Differential expression of early viral gene BmORF51 in Bombyx mori nucleopolyhedrovirus infection of resistant and susceptible silkworms

Abstract

Open reading frame 51 of Bombyx mori nucleopolyhedrovirus (Bm51) is a homologue of autographa californica multiple NPV ORF63. In this study, the expression profiles of Bm51 in the resistant strain NB and the susceptible strain 306 were characterized, and Bm51 gene was amplified from BmNPV genomic DNA by polymerase chain reaction (PCR) and cloned into Escherichia coli expression vector pET-30a (+). The recombinant His-tagged Bm51 protein was expressed in E. coli BL21 (DE3) and purified by metal chelating affinity chromatography to produce antibodies against Bm51 protein. The amino acid sequence of recombinant protein was confirmed by mass spectroscopic analysis. The transcription and protein product of early viral gene, Bm51, was detected at 6 h post-infection (p.i.) in resistant strain NB by quantitative real-time (qRT)-PCR and western blotting, and the expression of Bm51 in NB reached the maximal level at 36 h p.i. in NB, and then gradually decreased to undetectable level at 72 h p.i. In contrast, the Bm51 protein was undetectable until 12 h p.i. in susceptible strain 306 and the expression of Bm51 progressively increased during the 72 h post-infection.