全局转录工程选育抗铅表型酿酒酵母及其重金属抗逆分子机制研究

项目名称： 全局转录工程选育抗铅表型酿酒酵母及其重金属抗逆分子机制研究

项目编号： No.21506101

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 有机化学

项目作者： 朱丽英

作者单位： 南京工业大学

项目金额： 21万元

中文摘要： 生物修复治理化工行业重金属污染一直是生物化工研究领域的热门课题。酵母是重金属生物修复的最理想微生物之一，具有多种优良特性，但依然存在重金属抗性差的缺点。针对这一问题，本项目以模式真核生物酿酒酵母为出发菌，采用面向RNA聚合酶的全局转录机器工程这一定向进化新策略，通过引入随机片段交换法对影响RNA聚合酶与启动子结合偏好性和结合效率的关键转录因子随机突变，从而对细胞整体转录进行大规模的重组，构建多尺度细胞表型突变文库，以化工行业引起的典型重金属污染物铅为筛选压力，多轮定向筛选获得高抗铅的酵母突变株；进而联用RNA测序技术和表型芯片技术的交互式和可视化分析，鉴定原始菌和突变株中与重金属抗性相关的差异基因，获得相关基因表达水平及调控机理的基础数据，解析酿酒酵母突变株重金属抗逆的分子机制。本研究既为抗逆微生物细胞工厂的遗传改造提供新思路，又为重金属微生物修复的工业化应用奠定了坚实的理论基础。

中文关键词： 重金属抗性；酿酒酵母；全局转录机器工程；转录组测序；表型芯片

英文摘要： Bioremediation of heavy metal contamination from chemical industry has become the focus in the field of biochemical engineering research. Owing to its good features, yeast is thought to be one of the ideal type of microorganisms for bioremediation of heavy metal contamination, but it still suffers from low tolerance of heavy metal. This project aims to screen higher heavy metal tolerance Saccharomyces cerevisiae strain using global transcription machinery engineering (gTME). Large-scale recombination will be performed, and multi-scale phenotypic mutation library will be constructed, due to key transcription factor random mutation by random segments exchange. Lead will be selected as the screening stress, and the strains of high tolerance to heavy metal will be obtained with multiple times of directed screening. Differential genes associated with lead-tolerance will be found when comparing several mutants with wild type with the combing use of RNA sequencing and phenotype microarrays analysis. Meanwhile, the essential data of corresponding gene exression and regulation mechanism will be obtained, and the molecular mechanism of heavy metal resistance of Saccharomyces cerevisiae mutant strains will be analyzed. This research not only generates new idea for the genetic modification of tolerant microbial cellular factories, but also lays a solid theoretical foundation and offers technical support for the industrial application of microbial remediation of heavy metal.

英文关键词： Heavy metal resistance;Saccharomyces cerevisiae;Global transcriptional machinery engineering;RNA sequencing;Phenotype Microarrays