禽多杀性巴氏杆菌C_(48-1)外膜蛋白H基因真核表达载体的构建

Construction of eukaryotic expression vector of outer membrane protein H gene of avian Pasteurella multocida C_(48-1)

应用限制性内切酶BamHⅠ将片段ompH从重组质粒pMDT-omp H切下,非定向插入真核表达质粒pcDNA3中,经酶切分析、PCR鉴定及序列测定,证实成功构建了禽多杀性巴氏杆菌C48-1外膜蛋白H基因的重组真核表达质粒pcDNA3-ompH。将重组表达质粒转染至 COS7 细胞,经RT-PCR、SDS-PAGE及Western-blotting检测,证明了外膜蛋白H基因获得了瞬时表达。

The fragment omp H gene was obtained from the recombinant plasmid pEDT-ompH by (restrictive) enzyme BamHⅠ digestion,and then was inserted into eukaryotic expression vector pcDNA3. After restriction enzyme analysis, PCR identification and sequencing, the recombinant plasmid pcDNA3-ompH was constructed successfully. The recombinant plasmid was transfected into COS7 cells by (polybrene). The outer membrane protein H gene was expressed instantaneously in COS7 cells, which was (confirmed) by RT-PCR,SDS-PAGE and Western-blotting.