目前国内外对水稻瘤矮病毒(Rice gall dwarf virus,RGDV)的研究主要集中在其核衣壳蛋白和外衣壳蛋白的性质、结构及其与介体的亲和性、与介体传毒能力的关系等方面,本室在已有室内检测研究的基础上,利用RT-PCR和PVP-ELISA法对田间带毒...目前国内外对水稻瘤矮病毒(Rice gall dwarf virus,RGDV)的研究主要集中在其核衣壳蛋白和外衣壳蛋白的性质、结构及其与介体的亲和性、与介体传毒能力的关系等方面,本室在已有室内检测研究的基础上,利用RT-PCR和PVP-ELISA法对田间带毒叶蝉进行检测,并对2种方法的检测结果进行了比较.展开更多
To obtain the P8 protein of Rice gall dwarf virus (RGDV) with biological activity,its outer coat protein gene S8 was expressed in Spodoptera frugiperda (Sf9) insect cells using the baculovirus expression system.The S8...To obtain the P8 protein of Rice gall dwarf virus (RGDV) with biological activity,its outer coat protein gene S8 was expressed in Spodoptera frugiperda (Sf9) insect cells using the baculovirus expression system.The S8 gene was subcloned into the pFastBacTM1 vector,to produce the recombinant baculovirus transfer vector pFB-S8.After transformation,pFB-S8 was introduced into the competent cells (E.coli DH10Bac) containing a shuttle vector,Bacmid,generating the recombinant bacmid rbpFB-S8.After being infected by recombinant baculovirus rvpFB-S8 at different multiplicities of infection,Sf9 cells were collected at different times and analyzed by SDS-PAGE,Western blotting and immunofluorescence microscopy.The expression level of the P8 protein was highest between 48-72 h after transfection of Sf9 cells.Immunofluorescence microscopy showed that P8 protein of RGDV formed punctate structures in the cytoplasm of Sf9 cells.展开更多
RNA silencing is a conserved mechanism found ubiquitously in eukaryotic organisms.It has been used to regulate gene expression and development.In addition,RNA silencing serves as an important mechanism in plants' ...RNA silencing is a conserved mechanism found ubiquitously in eukaryotic organisms.It has been used to regulate gene expression and development.In addition,RNA silencing serves as an important mechanism in plants' defense against invasive nucleic acids,such as viruses,transposons,and transgenes.As a counter-defense,most plants,and some animal viruses,encode RNA silencing suppressors to interfere at one or several points of the silencing pathway.In this study,we showed that Pns12 of RGDV (Rice gall dwarf virus) exhibits silencing suppressor activity on the reporter green fluorescent protein in transgenic Nicotiana benthamiana line 16c.Pns12 of RGDV suppressed local silencing induced by sense RNA but had no effect on that induced by dsRNA.Expression of Pns12 also enhanced Potato virus X pathogenicity in N.benthamiana.Collectively,these results suggested that RGDV Pns12 functions as a virus suppressor of RNA silencing,which might target an upstream step of dsRNA formation in the RNA silencing pathway.Furthermore,we showed that Pns12 is localized mainly in the nucleus of N.benthamiana leaf cells.展开更多
文摘目前国内外对水稻瘤矮病毒(Rice gall dwarf virus,RGDV)的研究主要集中在其核衣壳蛋白和外衣壳蛋白的性质、结构及其与介体的亲和性、与介体传毒能力的关系等方面,本室在已有室内检测研究的基础上,利用RT-PCR和PVP-ELISA法对田间带毒叶蝉进行检测,并对2种方法的检测结果进行了比较.
基金supported by the National Science Foundation of China (30970135)The Key Project of Genetically Modified Organisms Breeding(2009ZX08009-044B)+1 种基金the Natural Science Foundation of Fujian Province of China (No.2006J0065)the Public-interest Scientific Institution Basal Research Fund of Fujian Province (2009R10029-3)
文摘To obtain the P8 protein of Rice gall dwarf virus (RGDV) with biological activity,its outer coat protein gene S8 was expressed in Spodoptera frugiperda (Sf9) insect cells using the baculovirus expression system.The S8 gene was subcloned into the pFastBacTM1 vector,to produce the recombinant baculovirus transfer vector pFB-S8.After transformation,pFB-S8 was introduced into the competent cells (E.coli DH10Bac) containing a shuttle vector,Bacmid,generating the recombinant bacmid rbpFB-S8.After being infected by recombinant baculovirus rvpFB-S8 at different multiplicities of infection,Sf9 cells were collected at different times and analyzed by SDS-PAGE,Western blotting and immunofluorescence microscopy.The expression level of the P8 protein was highest between 48-72 h after transfection of Sf9 cells.Immunofluorescence microscopy showed that P8 protein of RGDV formed punctate structures in the cytoplasm of Sf9 cells.
基金supported by the National Basic Research Program of China(Grant No. 2010CB126203)the National Transgenic Major Program(Grant Nos. 2009ZX08009-044B and 2009ZX08001-018B)+2 种基金the National Natural Science Foundation of China(Grant No. 30970135)the Fujian Province Education Department(Grant No. JB08078)Specialized Research Fund for the Ministry of Agriculture(Grant No. nyhyzx07-051)
文摘RNA silencing is a conserved mechanism found ubiquitously in eukaryotic organisms.It has been used to regulate gene expression and development.In addition,RNA silencing serves as an important mechanism in plants' defense against invasive nucleic acids,such as viruses,transposons,and transgenes.As a counter-defense,most plants,and some animal viruses,encode RNA silencing suppressors to interfere at one or several points of the silencing pathway.In this study,we showed that Pns12 of RGDV (Rice gall dwarf virus) exhibits silencing suppressor activity on the reporter green fluorescent protein in transgenic Nicotiana benthamiana line 16c.Pns12 of RGDV suppressed local silencing induced by sense RNA but had no effect on that induced by dsRNA.Expression of Pns12 also enhanced Potato virus X pathogenicity in N.benthamiana.Collectively,these results suggested that RGDV Pns12 functions as a virus suppressor of RNA silencing,which might target an upstream step of dsRNA formation in the RNA silencing pathway.Furthermore,we showed that Pns12 is localized mainly in the nucleus of N.benthamiana leaf cells.
