[Construction of a novel Bm NPV Bac to Bac system]

A Bi-Shuttle vector Bm-Bacmid was constructed by co-infecting Bm N cells with wild type genomic DNA from BmNPV and Ac-Bacmid DNA. It could not only replicate in E. coli cells as a large plasmid and but also remain infectious when induced into Bm N or Sf9 cells. Recombinant virus rBmHBe was obtained after transposition of a donor plasmid carrying Hepatitis Be antigen gene (HBeAg) into att Tn7, and was demonstrated by Southern blotting. SDS-PAGE analysis showed that HBeAg gene were highly expressed in Bm N cells. By ELISA testing, the highest antigenecity titer of HBeAg protein in cell cultural medium was up to a dilution of 1:32,000. Although HBeAg protein also presented in the Bm N cells the titer was only 1:2000. The HBcAg protein was much fewer than HBeAg (< 1:160) whatever in culture medium and in cells. The results showed that Bm N cells was able to recognize the signal peptide sequence and cut it correctly for HBeAg protein's excreting production.