人自身抗原CYP2D6 285 bp基因片段克隆、真核表达及免疫性鉴定
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王文凯, 李永哲, 刘国振
自身抗原;融合基因;细胞色素P4502D6;显性表位王文凯,李永哲,刘国振.人自身抗原CYP2D6285bp基因片段克隆、真核表达及免疫性鉴定. 世界华人消化杂志2006;14(14)13
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王文凯, 李永哲, 中国医学科学院中国协和医科大学 北京协和医院检验科 北京市 100730
刘国振, 中国科学院北京华大基因研究中心 北京市 101300
王文凯, 2003年北华大学医学院硕士研究生, 主要从事自身免疫性疾病研究.
国家自然科学基金资助项目, No. 30471617
国家863计划重大专项基金资助项目, No. 2002AARZ2011
通讯作者: 李永哲, 100730, 北京市, 中国医学科学院中国协和医科大学北京协和医院检验科. yongzhelipumch@yahoo.com.cn
电话: 010-65295416 传真: 010-65295416
收稿日期: 2006-01-11 接受日期: 2006-02-21
Expression of Cyclin D1, proliferating cell nuclear antigen in liver of C57BL/6 mice infected with Helicobacter pylori
Wen-Kai Wang, Yong-Zhe Li, Guo-Zhen Liu
Wen-Kai Wang, Yong-Zhe Li, Department of Clinical Laboratory, Peking Union Medical College Hospital, PUMC & CAMS, Beijing 100730, China
Guo-Zhen Liu, Huada Genomics Research Center, Chinese Academy of Sciences, Beijing 101300, China
Supported by
Correspondence to: Yong-Zhe Li, Department of Clinical Laboratory, Peking Union Medical College Hospital, PUMC & CAMS, Beijing 100730, China. yongzhelipumch@yahoo.com.cn
Received: 2006-01-11 Accepted: 2006-02-21
Abstract
AIM: To clone and express immunodominant epitope of human autoantigen cytochrome P450 2D6 (CYP2D6) in Saccharomyces Cerevisiae, and establish a new assay for detecting autoantibody LKM-1.
METHODS: We obtained CYP2D6 DNA fragment by polymerase chain reaction (PCR), using total liver cDNA library as the template. The PCR products were recombined into pEGH expression vector to construct the high efficiency recombinant expression vector in Saccharomyces Cerevisiae Y258. The positive clones were identified by PCR and induced by galactose. Glutathione-Sepharose 4B was used for purification of recombinant CYP2D6 protein. The expression products were analyzed by SDS-PAGE and Western blot as well as by matrix-assisted laser desorption inoization-time of flight mass spectrometry (MALDI-TOF-MS.
RESULTS:The PCR product was about 290 bp in size, which was in accordance with the predicted 285 bp ......
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