Identification and analysis of key miRNA for sex differentiation in hermaphroditic Juglans mandshurica Maxim.

Background: Juglans mandshurica Maxim. is a monoecious heteromorphic plant. The mechanism of miRNA-mediated sex differentiation and dioecious heterozygosity is unclear. Results: ： In this study, 310 known and 278 unknown miRNA were identified and 129 differentially expressed miRNA were screened using high-throughput sequencing technology for microRNA sequencing analysis of male and female flower buds of the protogyny and protandry of Juglans mandshurica . 638 miRNA target genes predicted using PSRobots and these target genes are significantly enriched in reproductive, floral development and phytohormone signaling pathways. Therefore, it is hypothesized that miR159c-3p, miR156d-3p, miR172d, miR171b, miR164a, miR167a, miR319a, miR399a, and miR396t et al. may be the key miRNA affecting sex differentiation in Juglans mandshurica, among which miR164a, miR166c, and miR171c-5p may be the key miRNA for heterosexualism differentiation in Juglans mandshurica . 9 randomly selected miRNAs with significant differences were subjected to real-time fluorescence quantitative PCR to verify the accuracy of high-throughput sequencing. Combined with transcriptome sequencing results for co-analysis, a differential miRNA-differential mRNA primary regulatory network was established. Conclusion: ： It was further confirmed that these miRNA play an important role in the sex differentiation of Juglans mandshurica. This study lays the research foundation for the eventual discovery of the molecular mechanism of sex differentiation and heterosexualism in J uglans mandshurica.