Study On Expression Of Plasmodium Falclparum Merozoite Protein1(C-Terminal19kD) In Baculovirus/Silkworm Surface Display System

Malaria is a main global acute parasitic vector-borne infectious diseases whichcaused by a parasite, Transmitted by Anopheles anthropophagus. Clinical symptomsof plasmodium merozoite occurs in blood-stage, the current hot spots in malariavaccine research was mainly based on blocking this fission stage of Plasmodiumclones to prevent malaria infection. After the merozoite spore infecting red bloodcells,The merozoite surface protein1C-fragment (MSP1_19C) is still exist in the redblood cells membrane surface, T-cells and B-cells can be induced by the organismimmune response, Therefore, this protein is one of the most important of the malariavaccine candidate antigens sites. Silkworm baculovirus surface display technology isdeveloped in recent years, it is a new form of eukaryotic display technology.We fusedThe exogenous gene and Bombyx mori baculovirus glycoprotein gene gp64, canrealize the purpose of displaying the target proteins in the virus membranesurface.This technology has become a research hotspot in the single antibody,polyclonal antibody preparation, new vaccine and other fields in recent years.This study is to connect the MSP1_19cgene segments to the eukaryotic vectorpFastBac1-gp64, we successfully construted eukaryotic donor plasmidpFastBac1-gp64-MSP1_19c. The donor plasmid pFastBac1-gp64-MSP1_19ctrans into E.Coli DH10Bac competent cells after transposable with Silkworm bacmid in E. ColiDH10Bac cells,So we can get recombine Bacmid-gp64-MSP1_19c. Using the method ofliposome-mediated transfection, The Bacmid-gp64-MSP1_19ctransfected in silkwormBmN cells. After assembling in the BmN cells, it forms a recombinant baculovirus,named BmNPV-gp64-MSP1_19cand the recombinant baculovirus copy and expandedin BmN cells. Detected by PCR and Western blot technique both in BmN cells andsilkworm, Bombyx Mori silkworm have been successfully transcribed and expressed.Use the third generation of BmNPV-gp64-MSP1_19cto secondary infections of thevirus（MOI=10） complex vaccination chrysalis, after7days collecting thepathogenesis of silkworm. Bi-directional electrophoresis and Western Blottingshowed that recombinant baculovirus MSP1_19cprotein expressed in Silkworm pupa,proof of recombinant baculovirus BmNPV-gp64-MSP1_19crestructuring success.Purification of recombinant virus particle immunity New Zealand male rabbit toproduce serum antibody of rabbits, After ProteinA antibody purification columnpurification, Discovered by ELISA detection antibody titer up to1:8,000, indicating that can effectively encourage the body to produce antibodies.This study confirmed the Bombyx mori baculovirus surface display technologycan effectively displayed Plasmodium falciparum protein MSP119cin Bombyx moribaculovirus envelope surface. And the recombinant virus as immunogens canstimulate the body to produce antibodies and immune cells, which laid the Foundationfor a new malaria vaccine.