山羊痘病毒A11R基因的克隆及生物信息学分析

Cloning and bioinformatics analysis of A11R gene of Goatpox virus

为了分析绵羊痘病毒A11R蛋白质的分子特征,提取了山羊痘病毒古浪分离株(GL)的基因组DNA,设计引物进行PCR扩增,将PCR产物连接到p GEM-T Easy载体后转化至大肠杆菌DH5α感受态细胞,筛选阳性克隆进行序列测定,利用生物信息学软件对A11R基因序列进行预测分析。结果显示,A11R基因序列由954个核苷酸组成的开放阅读框,编码317个氨基酸残基组成的多肽,蛋白质分子质量的理论值为36 091.7,理论等电点为5.14。A11R蛋白质的二级结构中,α螺旋占29.02%,β折叠占10.41%,其余60.57%为无规则卷曲。多序列比对分析显示,不同羊痘病毒分离株A11R序列高度保守,同源性在98%以上。进化树分析结果显示,GL株与NK株、TU株以及SA株在一个分支,说明它们之间具有较近的亲缘关系,上述研究结果为进一步研究A11R蛋白质的生物学功能和羊痘病毒早期蛋白质分子间相互作用奠定了基础。

To explore the molecular characteristics of A11 R from Goatpox virus（ GTPV）,genomic DNA was extracted from Goatpox virus Gu Lang strains. The specific primers were designed and used to amplify A11 R gene from genomic DNA by PCR. The PCR product was ligated into p GEM-T Easy vector. After transformation into Escherichia coli DH5α,the positive clones were sequenced and the sequences were analyzed with the bioinformatic softwares.The result showed A11 R gene sequence contained an open reading frame（ ORF） of 954 nucleotides and the deduced protein consisted of 317 amino acids with the theoretical molecular weight of 36 091.7 and isoelectric point of 5. 14.Analysis of secondary structure of protein A11 R revealed that α-helix,β-strand and loop accounted for 29. 02%,10. 41% and 60. 57%,respectively. Multiple sequence alignment showed A11 R gene from different Capripoxvirus isolates were highly conserved with similarity as high as 98%. Phylogenetic analysis demonstrated that,GL and NK,TU and SA were grouped in one branch,indicative a close genetic relationship.