摘要
通过对PRRSV Hn-1/06株ORF5基因序列分析,设计删除信号肽和跨膜功能区的ORF5序列的3对引物,应用重叠延伸PCR以PTG19-T-ORF5质粒为模板进行扩增目的片段,得到长度为410 bp的片段。然后将此基因亚克隆到原核表达载体PET32a,经筛选获得了阳性重组质粒PET32a-ORF5abc,进而在IPTG的诱导下成功表达,获得了大小约为34 kD的融合蛋白GP5abc-His,Western blotting检测结果证实表达的融合蛋白具有良好的生物学活性。
According to analysis of the ORF5 gene order in PRRSV Hn - 1/06 strain, three pairs of primers for the deletion of signal peptide and ORF5 sequence in transmembrane domain function area were designed, and PTGI9 -T -GP5 was used as a template, 410 bp segment was obtained by overlap extension PCR amplification. Then after being sub - cloned into the prokaryotie expression vector PET32a, the ORF5abc gene was successfully obtained with the inducement of 1.0 mmol/L IPTG. Western -blotting was performed to confirm that the expressed fusion protein GP5abc - His (about 34 kD) could speeifically react with antiserum against PRRSV.
出处
《江西农业学报》
CAS
2009年第1期91-94,共4页
Acta Agriculturae Jiangxi