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Journal of Tea Science ›› 2007, Vol. 27 ›› Issue (1): 61-66.doi: 10.13305/j.cnki.jts.2007.01.009

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The Construction of the Engineered Escherichia coli Strain for the Biosynthesis of Theanine

WANG Xian-bo, WANG Li-yuan, CHENG Hao*, ZHOU Jian, LIN Zhi   

  1. Key Lab of Tea Chemical Engineering, Ministry of Agriculture; Tea Research Institute, Chinese Acade my of Agricultural Sciences, Hangzhou 310008, China
  • Received:2006-07-11 Revised:2006-11-21 Online:2007-03-25 Published:2019-09-11

Abstract: γ-ggt was cloned by PCR from E.coli DH5α. Then, the PCR product of γ-ggt digested with two restriction enzymes, Kpn I and Xho I, was purified and ligated with the pET-32a vector digested with the same enzymes by T4 DNA ligase. Then the ligation product was transformed to E.coli BL21 and the engineered Escherichia coli strain was constructed successfully. The γ-Glutamyltranspeptidase was expressed with induction of 0.05βmmol/L IPTG in 32℃ incubation. The activity of 1βg fresh cells of engineered strain was 2.0βU/g,it is about 15 times higher than that of E.coli DH5α.Under the catalysis of cells of the engineered strain induced with IPTG, the yield of theanine from L-Gln and ethylamine was 29.40βg/L and the conversion rate of L-Gln as to theanine being 48.22%; it is about 100 times higher than that of E.coli DH5α.

Key words: γ-Glutamyltranspeptidase;, theanine, engineered strain, construction

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