Transcriptomic Analysis of the Uterine Mucosa Reveals a Gene Regulatory Network Associated with Reduced Eggshell Strength in Late-Laying Hens

The decline in eggshell strength (ESS) during the late laying period is an important issue that needs to be addressed in the laying hen industry. Given that eggshell formation depends heavily on uterine mucosa secretions, understanding the underlying transcriptomic changes is essential. To investigate this, eggshell phenotypes were compared between the peak laying period (30 weeks of age, 30 W) and the late laying period (65 weeks of age, 65 W) of White Leghorn laying hens. Furthermore, RNA sequencing of the uterine mucosa was performed, and the sequencing data were quality-controlled, aligned to the chicken reference genome (GRCg7b), quantified using StringTie, and analysed for differential expression using DESeq2. WGCNA was subsequently performed to identify candidate genes associated with eggshell quality. The results showed that ESS decreased from 31.31 ± 3.34 N in the 30 W group to 24.65 ± 2.11 N in the 65 W group (p < 0.001). Transcriptomic analysis identified 903 differentially expressed genes (DEGs) in the uterine mucosa, and these DEGs were significantly enriched in pathways related to eggshell mineralisation. By integrating WGCNA, protein–protein interaction (PPI) network analysis, and trait correlation analysis, six candidate genes were identified: GNAS, BMPR2, EDN2, ANXA1, PLCB2, and CX3CL1. In conclusion, this study identifies new candidate genes and provides a theoretical basis for elucidating the molecular mechanisms underlying the decline in eggshell quality in late-laying hens.