茶树二氢黄酮醇4-还原酶基因的克隆、表达及功能分析

doi:10.13305/j.cnki.jts.2013.03.010

Abstract

Abstract: Dihydroflavonol 4-reductase (DFR) is a key enzyme in the biosynthesis of catechins in tea plant. However, the functions and the zymologic properties of DFR were not deeply identified in recent researches. The open reading frame of DFR gene, which encoding a 347 amino acids protein, was cloned from tea plant (Camellia sinensis) by RT-PCR. The deduced protein molecular weight was 38.69βkD and its theoretical isoelectric point was 6.02. The gene was cloned into the expression vector SUMO for expression in prokaryotic cells. The SDS-PAGE results showed that the dihydroflavonol 4-reductase peoteins was expressed in Escherichia coli BL21. The optimal inducing conditions including time, temperature and IPTG concentration were studied. The deduced protein was purified and its activity was detected by HPLC-MS method. The results indicated that purified protein showed the DFR activity, catalyzed the reduction reaction of DHQ and DHM. The research provides a valuable foundation for better understanding the substrate specificity and enzymatic properties of CsDFR.

Key words: tea plant (Camellia sinensis), dihydroflavonol 4-reductase, prokaryotic expression, enzyme activity

CLC Number:

WANG Yun-sheng, XU Yu-jiao, HU Xiao-jing, JIANG Xiao-lan, YANG Qing, LI Wei-wei, LIU Ya-jun, GAO Li-ping, XIA Tao. Clone, Expression and Functional Analysis of Dihydroflavonol 4-Reductase Gene of Tea Plant (Camellia sinensis)[J]. Journal of Tea Science, 2013, 33(3): 193-201.

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Clone, Expression and Functional Analysis of Dihydroflavonol 4-Reductase Gene of Tea Plant (Camellia sinensis)

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