关键词: 中华蜜蜂, 囊状幼虫病毒, VP1蛋白, 系统进化, 原核表达

Abstract: 【Aim】 To understand the molecular evolution characteristics and genetic diversity of VP1 protein of Chinese sacbrood virus (CSBV) in Apis cerana cerana. 【Methods】 Eight cDNA sequences encoding VP1 proteins were cloned by RTPCR from Beijing isolates of Chinese sacbrood virus. 【Results】 Sequence analysis results showed that the open reading frame (ORF) of VP1 gene is 945 bp in length, encoding 315 amino acids with the predicted molecular weight of 35.42 kDa and the theoretical isoelectric point of 9.23, and the encoded protein has hydrophilicity and immunogenicity characteristics. Multiple sequence alignment indicated that the VP1 proteins from these CSBV Beijing isolates in different years share high amino acid sequence identity with occasional changes. The VP1 genes from Beijing isolates have high nucleotide sequence identity with those of Liaoning (LN) and Vietnam isolates (93%), India and Korea isolates (92%), while have the lowest nucleotide sequence identity with that of UK isolate (88%). The sequence analysis results also showed that the VP1 gene from Beijing isolates, other than from some isolates in other areas, has a specific amino acid characteristic which includes amino acid insertion. The nucleotide substitution rate among all Asian isolates was less than that between Asian and European isolates. A recombinant plasmid pEASY-E1-VP1, containing the coding sequence of VP1 protein, was constructed using pEASY-E1 as a fused expression vector, and VP1 protein was expressed successfully after induced with isopropyl-beta-D-thiogalactopyranoside (IPTG) in BL21(DE3)pLysS strain of Escherichia coli. 【Conclusion】 The results provide the basis for further studying the molecular mechanisms of the pathogenicity evolution of CSBV by confirming the existence of mutants in CSBV of different isolates.