Fecundity improvement is one of the most important economic traits for the swine industry as it significantly increases production efficiency. Intriguingly, chitosan oligosaccharide (COS), a biomaterial with an active amino group, could promote sow reproductive performance. Therefore, we investigated the effects of dietary COS supplementation on the gene expression differences in the ovaries of sows using the RNA-Seq method. This analysis obtained 13 960 051 and 14 564 863 clean reads in control ovary and COS ovary libraries, respectively. A total of 486 differentially expressed genes (DEGs) were thereby identified (FDR ≤ 0.001, |log₂ ratio| ≥ 1). There were 234 up-regulated and 252 down-regulated genes in the COS ovary samples compared with the control ovary samples. A large number of these DEGs were involved in the terms cellular process, cell & cell part and binding. Furthermore, pathway analysis indicated that these DEGs were significantly enriched in 34 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, including cell cycle, progesterone-mediated oocyte maturation, metabolic pathways, oocyte meiosis, and hematopoietic cell lineage among others. These results provided the molecular mechanisms of using COS feed additive for improving sow litter size and prolificacy.