Small RNA sequencing identify miRNAs involved in pomegranate female sterility

Pomegranate pistils were collected from 12-year-old ‘Tunisiruanzi’ (also named ‘Tunisia’) trees grown in nursery of the Zhengzhou Fruit Research Institute located in Zhengzhou, Henan, China, and managed using conventional methods. Solitary flowers located in perennial branches, which contained high ratio of BFs, were selected as the sampling location of BFs’ pistils. Lateral flowers located in annual branches, which were observed with high ratio of FMFs, were selected for FMFs’ pistils. On the other hand, our previous studies showed that the putative cause of pomegranate female sterility was ovule development aborting after the formation of the inner integument primordium . The key stage for the termination of FMF ovule development was during the bud vertical diameter (BVD) was 5.0–13.0 mm. Pistils of FMFs at the pre-stage of FMFs’ ovules aborted (BVD 3.0–5.0 mm), the key stage of FMFs’ ovules aborted (BVD 5.0–13.0 mm), the stage after FMFs’ ovules aborted (BVD 5.0–13.0 mm) were collected for related miRNAs mining, pistils of BFs at the corresponding periods were selected as controls. In order to denote the accessions clearly, we used TNSI, TNSII, and TNSIII represent the BFs’ (Bisexual flowers) pistils when their BVD was 3.0–5.0, 5.1–13.0, 13.1–25.0 mm, respectively. Similarly, the designations ATNSI, ATNSII, and ATNSIII were used to represent the FMFs’ (Functional male flowers) pistils when their BVD was 3.0–5.0, 5.1–13.0, 13.1–25.0 mm, respectively , each stage with three repetitions.