胃癌相关基因GCRG213正反义真核表达载体的构建及鉴定
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高利利, 吴本俨, 王孟薇, 黄海力, 伍银桥,
胃癌相关基因GCRG213;真核表达;胃癌细胞;基因转染,高利利,吴本俨,王孟薇,黄海力,伍银桥,尤纬缔,王卫华,高利利,通讯作者,Const
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高利利, 吴本俨, 王孟薇, 黄海力, 伍银桥, 尤纬缔, 王卫华, 解放军总医院南楼消化科 北京市 100853
高利利, 2005年首都医科大学博士毕业, 主治医师, 主要从事消化道肿瘤和动力的临床及基础研究.
国家自然科学基金资助项目, No. 30370635
通讯作者: 吴本俨, 100853, 北京市复兴路28号, 解放军总医院南楼消化科.
电话: 010-66939443
收稿日期: 2006-02-13 接受日期: 2006-03-20
Construction and identification of eukaryotic expression vector of gastric cancer associated gene GCRG213
Li-Li Gao, Ben-Yan Wu, Meng-Wei Wang, Hai-Li Huang, Yin-Qiao Wu, Wei-Di You, Wei-Hua Wang
Li-Li Gao, Ben-Yan Wu, Meng-Wei Wang, Hai-Li Huang, Yin-Qiao Wu, Wei-Di You, Wei-Hua Wang, Department of Gastroenterology, General Hospital of Chinese PLA, Beijing 100853, China
Supported by National Science Foundation of China, No. 30370635
Correspondence to: Dr. Wu-Ben Yan, Department of Gastroenterology, South Building, General Hospital of Chinese PLA, Beijing 100853, China
Received: 2006-02-13 Accepted: 2006-03-20
Abstract
AIM: To construct and identify the gastric cancer associated gene GCRG213 eukaryotic expression vector.
METHODS: The sense and anti-sense fragment of GCRG213 were obtained by polymerase chain reaction (PCR), which were GCRG213-a and GCRG213-b respectively. They were cloned into eukaryotic expression vector pcDNA3.1(+) after introduced the sites of restrictive endonuclease enzyme KpnI, BamHI and EcoRI, BamHI. The recombinant plasmid pcDNA3.1-a, pcDNA3.1-b and the vector pcDNA3.1 were transfected separately into MKN45 cells conducted by lipofectamineTM 2000. G418 selection was used to obtain the stably transfected cells. The expression of GCRG213 was detected at both mRNA and protein level with semi-quantitative reverse transcription PCR (RT-PCR) and Western blot, respectively.
RESULTS: After sequencing, sense GCRG213 and anti-sense GCRG213 were confirmed to be successfully cloned into eukaryotic expression vector pcDNA3.1, which were named pcDNA3 ......
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