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作物学报

作物学报 ›› 2009, Vol. 35 ›› Issue (6): 1156-1160.doi: 10.3724/SP.J.1006.2009.01156

• 研究简报 • 上一篇    下一篇

表达大豆GMCHI基因能够提高原核生物对耐低温性

程立宝1，李淑艳3，景新明2，何光源1,*   

  1. 1华中科技大学中英HUST-RRes基因工程和基因组学联合实验室，教育部分子生物物理重点实验室，生命科学与技术学院，湖北武汉430074；2中国科学院植物研究所植物园，北京100093；3孝感学院生命科学与技术学院，湖北孝感432000
  • 收稿日期:2008-11-05 修回日期:2009-01-28 出版日期:2009-06-12 网络出版日期:2009-03-23
  • 通讯作者: 何光源，E-mail:hegy@hust.edu.cn；Tel：027-87792271
  • 作者简介:程立宝，E-mail:chenglibao453@163.com；Tel:027-87792271
  • 基金资助:

    本研究由国家重点基础研究发展计划（973计划）项目（2002CB111302）资助。

Expression of GMCHI gene，isolated from Soybean，Enhances the Survival in Prokaryotes to Low Temperature Stress

CHENG Li-Bao1,LI Shu-Yan2,HE Guang-Yuan1*   

  1. 1China-UK HUST-RRes Genetics Engineering and Genomics Joint Laboratory,Huazhong University of Science Technology,Wuhan 430074,China；2Institute of Botany，Chinese Academy of Sciences，Beijing 100093，China；3Life Science and Technique College of Xiaogan University，Xiaogan432000，China
  • Received:2008-11-05 Revised:2009-01-28 Published:2009-06-12 Published online:2009-03-23
  • Contact: HE Guang-Yuan，E-mail:hegy@hust.edu.cn；Tel：027-87792271
  • About author:CHEN Li-Bao，E-mail:chenglibao453@163.com；Tel:027-87792271

PDF

1575

被引次数

2

摘要：

低温是种子萌发过程中常见的自然灾害，在我国北方经常发生，导致种子萌发率较差、降低植株活力等后果。利用cDNA-AFLP技术从低温(4℃)吸胀24 h的抗低温吸胀大豆品种中黄22 (低温吸胀24 h对萌发率无影响的品种)中分离出一个基因片段，命名为GMCHI (GenBank No. EU699765)，通过RACE方法得到全长为387 bp的cDNA序列。在NCBI数据库中的查询表明， GMCHI基因和数据库记录的基因序列同源性较低，因此可以断定GMCHI是在大豆中被发现的新基因。半定量RT-PCR显示GMCHI受ABA和PEG诱导。该基因与PET30A连接后转入原核细胞，经过IPTG诱导，6% SDS-PAGE电泳条带说明，GMCHI在大肠杆菌中能够表达。把诱导表达和非诱导表达的菌落在–20℃下2 h后，移至37℃培养20 d，发现对照的菌落完全死亡，而诱导表达GMCHI的菌落只有部分死亡，并长出新菌落。RT-PCR检测表明新菌落携带GMCHI基因，证明GMCHI基因在大肠杆菌中的表达提高了低温耐性。

关键词: 大豆, GMCHI, 菌落, cDNA-AFLP, 基因

Abstract:

Low temperature during germination being a natural disaster usually occurs in North of China, which results in low germination rate and low seedling vigor, and even ultimately severe loss in yield. A gene was isolated from soybean Zhonghuang 22 cultivar (tolerant to low temperature in imbibition) for 24 h under 4℃ via cDNA-AFLP, name as GMCHI, GenBank accession No. EU699765. The full-length sequence of GMCHI which was 387 bp in nucleotide was obtained by RACE method. The sequence of gene shared low similarity with that of genes documented in NCBI, we inferred that GMCHI was a new discovered gene in soybean. Semi quantitative RT-PCR result revealed that GMCHI was induced by ABA and IPEG.. The gene ligated with Prokaryotic expression vector was transformed into prokaryotic cell, the result of 6% SDS-PAGE induced by IPTG was expressed in Prokaryotes. The E. coli carrying GMCHI survived and showed tolerance to low temperature under the treatment of –20℃ for 2 h. RT-PCR validated that GMCHI in Prokaryotes improved the adaptation to low temperature.

Key words: Soybean, GMCHI, E.coli, cDNA-AFLP, Gene


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