Transcriptomic regulation of sequential eggshell mineralization in laying hens: molecular mechanism underlying eggshell quality and production performance implications

Eggshell quality is a key economic trait in commercial layer production, which profoundly affects economic profits, flock elimination rate, and feeding management. Frequent occurrence of cracked, thin-shelled eggs has long plagued intensive laying hen farming. This study aimed to dissect the molecular mechanisms and regulatory networks underlying staged eggshell mineralization across the pre-mineralization, initial mineralization, and rapid mineralization phases in laying hens, and explore their links to eggshell quality and shell defect in production. By integrating RNA-sequencing, temporal expression profiling and WGCNA, we screened core functional genes and signaling pathways in uterus that governing the variation of eggshell thickness. In pre-mineralization stage, high expression of Ovalbumin ( OVAL ) may inhibit premature calcium carbonate crystallization and avoid abnormal shell development. During rapid mineralization, genes encoding eggshell matrix proteins, ATPase, TRP (transient receptor potential), SLC (Solute carrier) and collagen family members coordinately maintained uterine cellular calcium homeostasis, sustained calcium supply for eggshell deposition, and ultimately determined eggshell thickness and strength. Meanwhile, calcium concentration in uterine fluid increased gradually along with mineralization and remained stable throughout rapid mineralization phase, providing a steady calcium supply basis for preventing eggshell defects in commercial laying hen production. Findings of this study provided targets and theoretical support for the breeding of laying hen breeds, the regulation of nutritional management, and the optimization of farming environment management.