人整合素分子α4亚基片段的克隆和表达  

Cloning and expression of the 1.2kb cDNA fragment of human integrin α4 subunit

作　　者：刘永全[1] 高杰英[1] 罗振革[1] 孔祥英[1] 梅俊杰[1] 彭虹[1] 

机构地区：[1]军事医学科学院微生物流行病研究所免疫室,北京市太平路27号北京100850

出　　处：《细胞与分子免疫学杂志》2001年第1期13-15,共3页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金资助项目!No.39789010

摘　　要：目的克隆并表达人整合素分子α４亚基１．２ｋｂｃＤ－ＮＡ片段。方法从 ＨＬ－６０细胞系中提取总 ＲＮＡ，以 ＲＴ－ ＰＣＲ法扩增人整合素分子 α４亚基片段 １． ２ ｋｂ ｃＤＮＡ（１７５３－ ２ ９３４ｂｐ），并将该片段亚克隆至表达载体ｐＧＥＸ－３Ｘ，用ＩＰＴＧ（诱导表达。结果克隆了 α４亚基 １．２ ｋｂ ｃＤＮＡ片段。序列分析表明，其所编码的氨基酸序列与文献相比较只有一处错义突变（Ａｒｇ→Ｇｌｎ），经分析该突变对抗原性影响不大。将该片段亚克隆至表达载体ｐＧＥＸ－３Ｘ，经ＩＰＴＧ诱导在大肠杆菌中获得高效表达。结论获得了α４亚基１．２ｋｂｃＤＮＡ片段及其原核表达产物，对研究α４整合素的功能具有重要的意义。Aim To clone and express the 1. 2kb cDNA fragment (1/753-2 934bp) of human integrin α4 subunit. Methods The 1. 2 kb cDNA fragment of human integrin α4 subunit was amplified from HL-60 total RNA by AT-PCR, then it was subcloned into expression vector pGEX-3X and induced with IPTG. Results The 1. 2 kb cDNA fragment of human integrin α4 subunit was cloned. The sequencing indicated that there was only one missense mutation (Arg→Gln) among the fragment, and this mutation won't affect antigenicity after analysed by GOLDKEY. Then the 1. 2 kb cDNA was subcloned into expression vector pGEX-3X. The α4 fragment was highexpressed in E. coli after induced with IPTG. Conclusion The 1. 2kb cDNA fragment of α4 subunit was obtained, and it was highexpressed in E. coli, it might be important for study on the function of α4 integrins.

关 键 词：整合素分子 α4亚基 RT-PCR CDNA 克隆 表达 

分 类 号：R392.11[医药卫生—免疫学] Q785[医药卫生—基础医学]