人整合素分子α4亚基片段的克隆及大肠杆菌的表达

Cloning of the fragment of α4 subunit of human integrin and it’s expression from E.coli

目的克隆并原核表达 α4亚基片段。方法从 IL- 6 0细胞系中提取总 RNA,以 RT- PCR方法分两段扩增人整合素分子 α4亚基片段 2 .0 kb c DNA,将两片段连接 ,并将其中的 1.7kb基因片段亚克隆至表达载体 PGEX- KG中 ,IPTG诱导表达。结果 RT- PCR扩增并克隆了α4的两段 c DNA,序列分析表明 :与文献报道的α4c DNA序列基本一致。两片段成功连接后 ,将其中的 1.7kb亚克隆至表达载体 PGEX- KG中 ,经诱导在大肠杆菌中得到高效表达。结论获得了α4亚基 2 .0 kbc DNA片段 ,及其中 1.

ObjectiveTo clone and express the fragment of α4 subunit.MethodsThe 2.0kb cNDA of human integrin α4 subunit fragment was amplified from HL60 total RNA by RTPCR as two seperate segments,after the two segments were linked together,1.7kb of it was subcloned into expression vector PGEXKG and induced by IPTG.ResulstThe sequence of our cDNA is identical with the reported α4 cDNA Then the two segments were linked together, and 17kb of it was subcloned into expression vector PGEXKGThe α4 fragment was overexpressed by Ecoli after inducing with IPTGConclusionThe 2.0kb cDNA fragment of α4 subunit.was obtained,and 1.7kb of it was overexpressed by E.coil,it’s very important to study the function of α4 integrins. [