Parallel profiling of mutants for chronological lifespan

战略研究重点：了解参与细胞衰老的主要参与者是揭示促进健康老龄化所需机制的关键。裂殖酵母和芽殖酵母是远亲生物，它们都是用于老化研究的特殊模式生物。因此，这两种酵母为衰老研究提供了互补的见解。在这个项目中，我们采用条形码测序的S。粟酒裂殖酵母的非必需基因的全基因组缺失文库，占基因组的80%以上，以帮助发现在不同环境条件下具有长寿命和短寿命的突变体。我们期待这些突变体进一步用于药物筛选实验。条形码是横向跨越KanMX4盒的独特的20个核苷酸序列，其插入每个缺失基因的区域中，因此，条形码在时间顺序测定期间充当直接定量突变体的分子标记。通过PCR捕获突变体计数，并在从DNA文库外推的测序数据中突出显示。指导该项目的方法包括但不限于条形码测序技术的设置和优化、DNA文库制备、用于细胞成像的荧光显微镜检查沿着FACS分析。

Strategic Research Priority: World Class Underpinning BioscienceUnderstanding the main players involved in cellular senescence is key to uncovering the mechanisms required for the promotion of healthy ageing. Fission yeast and budding yeast are distantly related organisms, both of which are exceptional model organisms used for ageing studies. The two yeasts therefore offer complementary insights into the ageing research. In this project, we employ barcode sequencing to S. pombe genome-wide deletion library of nonessential genes, accounting for more than 80% of the genome, in order to aid the discovery of mutants with long and short lifespans under different environmental conditions. We anticipate further utilization of these mutants for drug screen experiments. The barcodes are a unique 20-nucleotide sequence that laterally spans the KanMX4 cassette, which is inserted in the region of each of the deleted gene, and thus, the barcodes act as molecular marks with direct quantitation of the mutants during chronological assays. The mutant counts is captured by PCR and highlighted in the sequencing data extrapolated from DNA libraries. Methods that direct the project include, but not limited to, the setting and optimization of the barcode sequencing technique, DNA library preparation, fluorescence microscopy for cellular imaging along with FACS analysis.