费尔干猪毛菜液泡膜氢离子焦磷酸酶基因SfVP的克隆、序列分析及表达  

Cloning and Expression Analysis of Vacuolar H^+-PPase Gene from Salsola ferganica

作　　者：贺怡[1] 钱雯婕[1] 袁磊[1] 李榕[1] 谭兰[1] 谢全亮 李鸿彬[1,2] 

机构地区：[1]石河子大学生命科学学院,石河子832003 [2]石河子大学农业生物技术重点实验室,石河子832003

出　　处：《石河子大学学报（自然科学版）》2014年第3期319-324,共6页Journal of Shihezi University(Natural Science)

基　　金：新疆兵团种质资源创新专项(2012BB050);石河子大学创新团队项目(2011ZRKXTD-0603);石河子大学SRP项目(SRP20130068)

摘　　要：植物液泡膜氢离子焦磷酸酶(Vacuolar H+-PPase,VP)能够酸化液泡并为液泡的次级转运系统提供能量,在植物耐盐性方面发挥着着重要作用。本研究采用RT-PCR结合RACE方法,从费尔干猪毛菜中(Salsola ferganica)克隆得到液泡膜质子焦磷酸酶基因SfVP,该基因的cDNA全长2738 bp,包含1个2301 bp的完整开放读码框(ORF),编码766个氨基酸的多肽。氨基酸序列分析表明其属于H+-转运无机焦磷酸酶超家族成员,具有典型的H+-PPase结构域,其疏水性较强,含有12个跨膜螺旋结构。SfVP序列比对显示具有H+-PPase的3个保守序列CS1、CS2和CS3,其中CS1中含有保守的底物催化位点,与烟草的NtVP氨基酸序列相似性为95.3%。半定量RT-PCR结果表明,SfVP的转录表达丰度随着600 mmol/L NaCl处理时间的增加而升高,处理12 h后显著增加并一直维持在较高水平。Plant vacuolar H^＋-translocating inorganic pyrophosphatase（V-H^＋-PPase）can acidify vacuoles and power the vacuolar secondary active transport system,which plays an important role in plant salt tolerance.In this study,a vacuolar H^＋-PPase gene was cloned from Salsola ferganica by RT-PCR and RACE with a 2738 bp full-length cDNA sequence and a 2301 bp open reading frame（ORF）which encoding a putative polypeptide of 766 amino acids.Sequence analysis showed that SfVP protein belonged to the H^＋-translocating inorganic pyrophosphatase superfamily and had classic H^＋-PPase multidomain.SfVP protein contains 12 transmembrane structures with strong hydrophobicity.Sequence alignment analysis based on amino acid sequences indicated that SfVP had 3 conservative sequence regions including CS1,CS2 and CS3.CS1 contains conservative substrate binding sites,which showed 95.3% similarity with tobacco NtVP.Semi-quantitative RT-PCR results revealed that transcription level of SfVP increases gradually with the increase of treatment time under 600 mmol/L NaCl stress,and maintains a high express abundance after 12 hour treatment.

关 键 词：费尔干猪毛菜 液泡膜氢离子焦磷酸酶 RACE 盐响应表达 

分 类 号：Q781[生物学—分子生物学] Q949.745.1