亚洲玉米螟保幼激素结合蛋白基因OfJHBP的鉴定及在生长发育及滞育中的功能分析OA

[Aim]To identify the juvenile hormone binding protein(JHBP)gene of Ostrinia furnacalis(OfJHBP),and explore its function in growth,development and diapause of O.furnacalis.[Methods]Based on the transcriptome database of O.furnacalis larvae at different diapause statuses assembled by our laboratory(NCBI accession number:PRJNA1030200),the open reading frame(ORF)sequence of OfJHBP was cloned and identified by RT-PCR.The molecular weight and isoelectric point of OfJHBP of O.furnacalis were predicted using the Expasy-ProtParam online tool.The OfJHBP-encoded amino acid sequence of O.furnacalis was aligned with those of JHBPs from other insects using DNAMAN 6.0 software.A phylogenetic tree was constructed using the neighbor-joining(NJ)method in MEGA 11 software.The expression levels of OfJHBP in the 5th instar larvae of O.furnacalis at non-diapause,diapause and diapause-terminated stages were determined by qPCR.The expression level of OfJHBP was measured by qPCR at 48 h after OfJHBP interference by injecting dsRNA into the diapausing 5th instar larvae of O.furnacalis.The triglyceride content in larvae was detected at 96 h after OfJHBP interference by injecting dsRNA into the diapausing 5th instar larvae of O.furnacalis.The body weight,diapause rate,mortality rate and phenotypic changes of O.furnacalis were observed and determined daily within 9 d after dsRNA injection.[Results]The 726 bp ORF sequence of OfJHBP(NCBI accession number:XM_028317397.1)of O.furnacalis was obtained.OfJHBP encodes 241 amino acids.The predicted molecular weight of OfJHBP was 27.06 kD,and its theoretical isoelectric point was 5.99.OfJHBP shared the highest amino acid sequence identity(99.17％)with OnJHBP from O.nubilalis.The evolutionary relationship between OfJHBP and OnJHBP was the closest.OfJHBP was expressed in the 5 th instar larvae at different diapause stages,with the highest expression level observed in diapause stage and lower expression levels in non-diapause and diapause-terminated stages.Compared to the dsGFP-injected control group,dsOfJHBP injection resulted in a 78.32％down-regulation of OfJHBP expression level at 48 after injection,a 1.18-fold increase in the triglyceride content at 96 after injection,as well as a mortality rate of 46.67％,a proportional reduction in the body weight,a 53.33％decrease in the diapause rate,developmental abnormalities,various malformations leading to death at different degrees at 9 d after injection,and surviving individuals predominantly developing into black pupae that failed to complete adult eclosion.[Conclusion]OfJHBP is closely associated with the growth and development of O.furnacalis and likely plays a crucial role in diapause process of O.furnacalis.