利用杆状病毒表达系统表达鸡传染性法氏囊病病毒VP2蛋白

Expression of VP2 protein of infectious bursal disease virus using baculovirus expression system

从GenBank下载传染性法氏囊病病毒(IBDV)VP2基因序列,根据昆虫细胞偏爱密码子对IBDV VP2基因进行密码子优化,化学合成优化后的VP2基因序列,然后构建杆状病毒表达载体pTri Ex-4-VP2,构建成功的重组杆状病毒pTri Ex-4-VP2转染sf9细胞,制备病毒原种,采用SDS-PAGE和Western blot方法鉴定VP2的免疫原性。结果显示:细胞感染Bac-VP2后,其细胞裂解蛋白在58 ku附近有1个条带,且该条带与IBD阳性血清发生特异性反应;间接免疫荧光试验(IFA)结果显示VP2基因能在Sf9细胞中表达;动物攻毒保护试验中,2次免疫重组VP2蛋白后能诱导14日龄SPF鸡产生对IBDV强毒攻击,保护率为60%。本研究为进一步研究IBDV VP2基因工程疫苗提供了物质基础。

The gene sequence of VP2 protein of infectious bursal disease virus was downloaded from GenBank,codon was optimized based on the insect cell preference,and then the optimized gene sequence was chemically synthesized. The baculovirus expression vector pTri Ex-4-VP2 was constructed,transfected into sf9 cells and analyzed by SDS-PAGE and Western blot. As expected,the calculated protein were approximately 58 ku and could be specifically recognized by chicken positive sera against IBDV. The result of indirect immunofluorescence assay（ IFA） also indicated that VP2 protein has been expressed in sf9 cells. Subsequently,the SPF chickens were immunized with recombinant VP2 protein twice,followed by challenging with IBDV BC6-85 at 2 weeks post the boost vaccination. The result showed that 60% of the chickens were protected against the challenge with very virulent IBDV. The investigation provides a basis for development of a novel subunit vaccine using VP2 protein.