鸡BLB重组蛋白的原核表达及多抗制备  

Preparation of antiserum against chicken BLB protein with recombinant BLB expressed in E. coli

作　　者：李行[1] 高彩霞[1,2] 蔡文博[1] 杨柳[1,3] 武永淑[1] 张伟[1] 韩凌霞[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/实验动物中心,黑龙江哈尔滨150001 [2]甘肃农业大学动物科学技术学院,兰州甘肃730070 [3]东北农业大学动物医学院,黑龙江哈尔滨150030

出　　处：《中国预防兽医学报》2012年第1期68-70,共3页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家重点实验室基本科研业务费(NKLVBP200828)

摘　　要：为分析鸡主要组织相容性复合体(MHC)II类抗原β亚基BLB蛋白在鸡脾脏中的表达情况,本研究采用RT-PCR技术从鸡脾细胞中扩增了BLB基因的保守区第3外显子的部分序列,大小约为380 bp。将其克隆于表达载体pET-30a(+)中。在大肠杆菌BL21(DE3)中,经IPTG诱导表达,获得大小约为19 ku以包涵体形式存在的His-BLB重组蛋白。切取含有重组蛋白的SDS-PAGE胶免疫实验兔,获得抗血清。以制备的血清通过western blot在鸡脾脏中检测到MHC II类分子特异性条带,其大小约为28 ku,与BLB天然蛋白的大小相符合。本研究为进一步分析MHC-II类分子在鸡体内的蛋白表达水平提供了依据。To prepare the polyclonal antibodies against chicken BLB protein,a partial gene of chicken major histocompatibility complex（MHC） class II β chain（BLB） was amplified by RT-PCR from chicken splenocyte and cloned into pET-30a（＋） vector for expression in E.coli BL2l（DE3）.The recombinant protein was expressed with IPTG induction and the expressed protein was about 19 ku which mainly in the form of inclusion body.The expressed protein was crude purified by cutting protein containing slices from SDS-PAGE gel and immunized in rabbits to prepare antisera against BLB.The BLB proteins were detected in spleens of chickens by western blot.The result showed that the antiserum was able to detect specifically a 28 ku of BLB protein in the spleen,which was in accordance with the natural molecular weight of MHC-II protein.The positive serum could be used for further study of chicken MHC-II protein.

关 键 词：鸡主要组织相容性复合体 BLB基因 兔抗血清 蛋白质免疫印迹 

分 类 号：S852.5[农业科学—基础兽医学]