生物技术通报2017，Vol.33Issue(7)：150-154,5.DOI:10.13560/j.cnki.biotech.bull.1985.2016-1182
羊口疮病毒蛋白ORFV035的表达、纯化和多克隆抗体的制备
Expression and Purification of orf Virus Protein ORFV035 and Preparation of Polyclonal Antibody
摘要
Abstract
This work aims to express and purify recombinant orf virus encoded protein 035(ORFV035)in Escherichia coli and prepare a polyclonal antibody(pAb)against ORFV035 for immunoassays,as well as lay foundation for the further researches on the replication, assembly,morphogenesis,and mature process . The gene of ORFV035 was amplified by PCR from orf viral genome,and sub-cloned into the expression vector pET-30a(+),and recombinant plasmid pET-30a-035 was constructed by enzymatic digestion and ligation of BamH I and Hind III. Then the plasmid pET-30a-035 after double enzyme digestion and sequencing was transformed into Escherichia coli BL21, the transformants were induced by IPTG,and SDS-PAGE was used to identify the expression of the protein. The His-tagged fusion protein ORFV035 was ultrasonic-treated and then purified through Ni-chelating affinity chromatography. The purified ORFV035 was used to immune into BALB/c mouse for producing anti-ORFV035 pAb. The results showed that the recombinant plasmids pET30a-035 was successfully constructed,and the protein ORFV035 in E. coli BL21 was in inclusion body. The ORFV035-his fusion protein was obtained after washing, dissolving,and purifying the inclusion body,and the pAb anti-ORFV035 was prepared. The Western blot analysis showed that this pAb recognized natural ORFV035.关键词
羊口疮病毒/ORFV035/纯化/多克隆抗体Key words
orf virus/ORFV035/purification/polyclonal antibody引用本文复制引用
陈慧芹,王小平,罗树红,王丽洁,陈倩倩,郝文波..羊口疮病毒蛋白ORFV035的表达、纯化和多克隆抗体的制备[J].生物技术通报,2017,33(7):150-154,5.基金项目
国家自然科学基金项目(31672536,31170147) （31672536,31170147）
