摘要
以猪流行性腹泻病毒CH/JL毒株的RNA为模板,通过RT-PCR扩增获得的3个相互重叠的cDNA克隆覆盖了S基因,序列比对结果表明：PEDV CH/JL株S基因与CV777、Brl/87、JS、KPEDV和Chinju99毒株S基因核苷酸序列的同源性分别为96.97%、96.87%、96.41%、94.02%和93.93%,氨基酸序列的同源性分别为96.17%、95.88%、96.10%、92.36%和92.05%;分子进化树分析结果显示,PEDV CH/JL株S基因与JS毒株S基因亲缘关系最近,处于同一群。利用DNAstar Protean程序预测了PEDV CH/JL株S蛋白一个抗原表位区（83-276aa）,将其克隆到原核表达载体pGEX-6p-1后转化E.coliBL21（DE3）感受态细胞,在终浓度1.0mmol/L的IPTG诱导下获得了表达,Western blot结果显示,预测的抗原表位区GST融合蛋白能与猪流行性腹泻病毒多克隆抗血清反应,提示该抗原表位区含有线性抗原表位。
Three overlapping cDNA clones of the spike gene of porcine epidemic diarrhea virus CH/JL strain was amplified by RT-PCR. Sequence analysis showed that the S gene of CH/JL strain was 96.97%, 96.87%, 96.41%, 94.02% and 93.93% identical at nucleotide sequence level and 96.17%, 95.88%, 96.10%, 92.36% and 92.05% identical at amino acid sequence level to that of strains CV777, Brl/87, JS, KPEDV and Chinju99, respectively. CH/JL strain and JS strain belonged to the same group in the phylogenetic tree based on the nucleotide sequences of the S genes of PEDV strains. An antigenic epitope region of CH/JL strain S protein was predicted by using DNAStar protean software, and was expressed in E. coli BL21（DE3）as a GST fusion protein. Western blotting showed that the fusion protein reacted positively with the polyclonal antibodies against PEDV, suggesting that the predicted antigenic region contained linear antigenic epitopes of PEDV spike protein.
出处
《病毒学报》
CAS
CSCD
北大核心
2007年第3期224-229,共6页
Chinese Journal of Virology
关键词
猪流行性腹泻病毒
S基因
序列分析
线性抗原表位区
porcine epidemic diarrhea virus
spike protein gene
sequence analysis
linear antigenic epitope region
