Construction Of Recombinant BmNPV Containing EcobNPV Helicase Gene And Sequence Analysis Of BmCPV Genome Segment 2

In this study, we successfully cloned Ectropis obliqua nuclear polyhedrosis virus(EcobNPV) 73.7-78.9kb region applied PCR, and got the whole sequence of this region. Sequence analysis revealed that this region coding 3 proteins include odv-e25, odv-e28 and P143 of the virus. Motif analysis of P143 showed that this protein has a typical helicase uncoiling domain, the phylogenetic tree based on the amino acid sequence of P143 showed that EcobNPV belong to nuclear polyhedrosis virus, GROUPâ…¡, and the evolution of insect baculovirus seems to be host-associated.We successfully cloned and sequenced the whole sequence of the segment 2 of the Bombyx mori cytoplasmic polyhedrosis virus suzhou strain(BmCPV-suzhou), (GenBank Accession Number:GQ924586). Sequence analysis indicated that the segment 2 consist of a single open reading frame (ORF) coding RNA-dependent RNA polymerase (RDRP) of the virus and with the conserved terminal sequences AGUAA and GUUAGCC at the 5'and 3'end respectively. Phylogenetic tree of the family Reoviridae constructed based on the amino acid sequence of RDRP from different genus of Reoviridae indicate that the different strain of BmCPV and LdCPV-1,and DpCPV within the same group.EcobNPV is pathogenic to tea looper caterpillar, Ectropis obliqua, and has become an important biological pesticide against the pest. We constructed a recombinant transposition vector pFastBacTMDual-polh-helicase contain the BmNPV polyhedrin expression cassette and EcobNPV helicase expression cassette, and generation recombinant virus BmBacmid-polh-helicase applied the Bac-to-Bac system. Molecular biology identification and infection experiment indicated that the recombinant virus contains the EcobNPV helicase expression cassette, and could format polyhedra in the Cell Nucleus, and the recombinant virus contains double sets of helicase expression cassettes replicated normally.