Transcription analysis of serine protease gene BmSP25 in Bombyx mori and its immune response

ObjectiveSequence and transcription state of serine protease(SP) gene BmSP25 were analyzed, the im-mune response mechanism of its expression pattern against Bombyx mori nucleopolyhedrovirus(BmNPV) were defined, in order to provide theoretical basis for exploring the function of BmSPs in immune responses of B. mori. MethodThe se-quence of BmSP25 gene was cloned, bioinformatics was used to analyze the amino acid sequence, molecular weight, func-tional domain of the coding region of this gene. Multiple sequence alignment and phylogenetic analysis of BmSP25 amino acid sequence were conducted by GeneDoc and MEGA5.0 software. BmSP25 gene transcription in different tissues and at different development stages were analyzed by semi-quantitative PCR, and transcription level of BmSP25 mRNA in intesti-nal tissue inoculated by BmNPV were detected by real-time quantitative fluorescence PCR. ResultThe full length of BmSP25 open reading frame (ORF) was 885 bp, encoding 294 amino acids, and from the 1st to the 17th ones were signal peptides. The molecular mass without signal peptides were 29.1 kDa and theoretical isoelectric point was 7.8. BmSP25 protein was composed of 4 α-helices, 15 β-folds and some irregular coils. Homology alignment of amino acid showed that BmSP25 (BGIBMGA008514-PA) shared the highest homology(62.1%) with SP sequence of Mamestra configurata(GenBank acces-sion number: ADM35105). BmSP25 gene was specifically expressed in midgut tissues and continuously expressed at the whole larval stage of B. mori. The transcription level of BmSP25 gene was changed after BmNPV inoculation, BmSP25 mRNA was down-regulated at 6 h, but it was up-regulated obviously at 3, 12 and 24 h of inoculation. ConclusionBmSP25 plays an important role in the process of immune response from intrusion of BmNPV in B. mori. Since insect SP contains highly conserved specificity sites of substrate, substrate analogues and gene site-directed mutation can be utilized to control agricultural and forestry pests.