## 4 Discussion
### miRNAs Involved in the Development of _G. sinensis_ Pods  
Thus far, diverse miRNAs have been validated as playing indispensable roles in plant development, for instance, _miR856_, _miR859_, _miR160_, _miR163_, _miR164_, _miR165_, _miR166_, _miR167_, _miR171_, and _miR172_ were showed to regulate pods, seeds, root development, and phase transitions (Table S8). In this investigation, _miR856_, _miR166_, and miR396 were differentially expressed in _G. sinensis_ pods with different developmental stages. Among them, the expression of the total members in the _miR8396_ family generally exceeded the other two miRNA families. However, _miR8396_, with six members, was expressed differently in the three stages (Table S3). Intriguingly, we found that _miR8396b-5p_ and _miR8396b-3p_ were the most abundant during the early stage of pod development but dramatically decreased in the latter two stages. Conversely, _miR8396a_ was highly expressed in the third stage (Table S6). A similar expression pattern was also found in the _miR156_ family in _G. sinensis_, suggesting that different members in same family may have different roles in the process of pod development. According to GO annotation and KEGG enrichment analysis, the targets of _miR8396b_, _miR8396b-3p_, and _miR156d-3p_ were enriched in the top 20 molecular functions, biological processes, or metabolic pathways (Figures S3 and S4). The _miR8396_ family, conserved among _A. thaliana_, _Populus trichocarpa_, and _Medicago truncatula_, was reported to regulate the development of leaves, seedlings, pods, and flowers by suppressing the targets encoding GRFs (growth-regulating factors) [48; 49; 50; 51]. In the model legume _Medicago truncatula_, _miR8396b_ was demonstrated to reduce root growth and mycorrhizal associations by targeting six GRFs and two _bHLH79_-like target genes [51]. Since the genome of _G. sinensis_ was unavailable, based on the transcription sequence, the target of _miR8396b_ from the pods was enriched in Golgi vesicle budding, lipid translocation, and membrane budding. _miR156d-3p_ was enriched in plant hormone signal transduction, and its targets were predicted to encode auxin response factor 18-like (ARF, gene ID: _G. sinensis_ Unigene1136, unpublished transcriptome data). In _Solanum Lycopersicum_ L., miR156 was predicted to target _SPL/SBP_ to negatively regulate fruit development [52]. However, _miR156d-3p_ may have a different regulation module for pod development in _G. sinensis_. It is well known that RNA-binding proteins (RBPs) and miRNAs are regarded as important factors for the suppression of the expression of mRNAs affecting multiple cellular functions during mRNA post-transcription processes [53; 54]. Polyadenylation is a two-step nuclear process including cleavage of the pre-mRNA at the 3'-end and the polymerization of a polyadenosine (polyA) tail to promote mRNA stability [55]. However, polyadenylation, which widely occurs during the degradation of intermediates and the production of polyA, further promotes mRNA degradation [56]. AU-rich elements (AREs)that can bind to the target genes of _miR2658_ are a kind of cis-acting destabilizing elements that dictate mRNA degradation [57]. In this investigation, _miR2658_ was highly expressed in May and was predicted to regulate the polyadenylation-dependent mRNA catabolic process, implying that _miR2658_ might regulate the ARE-mediated mRNA delay. Moreover, _miR2658_ was only expressed selectively in May, which might better regulate the growth and development of pod tissues. Furthermore, the targets of _miR89736_ were also significantly enriched in the KEGG pathway of photosynthesis and plant-pathogen interactions (Figure S3). Similar to the reduction in _miR156_ that changes the time of vegetative phase by increasing photosynthesis [58], _miR9736_, with a sharp increase in September, might change vegetative development in _G. sinensis_ pods.