人源M2二联体靶抗原BO-E2融合蛋白的克隆表达与鉴定
原发性胆汁性肝硬化;二氧酸脱氢酶复合体E2;2-氧戊二酸脱氢酶复合体E2;BO-E2融合蛋白;蛋白印迹,周晔,姚定康,陈燕,蒋天舒,陈波,仲人前,邓安梅,通讯作者:,CDNAcloningandexpressionofautoantigenB
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周晔, 姚定康, 陈燕, 蒋天舒, 陈波, 仲人前, 邓安梅, 上海长征医院实验诊断科 上海市 200003
国家自然科学基金项目, No. 30471616
通讯作者: 邓安梅, 200003, 上海市凤阳路415号, 长征医院检验科. amdeng@yahoo.com
电话: 021-63583108
收稿日期: 2007-03-01 修回日期: 2007-07-22
CDNA cloning and expression of autoantigen B0-E2
Ye Zhou, Ding-Kang Yao, Yan Chen, Tian-Shu Jiang, Bo Chen, Ren-Qian Zhong, An-Mei Deng
Ye Zhou, Ding-Kang Yao, Yan Chen, Tian-Shu Jiang, Bo Chen, Ren-Qian Zhong, An-Mei Deng, Labortary of Diagnostics, Shanghai Changzheng Hospital, Shanghai 200003, China
Supported by: National Natural Science Foundation of China, No. 30471616
Correspondence to: An-Mei Deng, Labortary of Diagnostics, Shanghai Changzheng Hospital, 415 Fengyang Road, Shanghai 200003, China. amdeng@yahoo.com
Received: 2007-03-01 Revised: 2007-07-22
Abstract
AIM: To express BO-E2 recombinant fusion protein in Escherichia coli for use in early diagnosis of primary biliary cirrhosis (PBC).
METHODS: CDNA encoding BCOADC-E2 and OGDC-E2 were obtained by reverse transcription pdymerase chain reaction (RT-PCR), confirmed by DNA sequencing, subcloned non-directionally into the bacterial expression plasmid pET28a(+), and then transformed into E. coli BL21(DE3) to express the recombinant fusion protein, which was identified by SDS-PAGE and western blotting.
RESULTS: Results of nucleotide sequencing and restriction analysis showed that the recombinant plasmid successfully constructed three species. Three types of fusion protein were expressed by isopropyl-Beta-d-thiogalactopyranoside (IPTG) induction. The recombinant fusion protein exhibited the antigenicity of AMA-M2 by western blotting.
CONCLUSION: BO-E2 recombinant fusion protein can be used for in vitro diagnosis of PBC.
Key Words: Primary biliary cirrhosis; BCOADC-E2; OGDC-E2; BO-E2 recombinant fusion protein; Western blotting ......
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