The number of days (DN) when hens lay fertile eggs as well as the number of fertile eggs (FN) were produced after a single artificial insemination (AI), including the two duration of fertility (DF) traits. Indeed, they are the key production performance that associates with the production cost of hatching egg when its determination the interval between successive artificial inseminations. However, the relevant genes response for regulating the DF has not been uncovered yet. Therefore, we performed a weighted gene co-expression network analysis (WGCNA) to investigate the insight into co-expression gene modules on DF process in hens. The total mRNA was extracted from the utero-vaginal junction (UVJ, with the sperm storage function in hen’s oviduct which is the biological basis for DF) of 20 hens with several levels of DF traits, and performed transcriptome sequences of mRNA. As a result, three co-expression gene modules were identified to be highly correlated with DF traits. Moreover, the expression changes of top 5 hub genes in each module with DF traits were further confirmed in other 20 hens by RT-PCR. These findings highlighted the co-expression modules and their affiliated genes as playing important roles in the regulation of DF traits.