蝴蝶兰液泡型ATP酶E亚基基因的克隆及序列分析  

Cloning and Sequence Analysis of Vacuolar Proton-ATPase E Subunit Gene from Phalaenopsis amabilis

作　　者：袁秀云[1] 田云芳[2] 梁芳[1] 蒋素华[1] 许申平[1] 王默霏[1] 崔波[1] 

机构地区：[1]郑州师范学院生物工程研究所,河南郑州450044 [2]郑州师范学院生命科学学院,河南郑州450044

出　　处：《河南农业科学》2016年第6期104-110,共7页Journal of Henan Agricultural Sciences

基　　金：河南省科技攻关项目(092102110128)

摘　　要：E亚基是液泡型ATP酶(V-ATPase)多亚基复合体的重要组成部分,响应植物的非生物胁迫,对其基因的克隆及分析有助于阐释其逆境条件下的分子调节机制。根据蝴蝶兰低温诱导差异蛋白的序列设计简并引物,利用RT-PCR和RACE技术从蝴蝶兰叶片中克隆获得2条V-ATPase E亚基的c DNA序列,分别命名为Ph VHA-Ea和Ph VHA-Eb,Gen Bank登录号分别为KT758849和KT758850。Ph VHA-Ea和Ph VHA-Eb全长分别为960 bp和1 036 bp,二者均具有687 bp的开放阅读框,编码228个氨基酸,包含相同的v ATP-synt_E保守域;生物信息学分析表明,2个序列编码蛋白具有相似的二级结构和相同的亲水性,预测的三级结构也相同。系统进化树分析显示,蝴蝶兰Ph VHA-Ea和Ph VHA-Eb与单子叶植物液泡型ATP酶E亚基距离最近。E subunit is an important part of the multisubunit complex of V-ATPase,which responds to abiotic stress in plant. Cloning and analysis of the gene encoding V-ATPase E subunit is helpful to elucidate its molecular regulation mechanism under stress. In this paper,two c DNA sequences of VATPase E subunit genes from Phalaenopsis amabilis leaf were isolated by the methods of RT-PCR combined with RACE techniques using degenerate primers designed according to the sequence of the differentially expressed protein. They were named as Ph VHA-Ea and Ph VHA-Eb,the Gen Bank accession numbers were KT758849 and KT758850 respectively. The two sequences length were 960 bp and 1 036 bp with the intact open reading frame of 687 bp,encoding a polypeptide of 228 amino acids with the same domain of v ATP-synt_ E. Bioinformatics analysis indicated that the two sequences had similar secondary structure,the same feature of hydrophilic protein and the same three-dimensional structure. The phylogenetic analysis revealed Ph VHA-Ea and Ph VHA-Eb were closed to the V-ATPase E subunits gene from monocots.

关 键 词：蝴蝶兰 液泡型ATP酶 E亚基 RACE 序列分析 

分 类 号：S682.31[农业科学—观赏园艺]