摘要
旨在利用pLA载体将IBDV B87株VP2蛋白展示在干酪乳杆菌表面。首先通过RT-PCR扩增VP2基因片段,测序鉴定正确后,将目的片段构建在pLA穿梭质粒上,命名为pLA-VP2,然后将重组质粒电转化到干酪乳杆菌中,构建IBDV重组干酪乳杆菌。应用SDS-PAGE检测重组菌表达目的蛋白VP2,得到约90 kDa的融合蛋白,与预期结果相符。Western blot结果显示VP2蛋白可以与抗IBDV多抗血清发生特异性反应,具有很好反应原性。流式细胞仪的散射光检测器可以捕捉到pLA-VP2重组菌菌体表面的荧光,且重组菌荧光强度强于pLA菌对照组,应用正态分布函数计算pgsA-VP2融合蛋白的表达效率,约为80%。结果表明IBDV VP2蛋白成功展示在干酪乳杆菌表面。
Using pLA as a vector,fusion VP2 protein of infectious bursal disease virus（IBDV）would be expressed on the surface of Lactobacillus casei（L.casei）.First of all,the primers were employed to amplify the cDNA of the VP2 gene by polymerase chain reaction（PCR）.After sequencing,the target genes were inserted into the vector pLA and named pLA-V2.Then the recombinant plasmid pLA-VP2 was transformed into L.casei by electroporation,construction of IBDV recombinant Lactobacillus casei. The identified recombinant strans which expressed VP2 protein were detected by SDS-PAGE,Western blot and flow cytometry.SDS-PAGE and Western blot results showed that the molecular weight of about 90 kDa was achieved. Moreover,the recombinant VP2 protein had the good immunoreactivity. The light scattering detector of flow cytometry could capture the fluorescence of recombinant strains on the surface of the cell wall,where the fluorescence signal was stronger than control. The expression efficiency of fusion protein pgsA-VP2 was 80% as normal distribution curve.In conclusion,VP2 protein of IBDV had expressed on the surface of L.casei.
出处
《黑龙江八一农垦大学学报》
2014年第5期39-43,共5页
journal of heilongjiang bayi agricultural university
基金
黑龙江省研究生创新科研项目(YJSCX2013-11BYND)
