The Study Of Using DsDNA To Induce The Gene Of Viral Binding Protein Of Myzus Persicae Silence In Order To Stop The Transmission Of PVY

Plant virus can transform by victor through interaction. The protein RPS2of Myzus persicae is a ribosomal protein, and has been comfirmed that it is useful for Potato Virus Y transmission by Myzus persicae.Using the GenBank to find the protein sequence of RPS2that have been reported in Myzus persicae,and then get the gene sequence of RPS2in Myzus persicae.Primers were designed based on the experimental purp-oses,amplified the complete sequences of RPS2gene by RT-PCR. Conne-cted the complete sequences of RPS2to the PGEM-T easy vector and transformed into Escherichia coli (E. coli) strain named NM522in order to amplify and save it. At the same time, three different couple of primers were designed to amplify the forepart(RPS2-1),the midpiece (RPS2-2) and the end-piece(RPS2-3) of the complete sequences of RPS2by PCR using the complete sequences of RPS2as a template.In order to build the vector,we introducted EcoRl on the5’end and HindⅢ on the3’end of RPS2-1,introducted EcoRl on the5’end and BamHI on the3’end of both RPS2-2and RPS2-3.Three fragments were connected to the LITMUS28i plasmid by digestion, ligation, and other molecular experi-mental methods commonly used.Then,we got three prokaryotic expres-sion vectors which can express dsRNA.The recombinant vector plasmid was consistent with expected results by PCR analysis, restriction enzyme digestion and DNA sequencing.Transformed three recombinant vectors into Escherichia coli (E. coli) strain named HT115(DE3).There are two reverse T7promoter in LITMUS-28i plasmid, while HT115is RNA enzyme Ⅲ-deficient strains, so after IPTG induction, the cells of HT115can express dsRNA and the dsRNA can be extracted by the CTAB method.The size of fragment extracted, DNase and RNaseA restriction results can be confirmed that the three vectors can express the dsRNA as expected.It proved that the artificial diet with these three dsRNAs had no impact on growth and development of Myzus persicae. The experiments above laid the foundation for evaluateing the effect of PVY spread by Myzus persicae.