Collaborative Research: Genetic Analysis and Metabolic Engineering of Solventogenic Clostridia

本研究旨在通过基因改变厌氧溶剂型梭菌（Clostridium acetobutylicum及其相关种）的代谢，以确定其在生产溶剂（丁醇和丙酮）和其他氧化学物质方面的潜在应用。这需要进一步了解应用分子遗传学和重定向细胞代谢所需的相关方法。这项研究的目的是：(1)建立产生稳定的过量生产菌株所需的技术（这种菌株对于不使用任何抗生素的批量或连续过程是必要的，以保持遗传稳定性），(2)对被称为sol位点的一组基因的调控进行更详细的分子理解。(3)开发能够在其他宿主中表达梭菌基因或在梭菌中表达异源基因的方法，以形成各自细胞通常不产生的产品；(4)开发灵活的碳通量控制的附加工具。具体来说，这项研究将开发所需的方法：理想的基因表达（在强度和时间方面），单个或多个染色体基因失活，以失活或消除不希望的途径或特征，以及稳定的染色体整合（一个或多个拷贝）基因，以增加理想途径的蛋白质活性（可能是碳通量）。＊＊＊

9604562 Bennett This proposal is on research to genetically alter the metabolism of anaerobic solventogenic clostridia (Clostridium acetobutylicum and related species) to determine their potential applications for producing solvents (butanol and acetone) and other oxychemicals. This requires further understanding of the applied molecular genetics and associated methods needed to redirect the cellular metabolism. This research is aiming to (1) establish the techniques needed for the generation of stable overproducing strains (such strains are necessary for batch or continuous processes without using any antibiotics to maintain genetic stability), (2) develop a more detailed molecular understanding of the regulation of a cluster of genes known as the sol locus, (3) develop the means which would allow one to express clostridia genes in other hosts or heterologous genes in clostridia in order to form products not normally produced by the respective cells, and(4) develop additional tools for flexible carbon-flux control. Specifically, this research will develop methods needed for: desirable gene expression (in terms of strength and timing), single or multiple chromosomal gene inactivations in order to inactivate or eliminate undesirable pathways or traits, and stable chromosomal integration (of one or more copies) of genes in order to increase the protein activities (and presumably carbon fluxes) of desirable pathways. ***