关键词: ORFV, 免疫相关基因, 系统进化树, 同源性分析

Abstract: In this study, a highly virulent orf virus （ORFV） strain was isolated from the clinical samples of the diseased sheep. After 90 consecutive passages on goat fibroblasts, the obtained ORFV clinical strain was attenuated. The genomes of the highly virulent ORFV strain and the attenuated ORFV strain were extracted and designated as ORFV-QD and ORFV-RD, respectively. A total of 3 sets of specific primers were designed based on the whole genome sequence of ORFV published on GenBank and used to amplify the gene fragments of B2L, F1L and VIR in ORFV-QD and ORFV-RD. The obtained genomic fragments were cloned into pMD-19T vector and were subsequently transformed into DH5α competent cells. After identification, the positive recombinant plasmids were sequenced. The obtained target gene sequences were aligned, and the nucleotide sequence homology within the same gene between ORFV-QD and ORFV-RD was compared using DNASTAR software. Furthermore, the nucleotide sequence homology comparison, gene-based phylogenetic analysis and amino acid sequence homology comparison between the gene sequences obtained in this study and corresponding gene sequences from 13 ORFV whole genome sequences published on NCBI were carried out. The results showed that the nucleotide sequence homology of the B2L, F1L and VIR genes between the sequences obtained in this study and the reference sequences was 92.1%-98.4%, 96.1%-99.1%, and 94.6%-100%, respectively. The amino acid sequence homology analysis demonstrated that there were no obvious differences between ORFV-QD and ORFV-RD.

Key words: ORFV, immune-related genes, phylogenetic tree, homology analysis