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In plants, sex determination is a comprehensive process of correlated events,which involves genes that are differentially expressed in distinct developmental phases.Exploring sex determination in gymnosperm species will contribute to understanding sex determination and the evolution of sex chromosomes.In this study, we conducted RNA-sequencing of female and male buds (CY and XY) as well as ovulate strobilus and staminate strobilus (CH and XH) of Ginkgo biloba to gain insights into the genes potentially related to sex determination in this species.Approximately 60 Gb of clean reads were obtained from four cDNA libraries.De novo assembly of the clean reads generated 108 307 unigenes with an average length of 796 bp.Among these unigenes, 51 953 (47.97%) had at least one significant match with a gene sequence in one of the seven public protein and nucleotide databases searched.A total of 4 709 and 9 802 differentially expressed genes (DEGs) were identified in the XY vs.CY and XH vs.CH comparisons, respectively.Genes and transcription factors associated with plant hormone signal and transduction as well as DNA methylation were assumed to involved in sex determination of G.biloba.High expression level correlation of 26 randomly selected genes was found between the RNA-Seq and quantitative real-time PCR results.The transcriptome resources that we generated allowed us to characterize gene expression profiles, examine differential expression profiles, and identify functional genes that may be associated with sex determination in G.biloba.Our results illustrate the dynamic nature of sex determination in G.biloba at the transcriptome level and confirm previous observations that sex determination is a complex process.