甘蔗抗坏血酸过氧化物酶基因ScAPX1的克隆和表达分析  

Cloning and Expression Analysis of Peroxidase Gene(ScAPX1)from Sugarcane

作　　者：张保青[1] 邵敏[2,4] 黄玉新 黄杏[1] 宋修鹏[1] 陈虎[3] 王盛[2] 谭秦亮[5] 杨丽涛[2] 李杨瑞[1,2] ZHANG Bao-qing;SHAO Min;HUANG Yu-xin;HUANG Xing;SONG Xiu-peng;CHEN Hu;WANG Sheng;TAN Qin-liang;YANG Li-tao;LI Yang-rui(Sugarcane Research Institute,Guangxi Academy of Agricultural Sciences/Sugarcane Research Center,Chinese Academy of Agricultural Sciences/Guangxi Key Laboratory of Sugarcane Genetic Improvement/Ministry of Agriculture Key Laboratory of Sugarcane Biotechnology and Genetic Improvement(Guangxi),Nanning 530007;College of Agriculture,Guangxi University,Nanning 530005;Guangxi Forestry Research Institute,Nanning 530002;Nanning City Greening Project Management Center,Nanning 530011;Guangxi Subtropical Crops Research Institute,Nanning 530001)

机构地区：[1]广西农业科学院甘蔗研究所中国农业科学院甘蔗研究中心,广西甘蔗遗传改良重点实验室,农业部广西甘蔗生物技术与遗传改良重点实验室,南宁530007 [2]广西大学农学院,南宁530005 [3]广西林业科学研究院,南宁530002 [4]南宁市绿化工程管理中心,南宁530011 [5]广西壮族自治区亚热带作物研究所,南宁530001

出　　处：《生物技术通报》2019年第12期31-37,共7页Biotechnology Bulletin

基　　金：国家重点研发计划(2018YFD1000500);国家自然科学基金项目(31760415,31660356);广西科技计划(桂科AA17202042-1);广西农业科学院基本科研业务专项项目(桂农科2018JZ16，桂农科2017JZ19);广西科技基地和人才专项(桂科AD17195100);广西八桂学者专项(2013-03);国家现代农业产业技术体系广西甘蔗创新团队专项(gjnytxgxcxtd-03-01)

摘　　要：通过克隆甘蔗ScAPX1及分析其在低温胁迫中的表达,为深入研究甘蔗APX1在低温胁迫中的功能及为甘蔗抗寒育种的分子机理提供依据。以甘蔗叶片总RNA为模板,通过RT-PCR技术克隆甘蔗叶片ScAPX1的完整ORF序列,采用生物信息学软件分析所克隆基因的编码蛋白特性,并用荧光定量PCR分析该基因在2个抗寒性差异较大的甘蔗品种GT28和YL6低温胁迫下的表达模式。结果表明,克隆得到甘蔗ScAPX1(NCBI登录号为KC794939),其包含一个759 bp的完整开放阅读框,编码252个氨基酸。该基因编码的蛋白不含信号肽,为可溶性蛋白,无跨膜结构,定位于细胞质,与高粱氨基酸的同源性为98%,推测其为胞质型抗坏血酸过氧化物酶基因。qRT-PCR分析结果表明,随着低温(0-4℃)时间的延长,2个甘蔗品种ScAPX1的表达量均是先上升后下降,但表达量存在差异,在整个胁迫过程中,抗寒强品种GT28的表达量始终比抗寒弱品种YL6高。甘蔗ScAPX1积极响应低温逆境胁迫,该基因的诱导表达与甘蔗品种本身的抗寒性密切相关。Here cloning the ScAPX1 of sugarcane and analyzing its expression under low temperature stress is to provide a basis for further studying the function of APX1 gene in sugarcane under low temperature stress and investigating the molecular mechanism of breeding cold stress-resistant sugarcane.Having total RNA of sugarcane leaf as template,RT-PCR technique was used to clone the complete ORF sequence of ScAPX1 from sugarcane leaf,bioinformatics software was to analyze the characteristics of the encoding protein,and quantitative realtime PCR(qRT-PCR)method was applied to study the expressions of ScAPX1 gene under low temperature stress in two sugarcane varieties GT28 and YL6 with widely different cold resistance.The results showed that the ScAPX1 gene(GenBank accession number:KC794939)in sugarcane was cloned,which contained a complete open reading frame of 759 bp and encoded 252 amino acids.The protein encoded by this gene contained no signal peptide and no transmembrane structure,was a soluble protein and located in cytoplasm,and its homology with sorghum amino acid was 98%.It was inferred that ScAPX1 gene was a cytoplasmic ascorbic acid peroxidase gene(ScAPX).qRT-PCR analysis showed that with the extension of low temperature(0-4oC),the expression levels of ScAPX1 gene in two sugarcane varieties increased first and then decreased,but differed.In the whole process of cold stress,the relative expression level in GT28,a cold resistant variety,was always higher than that in YL6,a cold sensitive variety.This result suggests that the ScAPX1 gene of sugarcane is active in response to low temperature stress,and the induced expression of this gene is closely related to the cold resistance of sugarcane varieties.

关 键 词：甘蔗 ScAPX1 基因克隆 低温胁迫 表达分析 

分 类 号：R73[医药卫生—肿瘤]