[1]刘晓兵,南海洋,袁晓辉,等.大豆GmbZIP基因的克隆及EMSA分析[J].大豆科学,2015,34(01):32-35,41.[doi:10.11861/j.issn.1000-9841.2015.01.0032]
　LIU Xiao-bing,NAN Hai-yang,YUAN Xiao-hui,et al.Expression Analysis and EMSA of Soybean GmbZIP71 Gene[J].Soybean Science,2015,34(01):32-35,41.[doi:10.11861/j.issn.1000-9841.2015.01.0032]
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大豆GmbZIP基因的克隆及EMSA分析

《大豆科学》[ISSN:1000-9841/CN:23-1227/S] 卷: 第34卷 期数: 2015年01期 页码: 32-35,41 栏目: 出版日期: 2015-02-25
Title:
Expression Analysis and EMSA of Soybean GmbZIP71 Gene
作者:
刘晓兵1; 南海洋2; 袁晓辉2; 刘宝辉2; 孔凡江2
1.黑龙江省种子管理局，黑龙江 哈尔滨 150008；
2.中国科学院 东北地理与农业生态研究所/大豆分子设计育种重点实验室，黑龙江 哈尔滨 150081
Author(s):
LIU Xiao-bing1;  NAN Hai-yang2;  YUAN Xiao-hui2;  LIU Bao-hui2;  KONG Fan-jiang2
1.The Seed Management Bureau of Heilongjiang Province, Harbin 150008, China;?
2.Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences/The Key Laboratory of Soybean Molecular Design Breeding, Harbin 150008, China
关键词:
大豆; bZIP; 原核表达; EMSA
Keywords:
Soybean;  bZIP;  Prokaryotic expression;  EMSA
分类号:
S565.1
DOI:
10.11861/j.issn.1000-9841.2015.01.0032
文献标志码:
A
摘要:
通过PCR扩增的方法从大豆品种Harosoy中克隆到1个新的bZIP基因（GmbZIP71），基因表达分析表明GmbZIP71在叶片、生长点、花、花芽、荚和根等多个器官中表达。将GmbZIP71构建到原核表达载体pET29b上，导入大肠杆菌BL21（DE3）中，对其进行IPTG诱导。结果表明：在IPTG浓度为0.25 mmol?L1，诱导时间为4 h，诱导温度为28℃时，重组蛋白得到表达，分子量大约为48 kDa，SDS-PAGE电泳结果表明重组蛋白主要以包涵体形式存在，用His蛋白纯化系统回收得到GmbZIP71-His重组蛋白，EMSA（electrophoretic mobility shift assay）实验表明GmbZIP71重组蛋白可以在体外与ACGT顺式作用元件结合。
Abstract:
?Using PCR amplification method, we cloned a new bZIP homolog which named GmbZIP71 in soybean cultivar Harosoy, RT-PCR showed that GmbZIP71 transcripted in multiple organs such as leaves, shoot apices, flowers, flower buds, pods and roots. GmbZIP71 was constructed into prokaryotic expression vector pET29b and then transformed into E. coli BL21 (DE3) to get the expression protein for further study. The results indicated that an 48 kDa recombinant protein was expressed with the treatment of 0.25 mmol?L1?IPTG for 4 h at 28℃, the recombinant protein was confirmed to be mainly existed in inclusion body form by SDS-PAGE. The high-quality recombinant protein was obtained through His-tag nickle column purification system and the EMSA experiment indicated that GmbZIP71 protein could bind with ACGT ciselement in vitro.

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备注/Memo

基金项目：国家自然科学基金（31071445，31171579，31201222和31371643）；黑龙江省自然科学基金（ZD201001，JC201313）；中国科学院“百人计划”。

第一作者：刘晓兵（1974-），男，高级农艺师，主要从事种子生物学研究。E-mail：13936161608@163.com。
通讯作者：孔凡江（1972-），男，博士，研究员，博导，主要从事大豆分子遗传学研究。E-mail:kongfj@iga.ac.cn。

更新日期/Last Update: 2015-04-12