甘蓝型油菜植物螯合肽合酶基因BnPCS1a的分离与功能验证  

Isolation and functional characterization of phytochelatin synthase gene BnPCS1a in Brassica napus

作　　者：张雪洁 张治远 张仕泽 白宁宁[1] 刘敏 彭佳师 ZHANG Xuejie;ZHANG Zhiyuan;ZHANG Shize;BAI Ningning;LIU Min;PENG Jiashi(School of Life and Health Sciences,Hunan University of Science and Technology,Xiangtan,Hunan 411201,China;Xiaoxiang College,Hunan University of Science and Technology,Xiangtan,Hunan 411201,China;Hunan Key Laboratory of Economic Crops Genetic Improvement and Integrated Utilization,Hunan University of Science and Technology,Xiangtan,Hunan 411201,China;Key Laboratory of Ecological Remediation and Safe Utilization of Heavy Metal-Polluted Soils,Hunan University of Science and Technology,Xiangtan,Hunan 411201,China)

机构地区：[1]湖南科技大学生命科学与健康学院,湖南湘潭411201 [2]湖南科技大学潇湘学院,湖南湘潭411201 [3]湖南科技大学经济作物遗传改良与综合利用湖南省重点实验室,湖南湘潭411201 [4]湖南科技大学重金属污染土壤生态修复与安全利用湖南省高校重点实验室,湖南湘潭411201

出　　处：《植物生理学报》2023年第5期861-868,共8页Plant Physiology Journal

基　　金：湖南省自然科学基金(2022JJ20022);湖南省教育厅资助科研项目(20K055和22C0249)。

摘　　要：近年来,作为油菜(Brassica napus)主产区的南方地区土壤镉(Cd)等重金属污染问题凸显,对油菜的安全生产和产量品质构成现实威胁。分离油菜中Cd积累和耐受的关键基因资源并解析其作用机理对于应对重金属污染的油菜遗传改良具有重要的指导意义。通过酿酒酵母(Saccharomyces cerevisiae)遗传互补系统筛选了甘蓝型油菜品种‘Westar’的cDNA文库,分离得到一个得到能够介导酵母对Cd耐受的植物螯合肽合酶(phytochelatin synthase)基因BnPCS1a。克隆到目标基因后首先进行了生物信息学分析,继而将其在酿酒酵母中表达后发现还能够介导耐受锌(Zn)胁迫,但对铜(Cu)胁迫作用不明显。BnPCS1a在油菜中的表达亦受Cd和Zn胁迫的诱导上调。为了进一步验证其植物螯合肽合酶的功能,将其在缺乏PCS的裂殖酵母(Schizosaccharomyces pombe)中表达,发现能够完全互补其Cd敏感表型。进而,将其在拟南芥(Arabidopsis thaliana)AtPCS1基因的启动子驱动下表达于AtPCS1基因的突变体cad1-3中,能够互补cad1-3的Cd胁迫敏感表型。综上所述,我们分离的BnPCS1a基因具有植物螯合肽合酶的功能,可能在油菜抵御Cd和Zn胁迫的过程中发挥作用。The rising pollution of heavy metals such as cadmium(Cd)in the southern region,which is the main producing area of oilseed rape(Brassica napus)in China,have posed a threat to the safe production and yield quality of rapeseed.Isolating the key genes responsible for Cd accumulation and tolerance in oilseed rape and uncovering their functional mechanism are of great significance for guiding the genetic improvement to cope with heavy metal pollution.Here we screened the cDNA library of B.napus‘Westar’using yeast genetic complementation system,and isolated a phytochelatin synthase(PCS)gene BnPCS1a that can mediate Cd tolerance in yeast.After cloning the target genes,bioinformatics analysis was performed first.Subsequently,BnPCS1a were expressed in Saccharomyces cerevisiae,and the transformants showed more tolerant to Cd and Zn,but had no effect on excess Cu stress.The expression of BnPCS1a in‘Westar’was also up-regulated by Cd and Zn stress.To further verify its function,we expressed it in PCS-deficient Schizosaccharomyces pombe,and its Cd-stress sensitive phenotype can be completely complemented.Furthermore,expressing it in cad1-3,an Arabidopsis thaliana mutant of AtPCS1 gene,under driving of the promotor of AtPCS1 can also complement the Cd-stress sensitive phenotype to the level of wild type plants.The results show that the BnPCS1a can be functioned as a phytochelatin synthase gene,and play a role in detoxicification of excess Cd and Zn in B.napus.

关 键 词：镉 油菜 植物螯合肽 

分 类 号：S565.4[农业科学—作物学]