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云南农业大学学报(自然科学) 2017, Vol. 32 Issue (3) :410-420    DOI: 10.16211/j.issn.1004-390X(n).2017.03.004
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烟草丛顶病毒云南不同分离物全基因组序列测定与分析
吕婧1, 刘芳1, 谭冠林2, 魏薇1, 尹朝先1, 马龑1, 陈小姣1, 和理淮1,3, 陈海如1, 李凡1
1. 云南农业大学 农业生物多样性与病虫害控制教育部重点实验室, 云南 昆明 650201;
2. 云南农业大学 教务处 现代教育技术中心, 云南 昆明 650201;
3. 丽江市古城区农业局农业技术推广中心, 云南 丽江 674199
Determination and Analysis of the Complete Nucleotide Sequences of Different Isolates of Tobacco bushy top virus in Yunnan, China
LYU Jing1, LIU Fang1, TAN Guanlin2, WEI Wei1, YIN Chaoxian1, MA Yan1, CHEN Xiaojiao1, HE Lihuai1,3, CHEN Hairu1, LI Fan1
1. Key Laboratory of Agro-Biodiversity and Pest Management of Education Ministry of China, Yunnan Agricultural University, Kunming 650201, China;
2. Modern Education Technology Center, Yunnan Agricultural University, Kunming 650201, China;
3. Agricultural Technology Extension Center of Gucheng District, Lijiang 674199, China

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摘要 [目的]探明烟草丛顶病毒(Tobacco bushy top virus,TBTV)云南不同分离物之间的遗传差异并完成系统进化分析。[方法] 采用RT-PCR方法扩增获得云南不同地区10个TBTV分离物的全基因组序列。[结果] 10个TBTV云南分离物的全基因组序列均为4 152 bp,基因组结构由4个开放阅读框(ORFs)构成,ORF2通过与ORF1发生-1位的移码(frameshift)表达融合蛋白,推测ORF1末端的GGAUUUU特征序列是确定ORF1与ORF2是否发生移码翻译的依据。各TBTV云南分离物全基因组序列的核苷酸序列同源性为84.9%~99.1%,其中来自德宏的分离物(TBTV-YDHo)变异最大。TBTV不同分离物间存在重组现象,且分离物之间亲缘关系与重组的发生有关,重组主要发生在变异较大的ORF1~ORF2区段。TBTV分离物的各区段中,ORF1和ORF1~ORF2变异最大,而ORF4和3'UTR较为保守。系统进化分析结果表明:TBTV作为幽影病毒属(Umbravirus)的成员之一,其与同属中的罂粟花叶病毒(Opium poppy mosaic virus,OPMV)亲缘关系最近,核苷酸序列同源性为61.1%~61.6%;与CMoV和CMoMV亲缘关系较远,同源性为54%~55.1%。除幽影病毒属外,TBTV与番茄丛矮病毒科(Tombusviridae)其他属的病毒核苷酸序列同源性为25.1%~49.8%;Tombusviridae中Umbravirus与香石竹环斑病毒属(Dianthovirus)的RNA2亲缘关系最远,与其他病毒属成员的亲缘关系都相对较近。[结论] TBTV云南各分离物之间存在一定的分子变异,有的存在重组现象;TBTV与OPMV亲缘关系最近。
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吕婧
刘芳
谭冠林
魏薇
尹朝先
马龑
陈小姣
和理淮
陈海如
李凡
关键词烟草丛顶病毒   幽影病毒属   番茄丛矮病毒科   序列分析   重组     
Abstract: [Purpose] The aim of this study was to explore the genetic differences among different isolates of Tobacco bushy top virus (TBTV) in Yunnan and complete the phylogenetic analysis.[Method] The complete genomes of ten TBTV Yunnan isolates were amplified by RT-PCR.[Result] All of the genomic sequences of the ten TBTV isolates were 4 152 nucleotides (nt) in size, and contained four open reading frames (ORFs). ORF2 expressed the fusion protein through -1 frameshift with ORF1, and the sequence GGAUUUU was suggested to be the signature for the frameshift strategies of ORF1-ORF2 in TBTV. Sequence analysis results revealed the nt identity of the complete sequence of the ten TBTV isolates was ranged from 84.9% to 99.1%, and isolate TBTV-DHo was the most variant compared with the other TBTV isolates. Recombinant events were detected among the TBTV Yunnan isolates. Phylogenetic relationships of these isolates involved in the recombinant, and recombinant was mainly occurred in the variant regions of ORF1-ORF2. ORF1 and ORF1-ORF2 were divergent among the four TBTV ORFs, while ORF4 and 3'UTR were conserved. Phylogenetic analysis showed that TBTV was most closely related to Opium poppy mosaic virus (OPMV) among different umbraviruses with nt identity of 61.1% to 61.6%, while was more distant to Carrot mottle virus (CMoV) and Carrot mottle mimic virus (CMoMV) with nt identity of 54%-55.1% in genus Umbravirus. Among the members in the 13 genera of family Tombusviridae, TBTV shared 25.1%-49.8% nt sequence identity with other viruses except umbraviruses, while umbraviruses were distant with the RNA2 of Dianthoviruses in family Tombusviridae.[Conclusion] Sequence analysis indicated a high genetic diversity among the TBTV Yunnan isolates, and recombinant events occurred among these isolates. TBTV was most closely related to OPMV among different umbraviruses.
Keywordstobacco bushy top virus   Umbravirus   tombusviridae   sequence analysis   recombinant     
Received 2016-05-16;
Fund:云南省高校科技创新团队支持计划(云教科[2014]22号);国家自然科学基金(30660101,31571983);云南农业大学杰出人才专项资助计划(2015JY01)。
Corresponding Authors: 陈海如(1944-),男,云南昆明人,博士,教授,主要从事植物病毒学研究。E-mail:13908854041@126.com;李凡(1970-),男,云南凤庆人,博士,教授,主要从事植物病毒学研究。E-mail:fanlikm@126.com     Email: 13908854041@126.com;fanlikm@126.com
About author: 吕婧(1989-),女,陕西千阳人,硕士研究生,主要从事植物病毒学研究。E-mail:18487331622@126.com;刘芳(1985-),女,四川内江人,硕士,主要从事植物病毒学研究。E-mail:lyf104821@126.com
引用本文:   
吕婧, 刘芳, 谭冠林, 魏薇, 尹朝先, 马龑, 陈小姣, 和理淮, 陈海如, 李凡.烟草丛顶病毒云南不同分离物全基因组序列测定与分析[J]  云南农业大学学报(自然科学), 2017,V32(3): 410-420
LYU Jing, LIU Fang, TAN Guanlin, WEI Wei, YIN Chaoxian, MA Yan, CHEN Xiaojiao, HE Lihuai, CHEN Hairu, LI Fan.Determination and Analysis of the Complete Nucleotide Sequences of Different Isolates of Tobacco bushy top virus in Yunnan, China[J]  Journal of Yunnan Agricultural University, 2017,V32(3): 410-420
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