为了明确O型口蹄疫病毒不同宿主适应毒株P1和3A基因位点的差异性,研究其遗传变异趋势,找出不同宿主适应毒株在P1和3A基因水平上位点的变异情况,将O型口蹄疫病毒O/XJ/10-11株分别接种于牛、乳鼠、猪及BHK-21细胞,获得相应的宿主适应毒株,提取RNA,反转录并扩增P1和3A基因,目的片段产物经琼脂糖凝胶电泳回收,并将其克隆到pMD19-T载体上,筛选阳性菌落,测序并分析其基因序列.结果表明:牛、乳鼠、猪、BHK-21细胞四种宿主适应毒株,核苷酸和氨基酸序列均未发生缺失,主要抗原位点稳定;P1基因发生了部分变异,变异程度依次为VP1、VP2＞VP3＞VP4,VP4基因最为保守;3A基因较稳定,变异较少;不同宿主适应毒株的变异程度依次如下:猪适应毒株＞BHK-21细胞适应毒株＞牛适应毒株＞鼠适应毒株.O型口蹄疫病毒在不同宿主传代过程中造成VP1基因的变异,但主要抗原位点稳定;VP2基因的变异均位于抗原表位上;鼠适应毒株作为口蹄疫原始毒种的保存更为有利.%Difference of P1 and 3A gene sequences of different hosts-adapted strains type O Footand mouth disease virus (FMDV) were identified,and the trend of genetic variation were studied to find the genetic mutation of P1 and 3A genes of different hosts-adapted FMDV.FMDV O/XJ/10-11 strain was inoculated into bovine,neonatal rat,swine and BHK-21 cell line to obtain corresponding adapted virus strains.The RNA of FMDV in each adapted virus was taken as template for reverse transcription and amplification of P1(1A,1B,1C,1D)and 3A genes.Fragment was purified and recovered by agarose gel electrophoresis,cloned into the vector pMD19-T.Positive colonies were screened,sequenced,and the gene sequences were analyzed.The results were as follows:no deletions were found in nucleotide and amino acid sequences of different hosts-adapted virus strains,the main antigenic sites were conserved in different hosts;The P1 gene of the bovine,swine,neonatal rat,and BHK-21 cells adapted virus strains had some variations,the variation extent decreased in sequence of VP1,VP2 ＞ VP3＞ VP4,and VP4 was the most conserved gene region;3A gene was more stable and showed less variation;The most varied host-adapted FMDV was from swine,followed by BHK-21 cells,bovine and neonatal rat.The results showed that there is greater variability in the gene of VP1,however,the main antigenic sites of VP1 were conserved;VP 2 mutations were located in its antigenic epitope;as the storage of the original virus of FMDV,neonatal rat-adapted virus is more favorable.
展开▼
