茶树DHAR酶基因的克隆与非生物胁迫响应分析

利用RT-PCR技术从茶树‘龙井43’cDNA中克隆得到1个编码茶树脱氢抗坏血酸还原酶基因CsDHAR,该基因全长639 bp,编码212个氨基酸,属于谷胱甘肽转移酶（GST）超级家族成员,具较高保守性。系统进化分析结果表明CsDHAR蛋白与拟南芥DHAR1、DHAR2进化关系近,在不同物种间,与毛果杨、胡杨等植物亲缘关系较近;理化性质分析表明CsDHAR属于亲水性蛋白;结构分析表明CsDHAR蛋白具典型功能结构域,三级结构主要由α–螺旋和不规则卷曲构成。荧光定量PCR分析显示,CsDHAR在茶树叶片不同发育阶段表达量差异不显著;对‘龙井43’与‘安吉白茶’中CsDHAR进行荧光定量PCR分析,‘龙井43’在低温、高温处理4 h表达量最高、干旱和高盐处理24 h表达量最高;‘安吉白茶’在高温、高盐处理4 h表达量最高,干旱处理2 h表达量最高。不同逆境胁迫下CsDHAR均被诱导表达,表明该基因参与茶树响应逆境胁迫调节过程。

Cloning and Expression Analysis under Abiotic Stress of the DHAR Gene in Camellia sinensis

Using RT-PCR technology,the CsDHAR gene encoding dehydroascorbate reductase was cloned from the cDNA of Camellia sinensis‘Longjing 43’. CsDHAR gene was 639 bp in length,encoding 212 amino acids,belonging to glutathione transferase superfamily which is highly conservative. Phylogenetic analysis showed that CsDHAR protein was closely related to Arabidopsis DHAR1 and DHAR2. Among different species,CsDHAR protein was closely related to Populus trichocarpa and P. euphratica. Physicochemical analysis showed that CsDHAR protein was a hydrophilic protein. Structural analysis showed that CsDHAR protein has a typical functional domain,and the tertiary structure is mainly composed of alpha-helix and irregular curl. Quantitative real-time PCR analysis showed that CsDHAR expression levels were not significantly different at different growth stages in leaves of tea plant. CsDHAR gene expression profiles of C. sinensis‘Longjing 43’and‘Anji Baicha’were detected and analyzed by qRT-PCR.‘Longjing 43’had the highest expression level in 4 h under low and high temperature treatments,and the highest expression level in 24 h under drought and high salt treatments. The expression level of‘Anji Baicha’was the highest in 4 h under high temperature and high salt treatments,and the highest expression level in 2 h under drought treatment. Under different abiotic stress treatments,CsDHAR was induced,indicating that the gene was involved in the regulation process of stress response in tea plant.