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。

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

Abstract: 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 and it belongs to the non-secretory pro-teins.The amino acid identity of OsPP6C with AtFyPP1 and AtFyPP3 was 93.73%and 93.40%,respectively;the phylogenetic tree shows that OsPP6C has the closest genetic relationship with wheat TaPP 6C.The cis-acting element analysis in the promoter of this gene shows that besides the TATA box and CAAT box,the promoter also contains multiple light-responsive elements,various responsive elements in relation to hormones such as ABA,ethylene,aux-in,methyl jasmonate ( MeJA ) and gibberellic acid, and the regulatory elements involved in various abiotic stresses such as low temperature,heat stress and drought. This paper will lay a solid foundation for further studying expres-sion characteristics and functions of OsPP6 C gene in rice.

Key words: Oryza sativa, Protein phosphatase 6, Vector construction for overexpression, Bioinformatics analysis

中图分类号: 

• S511