羊口疮病毒强弱毒株毒力相关基因的序列比较研究

发布时间：2018-03-23 04:28

本文选题：羊口疮病毒　切入点：致弱　出处：《内蒙古农业大学》2017年硕士论文　论文类型：学位论文

【摘要】：本研究将青岛某发病羊场分离保存的ORFV/QD/2015株,在山羊皮肤成纤维细胞上连续传90代,根据Genebank上发表的羊口疮全基因序列设计并合成4对特异性引物并扩增ORFV/QD/2015强毒株与传代90代后毒株的ORF020、ORF112、ORF117和ORF127基因片段,并将其分别克隆到PMD-19T载体,转化到DH5α感受态细胞,重组质粒鉴定后进行序列测定,用DNA Star软件拼接测序结果,进行生物学信息分析。试验结果显示,羊口疮病毒可以在山羊皮肤成纤维细胞上培养,并产生细胞病变。将ORFV/QD/2015强毒株与传代90代毒株的质粒测序结果与Genebank已经公布的9组ORFV全基因序列中的ORF020、ORF112、ORF117、ORF127基因比对结果中ORFV/QD/2015强毒株与公布的毒株毒力相关基因的同源性高,传代90代毒株与公布的毒株毒力相关基因的同源性有所降低,强毒株与传代90代毒株4组毒力相关基因核苷酸比对结果显示,ORF020基因氨基酸发生7处变异;ORF112基因氨基酸发生52处变异;ORF117基因氨基酸发生14处变异;ORF127基因氨基酸发生15处变异。比对后发现核苷酸与氨基酸变异最多的为ORF112基因,最少的为ORF020基因,说明羊口疮病毒致弱后可能与这些毒力相关基因的变异有很大联系。两毒株间的抗原指数也有明显变异。这些变异为毒力相关基因在强弱毒之间造成的差异提供试验基础。
[Abstract]:In this study, the ORFV/QD/2015 strain isolated from an infected sheep farm in Qingdao was transferred to goat skin fibroblasts for 90 consecutive generations. According to the whole gene sequence of sheep mouth ulcer published on Genebank, four pairs of specific primers were designed and synthesized, and the ORF020 ORF112ORF117 and ORF127 gene fragments of ORFV/QD/2015 virulent strain and virulent strain were amplified and cloned into PMD-19T vector and transformed into DH5 伪 competent cells, respectively. The recombinant plasmid was sequenced and sequenced by DNA Star software, and the biological information was analyzed. The results showed that the goat mouth sore virus could be cultured on goat skin fibroblasts. The results of sequencing the plasmids of ORFV/QD/2015 virulent strains and those of subculture 90 strains were compared with the ORF020 ORF112ORF117 ORF127 gene sequence published by Genebank. The results showed that the virulence related genes of ORFV/QD/2015 virulence strains were highly homologous to those of the published virulence genes. The homology of virulence related genes between the 90 generation strains and the published strains was decreased. Nucleotide comparison of virulence related genes in 4 groups of virulence related genes between virulent strains and generation 90 strains showed that ORF020 gene had 7 amino acids and ORF112 gene had 52 amino acids, and ORF117 gene amino acids had 14 mutations and ORF127 genes had amino acids. ORF112 gene was found to have the most variation between nucleotide and amino acid. The least common is the ORF020 gene. These results suggest that the weakened sheep mouth sore virus may be closely related to the variation of these virulence related genes, and the antigenic index of the two strains also has obvious variation. These variations provide the experimental basis for the difference between the virulence related genes and the virulence.
【学位授予单位】：内蒙古农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S852.65

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