Reproductive efficiency of a heifer is directly dependent on the health of reproductive system especially ovary and uterus. Current experiment was carried out to screen candidate genes related to reproductive traits and detect molecular mechanisms of DEGs by using RNA-sequencing (RNAseq) and Gene Set Enrichment Analysis (GSEA) to understand the environmental adaptability, molecular breeding and analyze the reproductive performance. Furthermore, Illumina II high throughput sequncing technology was utilized in the current experiment to sequence transcriptomes and analysis related to pathway of different genes between uterus and ovary in Xiangxi cattle. The total number of ovary assembled was compared to the reference genome sequence, DEGs analysis, GO and KEGG enrichment analysis and candidate gene screening were performed. Based on the transcriptome data, seven cDNA libraries were constructed by utilizing total RNA. Out of seven contracted cDNA libraries, in every sequencing library, raw reads reached >133 Mb, and >94% clean reads remained. After comparing the Xiangxi cattle ovaries with the uterus, a total of 3635 mRNAs were found, in which 1608 were significantly up-regulated expressed and 2027 were down-regulated expression mRNAs. The mRNA expression of ADCY3, ADCY4, ADCY9, FSHR, BMP15, LHCGR, INSR, CYP11A1, CYP17A1, CYP19A1, STAR and SCARB1 in ovary of Xiangxi Cattle were significant different in ovarian and uterus tissues. Through GSEA and PPI network, we found the ovarian steroidogenesis pathway played significant role in signal regulation during the development of the ovarian tissue of the Xiangxi cattle