水稻OsPP6C基因的克隆、过表达载体构建与生物信息学分析  

Cloning,Construction of Overexpression Vector and Bioinformatics Analysis of OsPP6C Gene from Oryza sativa

作　　者：王青云[1,2] 王润青[2,3] 方聪燕[2] 侯佩[2] 苏亮[2] 李建平[2] 宋梅芳[2] 杨建平[2,3] 李雪梅[1] 吴大付[3] 

机构地区：[1]沈阳师范大学化学与生命科学学院,辽宁沈阳110034 [2]中国农业科学院作物科学研究所,北京100081 [3]河南科技学院资源与环境学院,河南新乡453003

出　　处：《华北农学报》2013年第5期59-65,共7页Acta Agriculturae Boreali-Sinica

基　　金：国家转基因生物新品种培育重大专项(2011ZX08010-002)

摘　　要：为克隆水稻蛋白磷酸酶6(PP6)催化亚基基因OsPP6C,并构建该基因的过表达载体以及进行生物信息学分析。采用RT-PCR技术从水稻中花11叶片cDNA中扩增OsPP6C基因全长,并构建与GUS融合的pBI121-OsPP6CGUS表达载体。通过生物信息学方法分析了OsPP6C蛋白的理化性质、与拟南芥AtPP6C(即AtFyPP)之间的同源性以及不同物种间的系统发育关系,此外,对OsPP6C基因的启动子进行了顺式作用元件分析。结果表明,该基因转录序列包含一个912 bp的开放阅读框,编码303个氨基酸残基,蛋白的分子量约为3.47 kDa,等电点为5.13;具有疏水性,但不存在信号肽,属于非分泌型蛋白。OsPP6C与拟南芥AtFyPP1和AtFyPP3氨基酸序列一致性分别为93.73%和93.40%,系统进化树显示OsPP6C与小麦TaPP6C的亲缘关系最近,而与其他物种亲缘关系相对较远。对该基因启动子中含有的顺式作用元件分析表明,该启动子中除TATA盒和CAAT盒外,还含有多种参与光应答、激素(ABA、乙烯、生长素、MeJA和赤霉素等)应答以及低温、热激和干旱等非生物胁迫因子应答的调控元件。为进一步研究水稻OsPP6C基因的表达特性和功能奠定了基础。The aims of this study are to clone the catalytic subunit gene of rice protein phosphatase 6 （OsPP6C）,construct the over-expressing vector of this gene and conduct the bioinformatics analysis .Using the RT-PCR technique,the full length of OsPP6C gene was amplified from the complementary DNA （cDNA）of rice Zhong-hua 11 variety,and it was fused with GUS to form pBI121-OsPP6C-GUS expression vector.Through bioinformatics methods,the physiochemical characteristics of OsPP6C protein,the homology among Arabidopsis PP6Cs（i.e.At-FyPPs）and OsPP6C,and the phylogenetic relationship among different species were analyzed;in addition,the cis-acting elements of the OsPP6C promoter were analyzed .The results show that the transcript sequence of this gene contains an open reading frame of 912 base pairs , this gene encodes a peptide of 303 amino acids , the molecular weight and isoelectric point of this protein is approximately 3.47 kDa and 5.13,respectively;the OsPP6C protein The aims of this study are to clone the catalytic subunit gene of rice protein phosphatase 6 （OsPP6C）,construct the over-expressing vector of this gene and conduct the bioinformatics analysis .Using the RT-PCR technique,the full length of OsPP6C gene was amplified from the complementary DNA （cDNA）of rice Zhong-hua 11 variety,and it was fused with GUS to form pBI121-OsPP6C-GUS expression vector.Through bioinformatics methods,the physiochemical characteristics of OsPP6C protein,the homology among Arabidopsis PP6Cs（i.e.At-FyPPs）and OsPP6C,and the phylogenetic relationship among different species were analyzed;in addition,the cis-acting elements of the OsPP6C promoter were analyzed .The results show that the transcript sequence of this gene contains an open reading frame of 912 base pairs , this gene encodes a peptide of 303 amino acids , the molecular weight and isoelectric point of this protein is approximately 3.47 kDa and 5.13,respectively;the OsPP6C protein has the hydrophobic property ,but there is no signal peptide in this protein

关 键 词：水稻 蛋白磷酸酶6 过表达载体构建 生物信息学分析 

分 类 号：Q78[生物学—分子生物学]