Detection of SPFMV and SPVG by using reverse dot blot hybridization system
[Objective]Two sweet potato viruses,feathery mottle virus (SPFMV) and virus G (SPVG),were cloned,and the reverse dot blot hybridization system was built.Then this system was used for detecting the sweet potato virus so as to provide references for producing high quality virus-free seedlings.[Method]According to the published nucleotide sequences of SPFMV and SPVG,two specific primers were designed and two fragments were cloned by reverse transcription-polymerase chain reaction (RT-PCR) from total RNA extracted from sweet potato leaf which were suspiciously infected with sweet potato virus.Then,the two fragments and Actin gene were used as the probes in this reverse dot blot hybridization system.[Result]Two fragments,about 310 and 500 bp were obtained.It was found by sequencing and blasting that they were coat proteins of SPFMV and SPVG with the homology of 97％ and 99％,respectively.Reverse dot blot hybridization of recombinant plasmids pUC-SPFMV and pUC-SPVG showed that single signals were attained from different viruses.No cross signals were found.Then the infected and virus-free sweet potatoes were detected by reverse dot blot hybridization using probes.The results showed that single signals were found in the affected samples while no signal was observed in virus-free samples,which accorded with RT-PCR results.[Conclusion]SPFMV and SPVG can be detected by this hybridized system and there was no cross signal from the sample with virus.So this system can be used to detect virus of virus-free sweet potato seedlings in the initial stage.
sweet potatoSPFMVSPVGvirus detectionreverse dot blot hybridization
国家科技期刊平台
NSTL
万方数据
维普
