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Expression And Localization Of Dendrolimus Puntatus Cytoplasmic Polyhedrosis Virus(DpCPV) P44 And P50

Posted on:2016-10-03Degree:MasterType:Thesis
Country:ChinaCandidate:H PengFull Text:PDF
GTID:2283330470465697Subject:Biochemistry and Molecular Biology
Abstract/Summary:PDF Full Text Request
Dendrolimus punctatus cytoplasmic polyhedrosis virus(DpCPV) is one kind of pathogenic microorganism of forest pests, dendrolimus punctatus, which is endemic species in China. The application of DpCPV in agriculture is mainly used as microbial pesticide, which is safe, efficient and non-polluting properties. At present, research on DpCPV focuses on the analysis of the nucleotide sequence and structure of the virus, but the studies of the structure and function of protein encoded by the genome is less. To study DpCPV structural protein modification mechanism, will help us to understand the mechanisms of viruses, and providing a theoretical basis for the prevention and control of Dendrolimus punctatus and the production of virus preparation.In this research, prokaryotic and eukaryotic expression vector of protein P44 and P55 from DpCPV have been constructed, respectively, to study the interaction between P44 and P55. And the intracellular localization of P50 proteins in insect cells was studied.The results showed that the protein encoded by DpCPV s7 and s8 were induced and expressed in E.coli, wiht molecular size consistent with the theory size. Polyclonal antibody was prepared by immunized rabbit, respectively. Protein P50 and P44 were coexpressed through Bac-s8-s7-eGFP in Sf9 insect cells. P50 is ~50 kDa, consistent with that of prokaryotic expression results. P44 is ~35 kDa, slightly smaller than that of expressed by prokaryotic system, which indicated that P44 could not self-cleavage and be modified. And the results showed that the protein P44 was not involved in the cutting modification of protein P50. P50 was found to be located in the cytoplasm by using a laser confocal microscope to investigate the locazition of P50-eGFP.We studied the interaction between proteis encoded by different DpCPV segment, and P50 protein positioned in insect cells has been explored. The obtained molecular data will help to study the splicing mechanism of P50 and the interaction between constitutive protein and non-structural protein of DpCPV in further studies. And P50 protein localization experiments result provides a reference method for other study of RNA viruses protein localization. Results of this experiment provided relevant experimental basis to explore the mechanisms of insect infestation by DpCPV.
Keywords/Search Tags:Dendrolimus punctatus cytoplasmic polyhedrosis virus(DpCPV), Polyclonal antibodies, Prokaryotic expression, Eukaryotic expression
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