猪流产嗜性衣原体MOMP基因的克隆及原核表达  

Cloning and prokaryotic expression of the major outer membrane protein coding gene of Swine Chlamydophila abortus

作　　者：李西玉[1,2] 许信刚[1] 周继章[2] 邱昌庆[2] 曹小安[2] 宫晓炜[2] 王光华[2] 

机构地区：[1]西北农林科技大学动物医学院,陕西杨凌712100 [2]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室,农业部兽医公共卫生重点开放实验室,甘肃兰州730046

出　　处：《西北农林科技大学学报（自然科学版）》2010年第7期33-38,43,共7页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家科技基础性工作专项(2008FY210200);陕西省科技攻关项目(2009K02-01);西北农林科技大学"青年学术骨干"支持计划项目(E111020901)

摘　　要：【目的】克隆猪流产嗜性衣原体青海株主要外膜蛋白(Major outer membrane protein,MOMP)基因,并进行序列分析及原核表达。【方法】根据GenBank公布的猪流产嗜性衣原体MOMP基因的核苷酸序列,设计并合成4条特异性引物,用套式PCR方法扩增猪流产嗜性衣原体青海株MOMP基因,将其克隆入pMD18-T载体中,进行测序及序列分析。然后将MOMP基因亚克隆入原核表达载体pGEX4T-1中,在大肠杆菌BL21(DE3)中用IPTG诱导表达,对表达产物进行SDS-PAGE和Western blot检测。【结果】扩增到猪流产嗜性衣原体青海株1170bp的MOMP全长基因。序列分析结果表明,该基因与已发表的猪流产嗜性衣原体B11001株和CP/12株核苷酸同源性均为99.7%。SDS-PAGE电泳可检测到分子质量约为66ku的融合蛋白,主要以包涵体形式存在。Western-blot分析表明,重组蛋白可被猪流产嗜性衣原体抗体识别。【结论】成功克隆了猪流产嗜性衣原体青海株MOMP基因,并进行了原核表达,表达的蛋白具有抗原活性。【Objective】The outer membrane protein(MOMP)gene of Swine Chlamydophila abortus(CA)Qinghai strain was cloned,analyzed and expressed in E.coli.【Method】According to the published complete nucleotide sequence of CA in GenBank,four primers specific to the full-length of MOMPgene were designed.The MOMPgene fragment amplified by nested PCR was cloned into pMD18-Tvector.The recombinant plasmid was sequenced and MOMPgene was compared with other GeneBank s CA strains.MOMPgene segment was amplified from recombinant pl...

关 键 词：猪 流产嗜性衣原体 MOMP蛋白 原核表达 

分 类 号：S852.67[农业科学—基础兽医学]