基因工程与基因组重排技术构建酿酒酵母高效木糖发酵菌株及甘油发酵菌株-生物化学与分子生物学专业论文.docxVIP

transformed to be a diploid ste2/ste2 strain ( the STE2 gene encodesα-factor receptor ), then subjected to EMS mutagenesis followed by meiotic recombination- mediated genome shuffling and evolutionary engineering. We obtained a strain with enhanced xylose-fermenting ablity named WCY1 which can consume 100 g/l xylose in 48 h with an ethanol yield of 0.395 g/l, and the maximum specific growth rate was 0.294 h-1.These figures exceed by a significant margin any other performance metrics on xylose utilization by S. cerevisiae reported to-date.. The fermentation in medium with mixed sugar of WCY1 showed that xylose- utilization was inhibited. Measurement of the xylose-utilizing enzymes activities showed no significant change suggesting a new genetic trait in the strain had been generated in favor of xylose metabolism. Finally, we tested the mating type of evolved strains. Result meets the expectation of the designed experiment which verified that genome shuffling is an efficient strategy to improve xylose fermenting property of Saccharomyces cerevisiae strain. Key words：Saccharomyces cerevisiae; glycerol; xylose; ethanol; DAK1; DAK2; GCY1; PHO13; xylitol; meiotic recombination; genome shuffling; fermentation 目 录 中文摘要 1 ABSTRACT 2 HYPERLINK \l _bookmark0 目 录 1 HYPERLINK \l _bookmark1 第一章 文献综述 1 HYPERLINK \l _bookmark2 1.1 生物乙醇发展现状 1 HYPERLINK \l _bookmark3 1.2 甘油的生物转化 2 HYPERLINK \l _bookmark4 1.2.1 微生物对甘油的利用2 HYPERLINK \l _bookmark5 1.2.2 以甘油为底物生产乙醇3 HYPERLINK \l _bookmark6 1.3 木糖-乙醇的生物转化5 HYPERLINK \l _bookmark7 1.4 酿酒酵母利用木糖菌株构建 6 HYPERLINK \l _bookmark8 1.4.1 代谢工程方法7 HYPERLINK \l _bookmark9 1.4.2 进化工程方法8 HYPERLINK \l _bookmark10 1.4.3 基因组重排与有性重组9 HYPERLINK \l _bookmark11 1.5 本文的研究背景、内容及目的 11 HYPERLINK \l _bookmark12 第二章 实验材料及方法 12 HYPERLINK \l _bookmark13 2.1 实验材料 12 HYPERLINK \l _bookmark14 2.1.1 菌株、质粒与引物12 HYPERLINK \l _bookmark15 2.1.2 实验仪器16 HYPERLINK \l _bookmark16 2.1.3 主要试剂17 HYPERLINK \l _bookmark17 2.1.4 培养基18 HYPERLINK \l _bo