核桃V-ATPase c亚基基因(JrVHAc4)的克隆和抗旱功能分析

doi:10.3969/j.issn.1000-2006.201811014

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南京林业大学学报（自然科学版） ›› 2019, Vol. 62 ›› Issue (02): 79-85.doi: 10.3969/j.issn.1000-2006.201811014

核桃V-ATPase c亚基基因(JrVHAc4)的克隆和抗旱功能分析

李大培^{1, 2}, 王艺¹, 张尚昆¹, 赵翔^{1, 2},赵焕元^{1, 2}, 刘玉梅^{1, 2},杨桂燕^{1, 2*}

(1.西北农林科技大学林学院,山阳核桃板栗试验示范站,陕西杨凌 712100; 2.西北农林科技大学林学院,陕西省经济植物资源开发利用重点实验室,陕西杨凌 712100)

出版日期:2019-03-30 发布日期:2019-03-30
基金资助:
收稿日期:2018-11-06 修回日期:2018-12-22
基金项目:国家自然科学基金项目(31800510); 陕西省自然科学基础研究计划项目(2018JQ3066); 大学生创新创业训练计划项目(201803072)。
第一作者:李大培(deplee1021@gmail.com)。
*通信作者:杨桂燕(yangguiyan@yahoo.com),讲师,ORCID(0000-0002-7669-6951)。

Drought resistance analysis of a V-ATPase c subunit(JrVHAc4)gene from Juglans regia

LI Dapei^1,2,WANG Yi ¹,ZHANG Shangkun ¹,ZHAO Xiang^1,2,ZHAO Huanyuan ^1,2,LIU Yumei^1,2,YANG Guiyan^{1,2 *}

(1. Walnut and Chestnut Experiment Station of Shanyang, College of Forestry, Northwest A & F University,Yangling 712100, China; 2. Key Laboratory of Economic Plant Resources Development and Utilization in Shaanxi Province, College of Forestry, Northwest A & F University, Yangling 712100, China)

Online:2019-03-30 Published:2019-03-30

摘要/Abstract

摘要： 【目的】V-ATPase是植物逆境响应的重要酶,有必要了解V-ATPase相关亚基参与核桃响应逆境胁迫的功能机制。【方法】从‘香玲’核桃转录组中克隆获得1条V-ATPase c 亚基基因(命名为JrVHAc4),通过分析启动子预测其逆境响应功能。对JrVHAc4基因进行干旱胁迫下的表达分析,同时构建JrVHAc4酵母表达载体转入酵母表达系统研究其抗旱功能。【结果】JrVHAc4基因开放读码框(ORF)全长495 bp,拟推导的蛋白分子量为16 527.61 u,包含164个氨基酸,理论等电点为8.62; 其上游1 047 bp启动子包含MYC、DOF、MYB等与干旱响应相关的顺式作用元件。干旱胁迫下,JrVHAc4基因在核桃叶和根中均被显著诱导,并存在差异。对JrVHAc4转基因酵母进行不同浓度甘露醇胁迫,与对照酵母相比,发现JrVHAc4基因的表达能显著提高转基因酵母的生长活性。【结论】JrVHAc4基因能响应干旱胁迫,并能提高酵母的抗旱能力,JrVHAc4可作为核桃抗逆重要候选基因。

Abstract: 【Objective】 V-ATPase is an important enzyme in plant stress response. This study is under stunll the functional mechanism of V-ATPase subunits in response to stress in walnuts. 【Method】 A V-ATPase c subunit gene(named JrVHAc4)was cloned from the ‘Xiangling’ walnut transcriptome. The stress response function was predicted by analyzing its promoter and expression under drought stress. Meanwhile, JrVHAc4 was inserted into a yeast expression vector pYES2 and transformed into yeast INVSC1 to study its drought resistance function. 【Result】 The open reading frame(ORF)of JrVHAc4 was 495 bp in length, the deduced protein was 16 527.61 u and contained 164 amino acids, whose theoretical isoelectric point(pI)was 8.62. An upstream 1 047 bp promoter segment was identified and found to contain abundant cis-elements, among which some are related to drought response, such as MYC, DOF and MYB. JrVHAc4 was significantly induced in leaves and roots by drought stress, and the induction in leaves and roots were different. Compared with control yeast, we found that expression of JrVHAc4 increased the growth activity of transgenic yeast significantly under different concentrations of mannitol stress. 【Conclusion】 JrVHAc4 can respond to drought stress and improve the drought resistance of transgenic yeasts, JrVHAc4 can be used as an important candidate gene for walnut resistance.

中图分类号:

Q786

李大培,王艺,张尚昆,等. 核桃V-ATPase c亚基基因(JrVHAc4)的克隆和抗旱功能分析[J]. 南京林业大学学报（自然科学版）, 2019, 62(02): 79-85.

LI Dapei,WANG Yi ,ZHANG Shangkun ,ZHAO Xiang^1,2,ZHAO Huanyuan ,LIU Yumei^1,2,YANG Guiyan^{1,2 *}. Drought resistance analysis of a V-ATPase c subunit(JrVHAc4)gene from Juglans regia[J].Journal of Nanjing Forestry University (Natural Science Edition), 2019, 62(02): 79-85.DOI: 10.3969/j.issn.1000-2006.201811014.

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核桃V-ATPase c亚基基因(JrVHAc4)的克隆和抗旱功能分析

Drought resistance analysis of a V-ATPase c subunit(JrVHAc4)gene from Juglans regia

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