海州香薷不同抗性种群液泡转化酶基因EhvINV序列趋异及表达差异分析

doi:10.13560/j.cnki.biotech.bull.1985.2016.10.021

生物技术通报 ›› 2016, Vol. 32 ›› Issue (10): 170-179.doi: 10.13560/j.cnki.biotech.bull.1985.2016.10.021

海州香薷不同抗性种群液泡转化酶基因EhvINV序列趋异及表达差异分析

蔡深文^{1, 3}, 徐仲瑞², 熊治廷^{2, 3}, 王加真⁴, 陈瑶⁴

1. 遵义师范学院资源与环境学院,遵义 563002;
2. 武汉大学资源与环境科学学院,武汉 430079;
3. 生物质资源化学与环境生物技术湖北省重点实验室,武汉 430079;
4. 遵义师范学院生命科学学院,遵义 563002

收稿日期:2016-04-19 出版日期:2016-10-25 发布日期:2016-10-12
作者简介:蔡深文,男,博士,副教授,研究方向：环境生物学;E-mail：caishenwen@163.com
基金资助:
国家自然科学基金项目（21477093,31270432）,生物质资源化学与环境生物技术湖北省重点实验室开放基金项目（HBRCEBL-2013-2014004）,遵义师范学院博士基金项目（遵师BS[2014]20号）

Sequence Divergence and Analysis of Expression Difference of Vacuolar Invertase Gene EhvINV from Different Resistant Populations in Elsholtzia haichowensis

CAI Shen-wen^{1, 3}, XU Zhong-rui², XIONG Zhi-ting^{2, 3}, WANG Jia-zhen⁴, CHEN Yao⁴

1. College of Resources and Environment,Zunyi Normal College,Zunyi 563002;
2. School of Resource and Environmental Sciences,Wuhan University,Wuhan 430079;
3. Hubei Biomass-Resource Chemistry and Environmental Biotechnology Key Laboratory,Wuhan 430079;
4. College of Life Science,Zunyi Normal College,Zunyi 563002

Received:2016-04-19 Published:2016-10-25 Online:2016-10-12

摘要/Abstract

摘要： 根据GenBank中海州香薷（Elsholtzia haichowensis）液泡转化酶基因EhNvINV（JX500755）和EhCvINV（JX500756）的序列设计特异性引物,克隆cDNA全长序列,并通过生物信息学分析基因及推导的蛋白序列,利用SWISS-MODEL进行同源建模,并与蔗糖分子进行模拟对接,实时荧光定量PCR分析EhNvINV和EhCvINV在铜胁迫下的转录表达。结果表明,海州香薷非抗性和抗性种群液泡转化酶蛋白EhNvINV和EhCvINV在114和346处存在氨基酸趋异位点,模拟3D结构相似,仅在趋异位点Glu114/Gln114和Leu346/Pro346处有差别。EhNvINV、EhCvINV和模拟突变体（EhNvINV-E114Q和EhNvINV-L346P）与蔗糖分子对接形成的活性中心构象基本一致,但在空间位置上存在细微差异。实时荧光定量PCR结果表明铜胁迫7 d后,EhCvINV的表达受铜的诱导,而EhNvINV受铜的抑制。

关键词: 海州香薷, 液泡转化酶, 铜胁迫, 转录表达

Abstract: Specific primers were designed to clone the cDNA sequences according to the EhNvINV（JX500755）and EhCvINV（JX500756）from GenBank. The DNA and deduced amino acid sequences were analyzed by bioinformatics methods. The three-dimensional structures were constructed by homologous modeling. The structures of vacuolar invertase from two populations of E. haichowensis and non-tolerant population with each single point mutation in complex with sucrose were simulated by AutoDock 4.0. Transcript expression of EhNvINVand EhCvINV under copper tress was analyzed by real-time PCR. The results showed that there were two divergent amino acids at position 114 and 346 between EhNvINV and EhCcINV. The three-dimensional structures were exactly similar between EhNvINV and EhCvINV. It showed difference at Glu114/Gln114 and Leu346/Pro346,which were divergent sites. The structures of catalytic active center of EhNvINV,EhCvINV,and simulated mutants（EhNvINV-E114Q,EhNvINV-L346P）binding with sucrose showed no significant differences. However,there were differences on spatial position. The result of real-time PCR indicated that the transcript expression of EhCvINV induced by copper stress after 7 days,however,the transcript expression of EhNvINV inhibited by copper stress after 7 days.

Key words: Elsholtzia haichowensis, vacuolar invertase, copper stress, transcript expression

蔡深文, 徐仲瑞, 熊治廷, 王加真, 陈瑶. 海州香薷不同抗性种群液泡转化酶基因EhvINV序列趋异及表达差异分析[J]. 生物技术通报, 2016, 32(10): 170-179.

CAI Shen-wen, XU Zhong-rui, XIONG Zhi-ting, WANG Jia-zhen, CHEN Yao. Sequence Divergence and Analysis of Expression Difference of Vacuolar Invertase Gene EhvINV from Different Resistant Populations in Elsholtzia haichowensis[J]. Biotechnology Bulletin, 2016, 32(10): 170-179.

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海州香薷不同抗性种群液泡转化酶基因EhvINV序列趋异及表达差异分析

Sequence Divergence and Analysis of Expression Difference of Vacuolar Invertase Gene EhvINV from Different Resistant Populations in Elsholtzia haichowensis

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