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目的:制备抗百合斑驳病毒(Lily mottle virus,LMoV)CP血清,为研究侵染不同植物LMoV之间的血清学关系以及大田检测LMoV奠定基础。方法:根据Genbank报道的百合斑驳病毒CP基因序列设计引物,扩增其外壳蛋白基因并分析序列。将LMoV CP基因插入表达载体pSBET,转化大肠杆菌BL21(DE3)Plys E菌株,通过IPTG诱导表达。经12%SDS-PAGE和5%～20%梯度SDS-PAGE两次纯化CP,免疫小鼠获得抗CP血清,采用Western blot分析抗体的特异性,采用ELISA分析抗体能否与天然LMoV病毒结合。结果:LMoV的CP与目前已报道的LMoV不同分离物CP基因核苷酸序列同源性为95%～99%,氨基酸序列同源性为98%～100%。制备的抗体对CP具有高度特异性,且能够与天然LMoV病毒离子结合。结论:本研究制备的抗血清可以作为百合斑驳病毒的检测。
Objective: To prepare anti-lily mottle virus (CPV) CP serum and lay the foundation for the investigation of the serological relationship between LMoV infection and field detection of LMoV. Methods: Primers were designed according to the CP gene sequence of lily mottle virus virus reported in Genbank. The coat protein genes were amplified and the sequences were analyzed. The LMoV CP gene was inserted into the expression vector pSBET and transformed into E. coli BL21 (DE3) Plys E strain for expression by IPTG. CPs were purified twice by 12% SDS-PAGE and 5% -20% gradient SDS-PAGE, and the mice were immunized to obtain anti-CP serum. The specificity of the antibodies was analyzed by Western blot. The antibodies were analyzed by ELISA for binding to the native LMoV virus. RESULTS: The CP of LMoV was 95% -99% homologous to CP of the different LMoV isolates reported so far, and the amino acid sequence homology was 98% -100%. The prepared antibodies are highly specific to CP and are capable of binding to native LMoV virus ions. Conclusion: The antiserum prepared in this study can be used as lily mottle virus detection.