兔出血症病毒VP60截短基因原核表达及诱导条件优化

Optimization of Induction Conditions for Prokaryotic Expression of Rabbit Hemorrhagic Disease Virus VP60 Truncated Gene

兔瘟是由兔出血症病毒(RHDV)引起的传染病,给肉兔养殖产业带来巨大损失。根据RHDV VP60基因序列进行抗原肽分析,选择该基因主要抗原肽段后,设计一对特异性引物进行PCR扩增,并插入pET32a(+)载体中构建重组质粒。经PCR鉴定、酶切鉴定和测序,结果表明p ET32a(+)-VP60截短基因质粒构建成功。对RHDV-VP60截短基因蛋白原核表达系统筛选诱导条件,结果表明:IPTG浓度为0.8 mmoL/L,诱导时间为8 h,诱导温度为37℃,表达量最高。研究为后续兔瘟亚单位疫苗和诊断试剂的研究奠定基础。

Rabbit plague is an infectious disease caused by rabbit hemorrhagic disease virus(RHDV),which brings huge losses to the rabbit breeding industry.According to the antigenic peptide analysis of RHDV VP60 gene sequence,after selecting the main antigenic peptide segment of the gene,a pair of specific primers were designed for PCR amplification and inserted into pET32a(+)vector to construct the recombinant plasmid.The results of PCR,restriction enzyme digestion and sequencing showed that the pET32a(+)-VP60 truncated gene plasmid was successfully constructed.The results showed that the expression level was the highest,which the concentration of IPTG was 0.8 mmoL/L,the induction time was 8 h,the induction temperature was 37°C.This study lays a foundation for the follow-up study of rabbit plague subunit vaccine and diagnostic reagents.