产D-乳酸重组大肠杆菌ptsG基因的敲除及其混合糖同步发酵

doi:10.13560/j.cnki.biotech.bull.1985.2015.12.032

生物技术通报 ›› 2015, Vol. 31 ›› Issue (12): 221-226.doi: 10.13560/j.cnki.biotech.bull.1985.2015.12.032

产D-乳酸重组大肠杆菌ptsG基因的敲除及其混合糖同步发酵

丁小云,顾健健,王永泽,赵锦芳,王金华,赵筱

湖北工业大学轻工学部发酵工程教育部重点实验室，武汉 430068

收稿日期:2014-12-17 出版日期:2015-12-19 发布日期:2015-12-19
作者简介:丁小云，男，硕士研究生，研究方向:发酵工程；E-mail:858709213@qq.com
基金资助:
国家自然科学基金项目(NSFC31070094)，湖北省自然科学基金项目(2011CDA008，2011CDB076)，湖北工业大学博士科研启动基金项目(BSQD12143)

The Knockout of Gene ptsG of Recombinant Escherichia coli Producing D-lactic Acid and the Simultaneous Fermentation of Mixed Sugars

Ding Xiaoyun, Gu Jianjian, Wang Yongze, Zhao Jinfang, Wang Jinhua, Zhao Xiao

Key Laboratory of Fermentation Engineering of Ministry of Education，Hubei University of Technology，Wuhan 430068

Received:2014-12-17 Published:2015-12-19 Online:2015-12-19

摘要/Abstract

摘要：

为构建能够同时高效利用五碳糖和六碳糖发酵产D-乳酸的重组大肠杆菌工程菌，以能高效利用五碳糖发酵产D-乳酸的大肠杆菌工程菌E.coli JH13为出发菌株，通过Red同源重组技术敲除葡萄糖跨膜转运基因ptsG。实验结果表明，ptsG缺陷菌株E.coli JH15在10%混合糖(5%葡萄糖和5%木糖)培养基中发酵，可同时利用五碳糖和六碳糖以完成发酵；而对照菌葡萄糖消耗完才利用木糖，发酵结束还有 18 g/L木糖残留；JH15乳酸产量为83.04 g/L，相比于对照菌株提高了25.86%；在稻草秸秆水解液中发酵，JH15同时利用葡萄糖、木糖和L-阿拉伯糖，乳酸产量为25.15 g/L，转化率为86.42%。JH15 作为能利用混合糖同步发酵产D-乳酸的大肠杆菌工程菌，它的成功构建为利用廉价的木质纤维素水解物为原料发酵生产D-乳酸提供参考依据。

关键词: 重组大肠杆菌工程菌, D-乳酸, ptsG基因, Red同源重组, 混合糖

Abstract:

In order to construct a recombinant engineering Escherichia coli strain that yields D-lactic acid in the simultaneous efficient fermentation of pentose and hexose, having an engineering E. coli JH13 that efficiently ferment the pentose to produce D-lactic acid as an original strain, a glucose transmembrane transporter gene ptsG was knocked out by the technique of Red homologous recombination. The fermentative results showed that in the 10% mixed sugars(5% glucose and 5% xylose), the ptsG-deleted strain E. coli JH15 simultaneously utilized pentose and hexose to complete the fermentation；however, the control strain started to utilize the xylose only after glucose was consumed up, and 18 g/L xylose still remained after the fermentation completed. The production of D-lactic acid by JH15 reached 83.04 g/L, and 25.86% higher than that by control strain JH13. The JH15 as a E. coli strain of producing D-lactic acid during simultaneous fermentation with mixed sugars, its construction provides a reference for producing the D-lactic acid in fermentation while utilizing the low-cost hydrolyzed components of lignocelluloses materials as raw material.

Key words: recombinant Escherichia coli, D-lactic acid, ptsG, Red homologous recombination, mixed sugars

丁小云,顾健健,王永泽,赵锦芳,王金华,赵筱. 产D-乳酸重组大肠杆菌ptsG基因的敲除及其混合糖同步发酵[J]. 生物技术通报, 2015, 31(12): 221-226.

Ding Xiaoyun, Gu Jianjian, Wang Yongze, Zhao Jinfang, Wang Jinhua, Zhao Xiao. The Knockout of Gene ptsG of Recombinant Escherichia coli Producing D-lactic Acid and the Simultaneous Fermentation of Mixed Sugars[J]. Biotechnology Bulletin, 2015, 31(12): 221-226.

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产D-乳酸重组大肠杆菌ptsG基因的敲除及其混合糖同步发酵

The Knockout of Gene ptsG of Recombinant Escherichia coli Producing D-lactic Acid and the Simultaneous Fermentation of Mixed Sugars

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