Identification of microRNAs correlated with citrus granulation based on bioinformatics and molecular biology analysis

• 12 potentially unreported miRNAs were predicted to exist in citrus.
• Two miRNA families (miR397, miR828) involve in lignin biosynthesis pathway.
• csi-miR397 and N-miR828 were negatively related to citrus granulation.
• csi-miR397/Cs6g06890.1 and N-miR828/Cs1g17590.1 modules relate to granulation.

Juice sac granulation leads to large postharvest losses of citrus fruits. Although the fundamental mechanism of this process remains not fully understood, some previous studies have clearly shown the close association between lignin metabolism and juice sac granulation. While miRNAs have been proved as main regulators of target gene expression which regulates lignin biosynthesis in some plant species, the role of miRNAs in citrus granulation was still unknown. To evaluate the role of miRNAs in granulation, homology search-based computational approach was employed to identify miRNAs in citrus, and 12 potentially unreported miRNAs in citrus were obtained. Through the identification and annotation of target genes, two miRNA families (miR397, miR828) were shown to be involved in lignin biosynthesis pathways. The expression pattern of miRNAs at different granulation levels was further investigated by real-time quantitative reverse transcription PCR (qRT-PCR). The results demonstrated that the expression of csi-miR397 and N-miR828 were negatively related to citrus granulation, with correlation coefficients of 0.888 and 0.944, respectively. A further study on their targets by qRT-PCR revealed the expressions of Cs6g06890.1 and Cs1g17590.1 were positively correlative to granulation, with correlation coefficients of 0.917 and 0.963, respectively. The correlation analysis among the expression of miRNA, its target, lignin content and granulation degree indicates a likely role of the csi-miR397/Cs6g06890.1 and N-miR828/Cs1g17590.1 modules were correlate to granulation through regulation of the internal lignin activity. The results of this study lay a foundation for miRNAs in regulating lignin processes which relate to citrus granulation on post-transcriptional level.