维生素B12（VB12）是一种重要的咕啉类化合物。目前，基于脱氮假单胞菌开发的好氧发酵工艺已成为国内外工业化生产VB12的主流。然而关于VB12好氧合成途径基因表达调控方面的研究尚无报道。基于脱氮假单胞菌菌株所开发的遗传工具甚少，从而限制了其代谢调控的相关研究。随着生物技术的发展，大肠杆菌由于其遗传工具丰富等诸多优点，已经广泛应用于基因表达调控以及各种医药化合物及化工原料合成的代谢工程研究。本项目将在大肠杆菌5-氨基乙酰丙酸合成途径调控研究的基础上，通过基因组装以及基因组整合在大肠杆菌内构建VB12好氧合成途径，进而系统分析VB12好氧合成途径的基因表达及其中间代谢产物，以解析关键基因及其调控节点。同时，通过多种策略分析各关键基因的优化表达对 VB12合成的影响。该研究结果将为构建高产VB12的工程菌株提供重要的指导作用。

Vitamin B12 (VB12) is an important kind of corrin compound. Currently, the developed aerobic fermentation process based on P. denitrificans has become the mainstream for VB12 production. However, to date, no research about gene expression and regulation of aerobic VB12 synthetic pathway was reported. At present, the genetic tools developed for operation in P. denitrificans were very limited, which further restricted study of the related metabolism. Along with the development of biotechnology, Escherichia coli, because of its many advantages, such as rich genetic tools developed, has been widely used in gene expression analysis and metabolic engineering for several medicine compound and industrial material production. Based on previous study on regulation and optimization of 5-aminolevulinic acid synthesis, we will construct the aerobic VB12 synthesis pathway in E. coli through gene assembly and genomic integration. By systematically analyzing the level of gene expression and intermediary metabolite, we will resolve the critical gene and the control note of the pathway. Meanwhile, we will also analyze the effect of optimized expression of the key enzyme on VB12 synthesis by means of several strategies. The results obtained will give a meaningful aid for constructing good characterized strain for VB12 microbial production.