绵羊痘病毒固原株L2R基因的克隆与生物信息学分析

摘要

To explore the molecular characteristics of protein L2R from sheeppox virus(SPPV), genomic DNA was extracted from SPPV GY strain.The specific primers were designed and used to amplify the L2R gene from the genomic DNA by PCR.Then the PCR product was ligated into pGEM-T Easy vector.After transformation into E .coli Trans 5α,the positive clones were se-quenced and the sequences were analyzed by the bioinformatic softwares.The result showed that L2R gene sequence contained an open reading frame (ORF)of 279 nucleotides and deduced pro-tein consisted of 92 amino acids with the theoretical molecular weight of 10.92 ku and isoelectric point was 6.56.Analysis of secondary structure of protein L2R revealed that α-helix,β-strand, random coil and extended strand were 69.57%,9.78%,8.70% and 11.96%,respectively.Analy-sis of multiple sequence alignment showed that L2R gene from different capripox virus isolates were highly conserved,phylogenetic analysis showed that GY and NK,TU and SA was in a branch,indicating with a close genetic relationship among them.The present results laid a foun-dation for further studies of biological functions of protein L2R and interaction among the early proteins of capripox virus.%为分析绵羊痘病毒 L2R 蛋白的分子特征，本试验提取了绵羊痘病毒固原株（GY）的基因组 DNA，设计L2R 基因引物，对 L2R 基因进行扩增，将扩增的基因连接到 pGEM-T Easy 载体后转化到大肠杆菌 Trans 5α感受态细胞，筛选阳性克隆，进行序列测序。利用生物信息学软件对 L2R 基因序列进行预测分析。结果显示，L2R 基因序列含有一个由279个核苷酸组成的开放阅读框，编码92个氨基酸残基组成的多肽，蛋白质分子质量理论值为10．92 ku，理论等电点为6．56。该蛋白质二级结构组成分别是α-螺旋占69．57％，β-折叠占9．78％，无规则卷曲占8．70％，延伸链占11．96％。多序列比对分析结果显示，不同羊痘病毒分离株 L2R 序列高度保守。进化树分析结果显示，GY 株与 NK、TU 及 SA 株在一个分支，表明它们之间亲缘关系较近。本试验结果为进一步研究 L2R 蛋白的生物学功能和羊痘病毒早期蛋白的分子相互作用奠定了基础。