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同安钮夜蛾核型多角体病毒Opdi108 基因的分子克隆和细胞定位(英文)
引用本文:常润磊,刘莉,韦春梅,郎国俊,许雯,毛珊珊,林同.同安钮夜蛾核型多角体病毒Opdi108 基因的分子克隆和细胞定位(英文)[J].昆虫学报,2012,55(4):403-411.
作者姓名:常润磊  刘莉  韦春梅  郎国俊  许雯  毛珊珊  林同
作者单位:华南农业大学林学院, 广州
基金项目:国家自然科学基金,广东省林业科技创新专项资金项目,国家高等学校本科特色专业建设点——林学项目
摘    要:【目的】同安钮夜蛾Ophiusa disjungens(鳞翅目, 夜蛾科)是危害桉树的主要害虫之一。同安钮夜蛾核型多角体病毒(OpdiNPV)是从O. disjungens中分离到的新病毒。本研究主要目的是从分子水平上了解该病毒。【方法】对一条PstⅠ的酶切片段进行了克隆、 测序和分析。【结果】在基因数据库中通过对碱基序列的比对, 发现了一个Ac108的同源基因, 命名为Opdi108 (GenBank登录号为EU 732666)。该基因的开放阅读框含有225对碱基, 其编码蛋白与其他已知杆状病毒同源物有16%~37%的氨基酸序列一致性。在起始密码子ATG上游有一个晚期启动子TAAG。C末端含有His-tag的Opdi108基因在大肠杆菌中Escherichia coli进行了表达, 融合蛋白的分子量为28.7 kDa。以荧光蛋白EGFP为标记物, 构建了Opdi108与EGFP的重组体, 利用Bac-to-Bac表达系统实现了与AcMNPV的重组, 用该重组病毒vEGFP-Opdi108感染斜纹夜蛾Trichoplusia ni 细胞。感染24和72 h后分别用共聚荧光显微镜观察, 发现Opdi108定位在细胞质内。【结论】本研究从分子生物学角度研-OpdiNPV, 为进一步利用该病毒防治包括同安钮夜蛾在内的害虫, 以及构建重组杀虫剂等奠定基础。

关 键 词:同安钮夜蛾  核型多角体病毒  Bac-to-Bac表达系统  细胞定位    Opdi108  

Molecular cloning and cellular localization of Opdi108 from Ophiusa disjungens nucleopolyhedrovirus
CHANG Run-Lei,LIU Li,WEI Chun-Mei,LANG Guo-Jun,XU Wen,MAO Shan-Shan,LIN Tong.Molecular cloning and cellular localization of Opdi108 from Ophiusa disjungens nucleopolyhedrovirus[J].Acta Entomologica Sinica,2012,55(4):403-411.
Authors:CHANG Run-Lei  LIU Li  WEI Chun-Mei  LANG Guo-Jun  XU Wen  MAO Shan-Shan  LIN Tong
Institution:College of Forestry, South China Agricultural University, Guangzhou 510642, China
Abstract:Aim] Ophiusa disjungens(Lepidoptera,Noctuidae) is one of the main insect pests that attack eucalyptus.A new nucleopolyhedrovirus(NPV) called Ophiusa disjungens NPV(OpdiNPV) was recently isolated from diseased larvae.The objective of this research was to understand OpdiNPV at the molecular level.Methods] A PstⅠ genomic fragment of OpdiNPV was cloned,sequenced and analyzed.Results] A homologue of Ac108 gene,assigned as Opdi108(GenBank accession number EU 732666),was found in this fragment.The open reading frame(ORF) of Opdi108 is 225 bp in length and its encoded protein has 16%-37% amino acid identities with other known baculovirus homologous proteins.Upstream of the start codon ATG of Opdi108,contains a late promoter motif TAAG.The ORF of Opdi108 with His-tag at C-terminus was expressed as a protein in Escherichia coli with a molecular mass of 28.7 kDa.EGFP-Opdi108 fusion protein was expressed in Trichoplusia ni(Tn) cells using Bac-to-Bac system,suggesting that Opdi108 was located mainly in the cytoplasm of infected cells at 24 h post-infection(p.i.) and 72 h p.i.Conclusion] This study provides a genetic basis for the development of the virus OpdiNPV as a biopesticide or an engineered pesticide to control O.disjungens and other lepidopteran insects.
Keywords:Ophiusa disjungens  nucleopolyhedrovirus  Bac-to-Bac expression system  cellular localization  Opdi108
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