Hypothesis testing of major transitions during diapause
The timing of termination of diapause in P. napi is regulated through the Prothoracicotropic hormone (PTTH)/20-hydroxyecdysone (20E) axis, where cold accumulation is needed in order to transition from diapause to post-diapause quiescence (Süess et al., 2022). A differential expression analysis was run comparing expression in both tissues at 24, 144, and 144 days in diapause, which are timepoints where diapause termination should be occurring based on prior work, and when pupae were also in cold temperatures (Lehmann et al., 2018; Lehmann et al., 2017). The differentially expressed genes (DEGs) from each list were then checked for overlap, and the overlap were considered candidate miRNAs.