Aphidius gifuensis Ashmead is an excellent solitary endoparasitoid natural enemy, which can control a variety of aphids effectively, especially the aphid Myzus persicae (Sulzer). Diapause plays an important role in the overwintering of A. gifuensis. By simulating the environmental conditions of winter via controlling temperature and photoperiod in the laboratory, we induced the insects to enter diapause for the purposes of enhancing adaptive capacity and parasitic ability of offspring, increasing the production of the biological control agents for pest management. Transcriptome sequencing analyses were employed to compare expression differences between no-diapause (ND), diapause (D) and post-diapause (PD) groups. After RNA extraction and purification, cDNA synthesis, and establishment of cDNA libraries, a total of 40477 unigenes were obtained from 9 samples. We detected 458 differentially expressed genes (DEGs) between ND and D, and 298 DEGs between D and PD. And 59 diapause-associated genes were revealed to be significantly different in D/ND and D/PD, but no significant difference was found in PD/ND. With analysis of GO and KEGG enrichment on diapause-related genes, we found that these genes were primarily associated with cold resistance, autoimmune defense, lipid metabolism, insect cuticle protein and transcriptional regulation, which are the genes regulating the diapause process of A. gifuensis.