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中国生物工程杂志

China Biotechnology
China Biotechnology  2014, Vol. 34 Issue (06): 61-67    DOI: 10.13523/j.cb.20140609
    
Accumulation of Aspartic Acid in Escherichia coli W3110 is Improved by Homologous Recombination
MA Huai-yuan, HUANG Fei, BAI Lin-han
Key Laboratory of Bio-resources and Eco-environment of Ministry of Education, College of Life Sciences, Sichuan University, Chengdu 610064, China
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Abstract  

There are three kinds of aspartate kinase in the metabolic pathway of Escherichia coli, including LysC, MetL and ThrA. Lysine, methionine and threonine are synthesized after aspartic acid phosphorylation pathway caused by aspartate kinase so that aspartic acid is not able to be accumulated to a high concentration in E. coli. Aspartic acid phosphorylation could partly inhibit by gene knock-out. Single gene mutants which lack LysC, ThrA and MetL respectively, are constructed from W3110.After chloroamphenicol resistance removed by pCP20, double gene mutants which lack LysC-ThrA and LysC-MetL respectively are constructed. All mutants are finally confirmed by check primer. These constructions are based on Red recombination system. Concentration of aspartic acid is determined by high performance liquid chromatography. The results show that all mutants except MetL single gene mutants are able to accumulate more L-aspartic acid than wild type. This will lay a foundation for strain improvement by metabolic engineering and contribute to production of aspartic acid by fermentation.



Key wordsEscherichia coli W3110      Red recombination      Gene knock-out      High performance liquid chromatography      L-aspartic acid     
Received: 10 April 2014      Published: 25 June 2014
ZTFLH:  Q784  
Cite this article:

MA Huai-yuan, HUANG Fei, BAI Lin-han. Accumulation of Aspartic Acid in Escherichia coli W3110 is Improved by Homologous Recombination. China Biotechnology, 2014, 34(06): 61-67.

URL:

https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20140609     OR     https://manu60.magtech.com.cn/biotech/Y2014/V34/I06/61


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