摘要
目的分析重庆地区中蜂囊状幼虫病病毒(Chinese sacbrood virus,CSBV)的遗传多样性。方法采用RT-PCR法从重庆地区中蜂囊状幼虫病病死幼虫体内扩增CSBV的2段结构蛋白和1段非结构蛋白的编码序列,并克隆至pMD18-T载体进行双向序列测定,所得序列在NCBI上进行BLAST搜索,以进行同源性分析;应用ClustalW2软件进行多序列比对分析;应用SNAP软件计算同义替换率(Synonymous substitution rate,dS)、非同义替换率(Nonsynon-ymous substitution rate,dN)及其比值(dS/dN);采用MEGA 4软件的邻接法(Neighbor-joining)中的最大可能性算法(Maximum likelihood method)构建系统进化树。结果 CSBV重庆分离株片段SB1-2、SB6-7和SB14-15的序列同源性高于99.06%,序列保守,3个片段序列均只发现少数点变异碱基,属于亚洲型,与东方蜜蜂广州分离株和韩国分离株的亲缘关系最近;CSBV重庆分离株存在特有的核苷酸和氨基酸位点,且与亚洲型分离株存在广泛的共有基序;亚洲型分离株的替换率高于欧洲型分离株。结论 CSBV分离株存在型特异的遗传变异规律。
Objective To analyze the genetic diversity of sacbrood virus isolates from Apis cerana cerana in Chongqing （CQ） Region, China. Methods The gene sequences encoding two partial structural protein and one partial non-structural protein of sacbrood virus isolates from A. c. cerana in Chongqing were amplified by RT-PCR and cloned into vector pMD18-T for dual sequencing. The obtained sequences were analyzed for homology to those in NCBI by using BLAST software, and aligned using ClustalW2 sequences alignment program. Synonymous substitution rate （dS）, non-synonymous substitution rate （dN） and the ratio of dS to dN were calculated by synonymous Non-synonymous Analysis Program （SNAP）. Phylogenetic trees were constructed using the maximum likelihood method in neighbor-joining in MEGA 4 soft- ware. Results The homology of CSBV-CQ isolates SB1-2, SB6-7 and SB14-15 was more than 99. 06%. The sequences of the three isolates were conserved, in which only a small quantity of point mutated bases were observed. All the isolates belonged to Asian genotype and were closely related to CSBV-GZ and Korean isolates from A. c. cerana. Several unique sites of nucleotides and amino acids appeared only in CSBV-CQ isolates, while a number of shared motifs were also observed between the Asian genotype isolates. However, the substitution rates in Asian genotype strains were higher than those in European genotype ones. Conclusion Type-specific genetic variations were observed in CSBV isolates.
出处
《中国生物制品学杂志》
CAS
CSCD
2013年第3期315-323,共9页
Chinese Journal of Biologicals
基金
现代农业产业技术体系建设专项资金资助(CARS-45-SYZ15)
重庆市畜牧科学院基本科研业务费项目(10615)
关键词
中蜂
囊状幼虫病病毒
遗传变异
多序列比对
替换率
系统发生
Apis cerana cerana
Sacbrood virus
Genetic variation
Sequence alignment
Substitution rate
Phylogenesis
