酿酒酵母抗乙醛胁迫关键基因识别及功能分析

It is known that acetaldehyde is toxic to yeast strains during alcoholic fermentation. It blocks cell growth and interferes with a wide range of metabolic activities. Identification genes responsible for acetaldehyde tolerance in the yeast Saccharomyces cerevisiae has great practical relevance. In this proposal, global transcription machinery engineering (gTME) and direct evolution methods will be applied to engineer and screen S. cerevisiae mutants with improved acetaldehyde tolerance, respectively. Crucial genes associated with acetaldehyde tolerance will be recognized by serial analysis of gene expression of the mutants. The function of each genes will be examined by individual gene knockouts on the phenotype. The results of this research will be a guideline to engineer S. cerevisiae strains with improved acetaldehyde tolerance.

乙醛对酿酒酵母具有较强的胁迫性，可抑制菌株生长，影响代谢的正常进行。识别酿酒酵母抗乙醛胁迫关键基因对工业菌株改良具有重要意义。本项目拟采用全转录机制工程（gTME）和定向驯化技术构建、筛选酿酒酵母抗乙醛胁迫表型突变菌株，利用基因表达系列分析（SAGE）技术对突变菌株和对照菌株的差异基因表达谱进行分析，并借助生物信息学软件识别出酿酒酵母抗乙醛胁迫的关键基因，最后通过基因敲除或过表达手段对识别出的关键基因进行功能验证。本项目的研究结果将有助于理解酿酒酵母抗乙醛胁迫应答的分子机制，为酿酒酵母工程菌株的基因改良提供理论依据。

乙醛是酵母酒精发酵的中间产物，高浓度乙醛对酿酒酵母菌株具有一定的胁迫性，影响发酵的正常进行。本项目以研究酿酒酵母抗乙醛胁迫关键基因识别为目标，通过易错PCR技术对酿酒酵母BY4741通用转录因子编码基因进行随机突变。构建突变文库并对突变菌株抗乙醛胁迫能力进行筛选。筛选培养基中乙醛浓度为2 g/L时，突变菌株的乙醛抗性与原始菌株差异不显著。在本试验条件下，通过易错技术产生转录因子编码基因随机突变，未能筛选到乙醛抗性显著提高的突变菌株。后续研究中需要改进突变方法及筛选技术，从菌株代谢过程调控出发，探索酿酒酵母乙醛胁迫相关基因。