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Objective To construct a recombinant secretory plasmid of Bacillus Calmette-Guerin(BCG) Ag85B-fused EB virus LMP2A and BZLF1 genes,and then transform it into BCG.Methods BCG-Ag85B signal sequence and Z2A gene were amplified from the genome of BCG and Z2A by PCR,respectively.BCG-Ag85B signal sequence was cloned into E.coli-BCG shuttle-vector pMV261 to obtain pMVS.The new recombinant plasmid of pMVZ2A was constructed by inserting the Z2A gene into pMVS and transformed into BCG by electrotransformation.The recombinant proteins were analyzed by SDS-PAGE.Results The cloned genes of BCG-Ag85B and Z2A were correctly inserted into the vector pMV261.The recombinant plasmid of pMVZ2A was identified by double restriction enzyme digestion,PCR amplification and gene sequencing.Conclusions pMVZ2A effectively expressed Z2A protein in BCG.The study provided a basis for BCG reconstruction and the development of new bivalent vaccine agaisnt EB virus and mycobacterium tuberculosis.
Objective To construct a recombinant secretory plasmid of Bacillus Calmette-Guerin (BCG) Ag85B-fused EB virus LMP2A and BZLF1 genes and then transform it into BCG. Methods BCG-Ag85B signal sequence and Z2A gene were amplified from the genome of BCG and Z2A by PCR, respectively. BCG-Ag85B signal sequence was cloned into E. coli-BCG shuttle-vector pMV261 to obtain pMVS. The new recombinant plasmid of pMVZ2A was constructed by inserting the Z2A gene into pMVS and transformed into BCG by electrotransformation. recombinant Proteins were analyzed by SDS-PAGE. Results of The cloned genes of BCG-Ag85B and Z2A were correctly inserted into the vector pMV261. The recombinant plasmid of pMVZ2A was identified by double restriction enzyme digestion, PCR amplification and gene sequencing. Conclusions pMVZ2A effectively expressed as Z2A protein in BCG. The study provided a basis for BCG reconstruction and the development of new bivalent vaccine agaisnt EB virus and mycobacterium tuberculosis.