Abstract:  【Aim】 Insect diapause is a critical eco-physiological adaptation for seasonal survival, but little is known about the molecular mechanisms of summer and winter diapause of Delia antiqua. Here, our main goal is to identify candidate genes linked to pupal diapause of D. antiqua. 【Methods】 A straightforward in silico approach was performed to predict the secreted diapause-related proteins based on the RNA-seq and digital gene expression datasets. Amino acid sequence similarity and basic physiochemical characteristics analysis were conducted. Gene expression levels of 12 selected genes from RNA-seq data were further validated in non-diapause pupae, and different developmental stages of summer and winter diapause pupae by qRT-PCR. 【Results】 A total of 38 putative secreted proteins were differentially regulated between the summer and winter diapause pupae of D. antiqua. Gene Ontology analysis revealed differentially expressed genes to be associated with odorant binding, chitin and lipid metabolic processes, innate immunity and development, etc. Among them, regulation of lipid mobilization and chitin-related metabolism was found to be important during diapause development. qRT-PCR analyses showed that the expression profiles of the 12 selected genes were in good agreement with results from RNA-seq (R=0.862, P=0.001). 【Conclusion】 This study establishes a robust pipeline for the discovery of diapause-related genes by secretome mining based on the high-throughput sequencing.

Key words: Delia antiqua, diapause, secreted protein, transcriptome, digital gene expression, RNA-seq