COX-2诱饵载体的构建及其在酵母双杂交系统中自激活作用的检测

首都医科大学学报 ›› 2011, Vol. 32 ›› Issue (1): 106-109.

COX-2诱饵载体的构建及其在酵母双杂交系统中自激活作用的检测

贺凌婕¹，张澍田^2*，朱圣韬²

1. 首都医科大学附属北京友谊医院急诊科； 2. 首都医科大学附属北京友谊医院消化内科，北京市消化疾病中心

收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2011-02-21 发布日期:2011-02-21
通讯作者: 张澍田

Construction and Identification of Yeast Two-hybrid Bait Vector pGBKT7COX-2

HE Ling-jie¹, ZHANG Shu-tian^2*, ZHU Sheng-tao²

1. Department of Emergency, Beijing Friendship Hospital, Capital Medical University; 2. Department of Digestive Diseases,Beijing Friendship Hospital, Capital Medical University, Beijing Digestive Diseases Center

Received:1900-01-01 Revised:1900-01-01 Online:2011-02-21 Published:2011-02-21
Contact: ZHANG Shu-tian

摘要/Abstract

摘要：

目的构建环氧化酶-2（cyclooxygenase-2，COX-2）的诱饵载体，为应用酵母双杂交系统(yeast two-hybrid system)筛选与COX-2相互作用的蛋白建立实验基础。
方法 PCR扩增COX-2基因编码区全长，引入酶切位点EcoRⅠ、BamHⅠ，将COX-2基因克隆至载体pGBKT7中，将构建好的诱饵载体pGBKT7-COX-2转化到酵母细胞Y190中，检测pGBKT7-COX-2在Mathmaker GAL4 Two-Hybrid System 3中有无自激活作用。
结果成功扩增了COX-2基因编码区全长，并克隆入pGBKT7中，测序结果正确，并成功转化到酵母细胞Y190中，检测无自激活作用。
结论成功构建了COX-2的酵母诱饵载体，为进一步运用酵母双杂交技术筛选与之相互作用的蛋白奠定了基础。

关键词: 环氧化酶-2, 酵母双杂交系统, 自激活

Abstract:

Objective To construct the bait vector of cyclooxygenase-2(COX-2) in yeast two-hybrid system 3, and to test it for its autonomous activation.
Methods COX-2 gene was amplified by polymerase chain reaction(PCR) from cyclooxygenase-2(COX-2) plasmid, and the EcoRⅠ and BamHⅠ sites were introduced into it. The COX-2 gene, after sequenced, was cloned into pGBKT7. These recombinant plasmids were transferred into Y190 and they activated the reporter genes.
Results The pGBKT7-COX-2 bait vector was successfully constructed and was identified that it had no autonomous activation.
Conclusion COX-2 gene has no self-activation, bait vector could be successfully constructed, the proteins interacting with COX-2 can be detected by the yeast two-hybrid system 3.

Key words: cyclooxygenase-2, yeast two hybrid system, autonomous activation

中图分类号:

Q 782

贺凌婕;张澍田;朱圣韬. COX-2诱饵载体的构建及其在酵母双杂交系统中自激活作用的检测[J]. 首都医科大学学报, 2011, 32(1): 106-109.

HE Ling-jie;ZHANG Shu-tian;ZHU Sheng-tao. Construction and Identification of Yeast Two-hybrid Bait Vector pGBKT7COX-2[J]. Journal of Capital Medical University, 2011, 32(1): 106-109.

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