%0 Journal Article
%T Construction of recombinant plasmids containing threonine operon and their effects on L-threonine accumulation<br>含苏氨酸操纵子重组质粒的构建及其对大肠杆菌L-苏氨酸积累的影响
%A 张雪
%A 闫继爱
%A 于雷
%A 张国强
%A 张芸
%A 陈宁
%A 温廷益
%J 微生物学报
%D 2009
%I 
%X Abstract: Objective] We reconstructed two recombinant plasmids and studied their effects on L-threonine accumulation of Escherichia coli W3110. Methods] We amplified the threonine operon containing ThrLp promoter, lead peptide thrL, thrA thrB and thrC genes by PCR from E. coli W3110 chromosome and ligated it into the pMD19 T-vector. Site-directed mutation were carried out by gene splicing by overlap extension PCR to release the feedback inhibition of aspartokinase I (thrA). Two recombinant plasmids WYE112 and WYE134 were transformed into E. coli W3110 by electroporation. Fed-batch cultures of E. coli W3110 were carried out in 5-Liter fermentors and the L-threonine concentration was measured by HPLC. Results] Fed-batch fermentation results showed that E. coli W3110 could accumulate little L-threonine (0.036 ± 0.004 g/L) but recombinant E. coli W3110 harboring the plasmid WYE112 containing a threonine operon exhibited a L-threonine production of 2.590 ± 0.115 g/L. Furthermore, L-threonine production reached 9.223 ± 1.279 g/L when the feedback inhibition of thrA was released. Conclusion] Overexpression of threonine operon can lead to the accumulation of L-threonine. Further release of feedback inhibition of aspartokinase I can enhance its accumulation.
%K Keywords: L-threonine
%K gene splicing by overlap extension PCR
%K Escherichia coli
%K fermentation
%K site-directed mutation<br>关键词：L-苏氨酸
%K 重叠延伸PCR
%K 大肠杆菌
%K 发酵
%K 定点突变
%U http://www.alljournals.cn/get_abstract_url.aspx?pcid=90BA3D13E7F3BC869AC96FB3DA594E3FE34FBF7B8BC0E591&jid=A3C6BA55AB623B90FA9104CFFC826F3C&aid=369BF9A3A9E6E152885EEB5B214BFAA0&yid=DE12191FBD62783C&vid=2A3781E88AB1776F&iid=94C357A881DFC066&sid=E543FC2C7CA75C74&eid=6341CCF6B158C5F9&journal_id=0001-6209&journal_name=微生物学报&referenced_num=3&reference_num=10