Large-scale deletion of non-essential genes of Saccharomyces cerevisiae

Saccharomyces cerevisiae is an important model eukaryotic organism with a known complete genome sequence and only 17.4% of genes are essential for cell viability. An attempt to create a cell with a minimal genome, the non-essential genes pinpointed by bioinformatics tools and then were deleted. Here, we report the large-scale deletion of non-essential genes of S. cerevisiae. The report is divided into three parts. Firstly, the relevant research progress of synthetic biology and minimal genome are introduced, and the principles of gene knockout and technical route of Cre/LoxP, Latour, MIRAGE, TALEN and CRISPR/Cas9 system are reviewed. Secondly, the present results of this work in our laboratory are described. The S. cerevisiae strain JM (matα, Dura3) was used in this study. Latour system was applied for ≥10 kb large scale deletion and eight deletants with 8 kb, 11 kb, 17 kb, 18 kb, 23 kb, 24 kb, 38 kb, 40 kb fragments excision respectively were genarated. Improved MIRAGE system was applied for ≤10 kb DNA fregment deletion and two deletants with 4 kb, 5 kb fragments deletion separately were obtained. Superposition of DNA fragments deletion were performed using both chromosome XⅢ and XV as the target on this basis. We obtained a deletant with five DNA fregment, 124 kb, excision and a deletant with four fragments, 133 kb, excision. Finaly, work arrangements are described. More non-essential genes will be knockout and we will explore the feasibility of using the CRISPR-Cas9 system to perform large-scale deletions.