Comparison of ORFV011 Genes of Orf virus Vaccine strain and ORFV-PN Strain
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摘要:
目的 比较分析羊口疮病毒疫苗株与ORFV-PN株ORFV011基因的分子特点、编码蛋白的生物学功能差异，为福建省羊口疮的科学防控提供理论指导。 方法 对羊口疮疫苗株和ORFV-PN株ORFV011基因进行克隆和测序，运用生物信息学相关软件比较分析两者测序结果的差异。 结果 测序结果显示，疫苗株和ORFV-PN株ORFV011基因均由1 137个核苷酸组成，编码378个氨基酸。核苷酸序列和氨基酸序列相似性比较显示，ORFV-PN与疫苗株和弱毒株D1701株ORFV011基因核苷酸序列相似性分别为98.6%和98.4%，氨基酸序列相似性均为98.7%。与疫苗株相比，ORFV-PN株共有16处核苷酸变异和5处氨基酸变异。在蛋白二级结构上，ORFV-PN株和疫苗株α-螺旋占比分别为34.13%（129/378）和30.42%（115/378），β-转角占比分别为6.08%（23/378）和6.88%（26/378），无规则卷曲占比分别为37.30%（141/378）和39.15%（148/378），β-折叠占比分别为22.49%（85/378）和23.54%（89/378）。蛋白三级结构上，ORFV-PN株预测的三维结构与疫苗株的三维结构在α-螺旋和无规则卷曲上有一定的差异。 结论 福建省羊口疮病毒ORFV011基因出现变异，应当引起广大兽医工作者的重视，有关部门应当加强对羊口疮的监测。 Abstract:Objective To provide theoretical basis for prevention and control of Orf in Fujian, the molecular characteristics and biological functions of ORFV011 genes of the vaccine strain and ORFV-PN strain were compared. Methods ORFV011 genes from the two strains were cloned and sequenced for comparison and analysis using bioinformatics software. Results The sequencing results indicated that both genes composed of 1 137 nucleotides coding 378 amino acids. The nucleotide sequence similarity of ORFV011 of the ORFV-PN strain was 98.6% with that of the vaccine strain, while 98.4% with that of the lentogen strain D1701. The amino acid sequence similarity of ORFV011 of the ORFV-PN strain with either the vaccine strain or the D1701 strain was the same at 98.7%. There were 16 nucleotide mutations and 5 amino acid variations in the gene of the ORFV-PN strain as compared to that of the vaccine strain. On the secondary structure, the encoded protein by ORFV011 gene of the ORFV-PN strain and the vaccine strain had 34.13% (or 129/378) and 30.42% (or 115/378) alpha helixes, respectively; 6.08% (or 23/378) and 6.88% (or 26/378) beta turns, respectively; 37.3% (or 141/378) and 39.15% (or 148/378) random coils, respectively; and, 22.49% (or 85/378) and 23.54% (or 89/378) extended strands, respectively. The predicted three-dimensional structure of ORFV011 of the ORFV-PN strain showed differences on the alpha helixes and random coils as compared with that of the vaccine strain. Conclusion It appeared that ORFV011 of the ORFV-PN strain deviated from that of the vaccine strain in sequence as well as secondary and three-dimensional structures. Thus, special attention must be placed in monitoring and controlling the Orf. -
Key words:
- Orf virus /
- vaccine strain /
- ORFV-PN strain /
- ORFV011 gene
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图 1 ORFV011基因的PCR扩增
注：M：2000 DNA Ladder；1：疫苗株DNA；2：ORFV-PN株DNA；3: 阴性对照。
Figure 1. PCR amplification of ORFV011 genes
Note: M:2000 DNA Ladder; 1: DNA of vaccine strain; 2:DNA of ORFV-PN strain; 3: negative control.
图 2 ORFV011重组质粒双酶切鉴定结果
注：M：5000 DNA Ladder；1：疫苗株重组质粒；2：ORFV-PN株重组质粒。
Figure 2. Identification of ORFV011 recombination plasmid by restriction endonuclease digestion
Note： M: 5000 DNA Ladder; 1:the recombination plasmid of vaccine strain; 2: the recombination plasmid of ORFV-PN strain.
图 4 疫苗株和ORFV-PN株ORFV011蛋白抗原位点预测
Figure 4. Predicted antigenic indices on ORFV011 in vaccine and ORFV-PN strains
图 5 疫苗株（A）和ORFV-PN株（B）ORFV011蛋白结构域三维结构预测
Figure 5. Predicted 3D tertiary structures of ORFV011 in vaccine strain (A) and ORFV-PN strain (B)
表 1 ORFV参考毒株
Table 1. ORFV reference strains
毒株Virus 登录号GenBank 分离地Isolation site 时间Time NZ2 DQ184476 新西兰New Zealand 2005 OV-SA00 AY386364 美国USA 2003 OV-IA82 AY386363 美国USA 2003 D1701 HM133903 德国Germany 2010 NA1-11 KF234407 中国广东Guangdong, China 2013 Shanxi HQ202153 中国山西Shanxi, China 2010 Jilin KJ808074 中国吉林Jilin, China 2011 Guizhou KP994595 中国贵州Guizhou, China 2010 Xinjiang2 KF666565 中国新疆Xinjiang, China 2012 FJ-NP KC568369 中国福建Fujian, China 2013 FJ-GT KC568394 中国福建Fujian, China 2009 China vaccine JQ904789 中国甘肃Gansu, China 2012 
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表 2 疫苗株和ORFV-PN株ORFV011蛋白二级结构预测
Table 2. Predicted secondary structures of ORFV011 in vaccine and ORFV-PN strains
蛋白二级结构预测 Secondary structure prediction α-螺旋 Alpha helix β-转角 Beta turn 无规则卷曲 Radom coil β-折叠 Extended strand ORFV-PN株 ORFV-PN strain 34.13%（129/378） 6.08%（23/378） 37.3%（141/378） 22.49%（85/378） 疫苗株 Vaccine strain 30.42%（115/378） 6.88%（26/378） 39.15%（148/378） 23.54%（89/378）
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