细胞自身分化抗原-1启动子区域的克隆及其特性分析

doi:中国-澳大利亚政府互换项目奖学金资助课题

细胞自身分化抗原-1启动子区域的克隆及其特性分析

韩淑华^1,2，陈丽萍³，迟宝荣¹

1.东南大学中大医院呼吸内科，江苏南京 210009；2.吉林大学第一医院消化内科, 吉

收稿日期:2007-06-20 修回日期:1900-01-01 出版日期:2008-05-28 发布日期:2008-05-28
通讯作者: 迟宝荣

Cloning and characterization of promoter region of cell division autoantigen 1

HAN Shu-hua^1,2, CHEN Li-ping³, CHI Bao-rong²

1.Deparment of Respiratory Medicine, Zhongda Hospital,Southeast University,Nanjing 210009, China;2.Department of Digestive Medicine, First Hospital,Jilin University, Changchun 130021,China;3.Department of Electrodiagnosis ,First Hospital, Jilin University, Changchun 130021,China

Received:2007-06-20 Revised:1900-01-01 Online:2008-05-28 Published:2008-05-28
Contact: CHI Bao-rong

摘要/Abstract

摘要： 目的：克隆不同长度的细胞自身分化抗原-1（CDA1)启动子区域并测定它们的活性，为进一步研究不同长度细胞自身分化抗原-1启动子（CDA1P）区域的功能打下基础。方法：以小鼠肝脏的全基因序列为模板，用PCR方法获得不同长度的目的片段，连接到pGL3-basic真核表达载体（pGL3-basic-mCDA1P），纯化pGL3-basic mCDA1P质粒后，瞬时转染到Leuwis肺癌细胞和单核巨噬细胞(RAW264.7)，48 h后收集转染细胞，测定萤光素酶活性，明确不同长度的CDA1P的活性。结果：①通过PCR法成功获得不同长度的CDA1P DNA片段，并用酶切法证实；②成功构建携带有CDA1P的pGL3-basic真核表达载体，并通过酶切法及测序证实； ③转染Leuwis肺癌细胞和单核巨噬细胞后测定萤光素酶活性发现不同长度的CDA1P活性不同，相同长度的CDA1P在Leuwis肺癌细胞和单核巨噬细胞中活性亦不相同。结论：CDA1P在不同细胞中活性可能存在差异。

关键词: 启动子, pGL3-basic真核表达载体

Abstract: Abstract:Objective To clone different lengths cell division autoantigen1( CDA1) promoters(CDA1P) and measure their activities in order to provide basis for fumctional study of CDA1P. Methods Different lengths CDA1P were amplified by PCR using mouse liver genomic as template and cloned into BglⅡ digested pGL3-basic plasmid to produce recombinant plasmid pGL3-basic-mCDA1P, then they were transfected into Lewis lung cell line (LLC) and RAW 264.7 cells differently, these cells were collected after transfected for 48 h, finally luciferase activities of pGL3-basic-mCDA1P were measured. Results ①Different lengths CDA1P were cloned and characterized by restriction enzymes. ②Different lengths CDA1P were ligated into pGL3-basic vector,and were proved with PCR and DNA sequencing .③Activities of pGL3-basic-mCDA1A were different in the two kinds of Leuwis and mononuclear macrophage cells (RAW264.7 cells). Conclusion The different activities of CDA1P may be related with different cellular types.

Key words: promoter, pGL3-basic plasmid

中图分类号:

韩淑华，陈丽萍，迟宝荣. 细胞自身分化抗原-1启动子区域的克隆及其特性分析[J]. J4, 2008, 34(3): 420-424.

HAN Shu-hua, CHEN Li-ping, CHI Bao-rong. Cloning and characterization of promoter region of cell division autoantigen 1[J]. J4, 2008, 34(3): 420-424.

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