过表达OsCatC基因水稻的获得及其耐盐性机理分析  

Obtainment of OsCatC Overexpressing Transgenic Rice and Analysis of Its Mechanism Underlying Salt Tolerance

作　　者：邓勇 刘聪 田野 刘聪美 刘选明[1] 林建中[1] DENG Yong;LIU Cong;TIAN Ye;LIU Congmei;LIU Xuanming;LIN Jianzhong(Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation,College of Biology, Hunan University, Changsha 410082, China)

机构地区：[1]湖南大学生物学院植物功能基因组学和发育调控湖南省重点实验室,长沙410082

出　　处：《西北植物学报》2021年第1期1-8,共8页Acta Botanica Boreali-Occidentalia Sinica

基　　金：国家自然科学基金(31571635,31871595);湖南省科技重大专项(2018NK1010);湖南省自然科学基金(2020JJ4004);长沙市科技计划(kq1801001);杂交水稻国家重点实验室(湖南杂交水稻研究中心)开放课题(2019KF02);广西水稻遗传育种重点实验室开放课题(2018-05-Z06-KF05)。

摘　　要：过氧化氢酶C(Catalase C,CatC)作为重要的抗氧化酶,在水稻发育和胁迫响应方面起重要作用。为了探究CatC在盐胁迫响应中的功能及其作用机制,该研究构建了OsCatC过表达转基因水稻,并比较了其耐盐性和相关抗逆生理指标的变化。结果表明:(1)成功构建过量表达载体pCUbi1390-OsCatC-Flag,并经农杆菌介导的愈伤组织转化获得了30个独立转基因株系(T0),Western blot鉴定T1代幼苗共获得2个OsCatC过表达株系(OE-10、OE-18);qRT-PCR分析发现,OE-10和OE-18株系的OsCatC转录水平显著高于野生型(WT),证明OsCatC基因已成功过表达于转基因株系(OE-10、OE-18)中,且能正常翻译为融合蛋白CatC-Flag。(2)正常水培条件下,OE-10和OE-18与WT的水稻幼苗长势无明显差异,200 mmol·L-1 NaCl处理7 d后再恢复水培10 d时,OE-10和OE-18幼苗的存活率为20%~25%,而WT幼苗绝大部分则干枯死亡,存活率仅为5%左右。(3)与WT相比,OsCatC过表达水稻(OE-10和OE-18)幼苗的耐盐性显著增强,其盐胁迫下的相对电导率、丙二醛和H_(2)O_(2)含量显著降低,过氧化氢酶活性显著升高;4μmol·L^(-1)甲基紫精(MV)处理7 d后,OE-10和OE-18生长受抑制程度显著小于WT,且幼苗的长度均显著大于WT,表现出较强的氧化胁迫耐受性。研究发现,CatC主要通过降解盐胁迫下过量积累的H_(2)O_(2),减轻氧化损伤,从而提高水稻的耐盐性,证明过表达OsCatC能显著提高水稻对盐胁迫的耐受性,且OsCatC是一个非常好的水稻耐盐候选基因。Catalase C(CatC),as an important antioxidant enzyme,plays important roles in rice development and stress response.In order to explore the function and mechanism of CatC involved in the response to salt stress in rice,we constructed OsCatC overexpressing transgenic rice plants,and analyzed their salt tolerance and related physiology in this study.The results demonstrated that:(1)an expression vector pCUbi1390-OsCatC-Flag was successfully constructed,thirty independent transgenic lines(T0)were obtained via Agrobacterium-mediated callus transformation,and two OsCatC overexpressing transgenic lines(OE-10,OE-18)were identified by Western blot analysis of T1 seedlings.Meanwhile,qRT-PCR analysis revealed that the transcriptional levels of OsCatC in OE-10 and OE-18 were significantly higher than that of wild-type(WT)plants,indicating that OsCatC was successfully overexpressed and normally translated into the fusion protein CatC-Flag in the transgenic lines(OE-10,OE-18).(2)Under normal hydroponic conditions,no significant difference was observed in the seedling growth between WT and transgenic lines OE-10 and OE-18,while treated with 200 mmol·L^(-1) NaCl for 7 d and recovered for 10 d,the survival rates of OE-10 and OE-18 seedlings remained 20%-25%,whereas the majority of WT seedlings dried up and died,and the survival rate was only about 5%.(3)Compared with WT,OsCatC overexpressing seedlings(OE-10,OE-18)exhibited higher tolerance to salt stress with lower relative electrolyte leakage,malondialdehyde(MDA)and H_(2)O_(2) contents,and their catalase activities were markedly increased.The growth inhibition degree of OE-10 and OE-18 was significantly lower than that of WT,and their seedling length was markedly higher than that of WT after treatment with 4μmol·L-1 methylviolet(MV)for 7 d,indicating that the transgenic lines had a stronger tolerance to oxidative stress.This study discovered that CatC reduces oxidative damage mainly by degrading the excessive accumulation of H_(2)O_(2) under salt stress,thereby improving the s

关 键 词：水稻 过氧化氢酶C 盐胁迫 氧化胁迫 

分 类 号：Q785[生物学—分子生物学] Q786