四翅滨藜Osmotin-like Protein 基因的克隆、序列分析及其原核表达

生物技术通报 ›› 2013, Vol. 0 ›› Issue (2): 106-110.

四翅滨藜Osmotin-like Protein 基因的克隆、序列分析及其原核表达

王升阳张勇王健刘言志刘金亮潘洪玉

（吉林大学植物科学学院，长春 130062）

收稿日期:2012-08-15 修回日期:2013-02-27 出版日期:2013-02-26 发布日期:2013-02-27
作者简介:王升阳，男，研究方向：植物抗逆基因工程；E-mail ：sywang8209@mails.jlu.edu.cn张勇，男，研究方向：植物抗逆基因工程；E-mail ：zhangyong199010@gmail.com
基金资助:
“十二五”国家科技支撑计划（2012BAD19B04），农业部转基因生物新品种培育重大专项（2009ZX08009-062B）

Cloning and Sequence Analysis of a Osmotin-like Protein Gene from Atriplex canescens and Its Prokaryotic Expression

Wang Shengyang Zhang Yong Wang Jian Liu Yanzhi Liu Jinliang Pan Hongyu

（College of Plant Sciences，Jilin University，Changchun 130062）

Received:2012-08-15 Revised:2013-02-27 Published:2013-02-26 Online:2013-02-27

摘要/Abstract

摘要： 在构建四翅滨藜（Atriplex canescens）全长cDNA 文库中通过随机克隆测序并进行EST 分析基础上，得到四翅滨藜osmotin-like protein 的1 个cDNA 序列，命名为AcOLP。AcOLP cDNA 包含一个全长为687 bp 完整开放阅读框，编码229 个氨基酸，属于GH64-TLP-SF 超家族，是一种病程相关5（PR-5）蛋白，其核酸序列与大洋洲滨藜（Atriplex nummularia）的osmotin-likeprotein 基因的同源性为94%，对应编码氨基酸序列同源性为87%。将得到的序列提交GenBank，序列号为JN632587.1。与其他植物PR-5 蛋白的氨基酸序列比对，AcOLP 具有保守的半胱氨酸残基，与二硫键的形成有关。对AcOLP 与其他植物的氨基酸序列的进化分析表明，其与大洋洲滨藜的亲缘关系较近。将AcOLP 基因与原核表达载体pET-28a 连接，进行融合表达，在大肠杆菌BL21（DE3）中诱导表达出分子质量约29 kD 的蛋白。

关键词: 四翅滨藜, Osmotin-like protein, 基因克隆, 序列分析, 原核表达

Abstract: A osmotin-like protein gene was isolated based on the library from Atriplex canescens and its EST analysis, and named as AcOLP. The full length of AcOLP contained an open reading frame of 687 bp. It encoded a polypeptide of 229 amino acids, belonging to GH64- TLP-SF superfamily as a kind of PR-5 proteins. The AcOLP had 94% nucleotide sequence homology and 87% amino acid sequence homology to the sequence of osmotin-like protein from Atriplex centralasiatica, respectively. The accession number of AcOLP in GenBank was JN632587.1. Comparison of amino acid sequence with PR-5 proteins from various plants showed AcOLP possessed 16 cysteine residues that were conserved at their invariant and were presumably involved in disulfide bonding. Phylogenic analysis on the amino acid sequence of AcOLP with other plants showed that Atriplex canescens was closely related to Atriplex nummularia. AcOLP was inserted into the prokaryotic expression vector of pET-28a and expressed its fusion protein（about 29 kD）in Escherichia coli BL21（DE3）.

Key words: Atriplex canescens, Osmotin-like protein, Gene clone, Analysis of sequence, Prokaryotic expression

王升阳, 张勇, 王健, 刘言志, 刘金亮, 潘洪玉. 四翅滨藜Osmotin-like Protein 基因的克隆、序列分析及其原核表达[J]. 生物技术通报, 2013, 0(2): 106-110.

Wang Shengyang, Zhang Yong, Wang Jian, Liu Yanzhi, Liu Jinliang, Pan Hongyu . Cloning and Sequence Analysis of a Osmotin-like Protein Gene from Atriplex canescens and Its Prokaryotic Expression[J]. Biotechnology Bulletin, 2013, 0(2): 106-110.

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