Marc-145细胞CD151蛋白主要抗原表位区的表达及其与猪繁殖与呼吸综合征病毒感染关系的研究

为研究CD151与猪繁殖与呼吸综合征病毒(PRRSV)感染的关系,根据GenBank中已发表的CD151蛋白的基因序列设计并合成一对特异性引物,从Marc-145细胞中扩增出294 bp的CD151基因片段并克隆入pGEX4T-3载体,转化人大肠杆菌用IPTG进行诱导表达,经SDS-PAGE和Western blot对表达产物进行鉴定.表达蛋白纯化后用于免疫小鼠制备抗CD151蛋白血清,用ELISA和IFA检测抗血清的效价及特异性.将抗CD151蛋白血清与Marc-145细胞孵育后再感染PRRSV,观察细胞CPE验证该血清对PRRSV的阻断效果.结果表明成功构建了pGEX4T-3-CD151,获得了相对分子质量为37 ku的重组CD151蛋白.制备的抗血清特异性结合Marc-145细胞并有效阻断PRRSV感染Marc-145细胞.这些研究结果为进一步阐明CD151与PRRSV感染的关系提供一定的理论基础.

Expression of Main Epitope Domains of CD151 from Marc-145 Cells and Relationship with Infection of Porcine Reproductive and Respiratory Syndrome Virus

To study the role of CD151 in porcine reproductive and respiratory syndrome virus(PRRSV) infection,primers specific to CD151 were designed according to the similar gene sequences submitted in GenBank.Total RNA was extracted from Marc-145 cell and a 294 bp of CD151 was amplified by RT-PCR.The target gene was cloned into pGEX4T-3 vector,then was transformed into E.coli and expressed with IPTG inducer.SDS-PAGE and Western blot were employed to identify the expressed protein.The protein was purified and used to immunize mice.The antiserum titres were determined by the ELISA and IFA,and were used to block PRRSV infection of Marc-145 cells.The results showed that CD151 gene was amplified and cloned into pGEX4T-3-CD151 vector.The recombinant CD151 protein with MW of 37 kDa was produced.The antiserum reacted specifically with the recombinant protein and blocked PRRSV infection of Marc-145 cells.These results provide useful information for further understanding the role of CD151 in PRRSV infection.