基于反向代谢工程的红酵母类胡萝卜素代谢调控分子机制研究

项目名称： 基于反向代谢工程的红酵母类胡萝卜素代谢调控分子机制研究

项目编号： No.31501464

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 农业科学

项目作者： 林心萍

作者单位： 大连工业大学

项目金额： 19万元

中文摘要： 微生物法生产天然类胡萝卜素潜力巨大，然而其代谢调控的关键基因及调控分子机制尚不明晰。本项目拟基于粘红酵母基因组测序的完成和遗传操作系统成功构建的工作基础，首先，采用反向代谢工程策略，以从表型到基因型的研究思路，利用非靶向基因组编辑技术，获得具有特异颜色表型的突变菌株，结合染色体步移及生物信息学手段，确定类胡萝卜素代谢调控的关键基因；其次，利用凝胶迁移率试验、基因过表达、基因敲除、RT-qPCR等分子生物学手段，解析关键基因的功能及调控分子机制；最终，绘制由这些基因构成的类胡萝卜素代谢调控网络。本项目将为红酵母类胡萝卜素生物合成途径的调控及产量的增加提供靶点信息，为调控机制的解析和应用奠定理论基础。项目成果将推动基于微生物的类胡萝卜素产业发展，为现代食品工业注入新活力。

中文关键词： 类胡萝卜素；红酵母；反向代谢工程；关键基因；分子机制

英文摘要： Microorganism hold great potential to produce natural carotenoids, however, relatively little is known about the regulation molecular mechanisms of its key genes. Based on the completion of Rhodotorula glutinis genome sequence and the establishment of its transformation system, this project intends to explore these key genes through a from phenotype to genotype strategies by inverse metabolic engineering. Firstly, a mutant library is constructed by Agrobacterium-mediated transformation, then strains with specific colors is picked out and subjected to genome walking to find out the flanking sequence of T-DNA. By bioinformatics analysis, key genes of carotenoids biosynthetic regulation are identified. After that, molecular biology technologies such as EMSA, gene overexpression, gene knockout, RT-qPCR are used to analyze the genes’ function and regulation mechanism. Finally, a carotenoids regulatory networks consisted of these key genes is drawn to provide target information for carotenoid biosynthetic regulation in red yeast. The result is going to lay the theoretical foundation for the comprehension and application of carotenoid biosynthetic engineering and regulatory mechanisms. The project will promote the development of microbial carotenoids industrial and give prosperity to the modern food industry.

英文关键词： Carotenoids;Red Yeast;Inverse Metabolic Engineering;Key Genes;Molecular Mechanism