《植物生理学报》 2014, 50(10): 1585-1592

羊草水孔蛋白基因LcPIP的克隆与表达分析

张旸, 孙天旭, 汤明威, 郭勃予, 解莉楠, 李玉花*

东北林业大学生命科学学院, 哈尔滨150040

通信作者：李玉花；E-mail: lyhshen@126.com；Tel: 0451-82191783

摘　要：

本研究基于羊草水孔蛋白EST序列, 应用RACE克隆技术从盐胁迫的羊草中克隆了cDNA全长为1 204 bp的水孔蛋白基因, 其GenBank登录号为KJ459872。经生物信息学分析, 该基因开放阅读框为879 bp, 其编码的蛋白含有292个氨基酸, 蛋白质分子量为30.93 kDa, 理论等电点(pI)为7.00, 与已知的小麦、大麦和玉米等单子叶植物来源的同类基因同源性较高, 相似性为80%~98%, 与小麦TaPIP1的遗传关系最近, 与PIP1家族聚为一类。荧光定量PCR分析显示, 在200 mmol•L^-1的Na₂CO₃胁迫不同时间后, 羊草根中LcPIP基因的表达量在6 h时受到抑制, 12~24 h之间表达量明显上升, 但胁迫持续48 h后, LcPIP表达量降至极低水平。羊草叶片的LcPIP基因表达量逐渐上升, 48 h达到最高峰, 72 h表达量下降。

关键词：RACE; 质膜内在蛋白; 生物信息学; 羊草; 表达分析

收稿：2014-07-31   修定：2014-08-27

资助：“十二五”农村领域国家科技计划课题项目(2013AA102706)。

Cloning and Expression Analysis of LcPIP cDNA from Leymus chinensis

ZHANG Yang, SUN Tian-Xu, TANG Ming-Wei, GUO Bo-Yu, XIE Li-Nan, LI Yu-Hua*

College of Life Sciences, Northeast Forestry University, Harbin 150040, China

Corresponding author: LI Yu-Hua; E-mail: lyhshen@126.com; Tel: 0451-82191783

Abstract:

Based on the PIP EST sequence of Leymus chinensis, a full length cDNA named LcPIP from L. chinensis under salinity stress was cloned by RT-PCR RACE. Sequence analysis indicated that the gene contained 1 024 bp nucleotide, and an open reading frame (ORF) encoding 292 amino acids. The molecular weight of the protein was 30.93 kDa and theoretical pI was 7.00. Homology analysis showed that the deduced amino acid sequence of LcPIP shared 85%–98% identity with PIP from monocotyledon, such as Triticum aestivum, Hordeum vulgare and Zea mays, had a closest genetic relationship with wheat TaPIP1, belonged to the PIP1 family. The expression of LcPIP was analyzed by fluorogenic quantitative PCR under Na₂CO₃ stress. With the extension of Na₂CO₃ stress time, the expression of LcPIP dropped from 0 to 6 h in the roots, and then decreased, the peak of expression level arrived at 12 h; but dropped after 24 h. In the leaves, the expression of LcPIP decreased gradually, and the peak of expression level arrived at 48 h; and then the LcPIP expression dropped after 48 h.

Key words: RACE; PIP; bioinformatics; Leymus chinensis; expression analysis