人免疫球蛋白轻链恒区基因的体外扩增及其在大肠杆菌中的分泌表达

PCR-AMPLIFICATION AND SECRETION EXPRESSION OF HUMAN IMMUNOGLOBULIN KAPPA CHAIN CONSTANT REGION

本文应用聚合酶链反应(PCR)技术体外大量扩增人免疫球蛋白轻链恒区基因(人C_k基因),并将该基因克隆至分泌型表达载体pINⅢ-ompA2,转入大肠杆菌中表达。应用水休克法抽提大肠杆菌的周质蛋白,经SDS-PAGE电泳,免疫印迹法分析,结果表明人恒区基因在大肠杆菌中得到表达,并能分泌至细菌的周质空间。约50％的表达蛋白其ompA信号肽已被切除。

In this paper, human immunoglobulin kappa chain constant region (human C_kgene) was amplified by PCR with a set of primers and cloned into secretion expressionvector pINIII-ompA2 in frame, then expressed in three kinds of E. coli strains. SDS-PAGE and immun-blot analysis of periplasmic proteins in E. coli, which wereextracted by osmotic shock showed that the expressed product of human C_k gene wassecreted to bacteria periplasmic space and about 50% of them was without ompAleader. This work will favour tke construction of human/mouse chimeric antibody inEacherichia coli.