中间锦鸡儿<em>DHN1</em>基因克隆及表达分析

doi:10.13523/j.cb.20160413

中国生物工程杂志

2016, Vol. 36

Issue (4): 88-96 DOI: 10.13523/j.cb.20160413

研究报告

中间锦鸡儿DHN1基因克隆及表达分析

万永青¹, 杨洋¹, 张春林¹, 万东莉², 杨爱琴³, 杨杞¹, 王瑞刚¹, 李国婧¹

1. 内蒙古农业大学生命科学学院呼和浩特 010018;
2. 中国农业科学院草原研究所呼和浩特 010010;
3. 内蒙古伊金霍洛旗经济作物工作站鄂尔多斯 017200

Cloning and Expression Analysis of CiDHN1 Gene in Caragana intermedia

WAN Yong-qing¹, YANG Yang¹, ZHANG Chun-lin¹, WAN Dong-li², YANG Ai-qin³, YANG Qi¹, WANG Rui-gang¹, LI Guo-jing¹

1. College of Life Sciences, Inner Mongolia Agricultural University, Hohhot 010018, China;
2. Grassland Research Institute, Chinese Academy of Agricultural Sciences, Hohhot 010010, China;
3. Inner Mongolia Yijinhuoluo Economic Crops Workstation, Erdos 017200, China

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摘要：

脱水素(DHNs, dehydrins)是LEA II亚家族蛋白,在种子发育后期大量积累,并受不同逆境胁迫处理诱导表达。在中间锦鸡儿干旱胁迫抑制性削减杂交文库中筛选到了一段 DHN1 序列,利用RACE技术克隆获得基因全长序列。序列比对分析表明, CiDHN1 具有开放阅读框891bp,起始密码子为ATG,终止密码子为TAG,编码297个氨基酸,含有5个Y片段和一个K片段,与CiDHN1相似性最高的为山茱萸科主教红端木(Cornus sericea),相似性为39%。系统进化分析显示CiDHN1单独聚为一支,推测该蛋白为新的功能未知蛋白。亚细胞定位结果表明CiDHN1定位在细胞质、质膜和细胞核。荧光定量PCR结果显示 CiDHN1 的表达受冷、脱水和NaCl等非生物胁迫的诱导。 CiDHN1 过量表达拟南芥后,其中表达量最强的株系对200 mmol/L的NaCl处理较为敏感。CiDHN1在中间锦鸡儿抵抗逆境胁迫中的功能有待于进一步研究。

关键词： 基因克隆; DHN; 表达分析; 亚细胞定位; 中间锦鸡儿

Abstract:

Dehydrins (DHNs; LEA D11 family) are subfamily of group 2 LEA proteins that accumulate to high levels during late stages of seed development and in vegetative tissues subjected to abiotic treatment. A fragment of DHN1 gene was isolated from the suppression subtractive hybridization (SSH) library of Caragana intermedia. The full length sequence of DHN1 was obtained by RACE cloning. Sequence analysis indicated the full length cDNA of CiDHN1 was 1 140 bp, with open reading frame 891bp, deduced encoding 297 amino acids including five Y-segments and one K-segment, the most similarity protein was DHN of Cornus sericea with similarity 39%. CiDHN1 clusters as a unique branch by phylogenetic analysis and was referred as a novel protein with unknown function. Subcellular localization observed CiDHN1 was localized in the plasma membrane and cytoplasm. The transcripts of CiDHN1 were induced under different abiotic stresses, such as cold, dehydration and NaCl. Overexpression of CiDHN1 in Arabidopsis showed susceptible phenotype under 200mmol/L NaCl treatment. The roles of CiDHN1 in Caragana intermedia resist to environmental stresses need further investigation.

Key words: DHN Expression analysis Subcellular localization Gene clone Caragana intermedia

收稿日期: 2015-11-02 出版日期: 2015-12-01

ZTFLH:

Q819

基金资助:

国家自然科学基金(31360169)、内蒙古自然科学基金(2013MS0308)、内蒙古自治区科技创新人才团队、产业创新人才团队项目资助项目

通讯作者: 李国婧 E-mail: liguojing@imau.edu.cn

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引用本文:

万永青, 杨洋, 张春林, 万东莉, 杨爱琴, 杨杞, 王瑞刚, 李国婧. 中间锦鸡儿DHN1基因克隆及表达分析[J]. 中国生物工程杂志, 2016, 36(4): 88-96.

WAN Yong-qing, YANG Yang, ZHANG Chun-lin, WAN Dong-li, YANG Ai-qin, YANG Qi, WANG Rui-gang, LI Guo-jing. Cloning and Expression Analysis of CiDHN1 Gene in Caragana intermedia. China Biotechnology, 2016, 36(4): 88-96.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20160413 或 https://manu60.magtech.com.cn/biotech/CN/Y2016/V36/I4/88

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