combinatorial loss of mini-chromosomes. During culture, cells with various combinations of mini-chromosomes arose, indicating that genomic reorganization could be achieved by splitting chromosomes to generate mini-chromosomes followed by their combinatorial loss. We found that although non-essential mini-chromosomes were lost in various combinations in ZN92, one mini-chromosome (18kb) that harbored 12 genes was not lost. This finding suggests that the loss of some combination of these 12 non-essential genes might result in synthetic lethality. We also found examples of genome-wide amplifications induced by mini-chromosome loss. In SH6484, the mitochondrial genome, as well as the copy number of genomic regions not contained in the mini-chromosomes, was specifically amplified. We conclude that PCS allows for genomic reorganization, in terms of both combinations of mini-chromosomes and gene dosage, and we suggest that PCS could be useful for the efficient production of desired compounds by generating yeast strains with optimized genomic constitutions.