Identification of Genes Affected Blue Eggshell Coloration in Xuefeng Black-Bone Chickens

Blue eggshells have become economically valuable due to their higher popularity among consumers when compared with eggs of other colors, particularly in East Asia. However, eggshell colors vary widely and gradually become lighter with the increase of age. Therefore, to determine the association between DNA methylation and gene expression on the molecular mechanism of eggshell color variation in Xuefeng Black-bone chickens, we collected the shell glands from chickens that produced dark blue eggshells and light blue eggshells, respectively, and performed whole genome bisulfite sequencing (WGBS) and RNA sequencing (RNA-Seq) on them. The results showed that in the context of CG, DNA methylation levels were the highest compared to CHG and CHH contexts, and were negatively correlated with gene expression levels in three different gene regions in all samples, while there was no significant correlation in the contexts of CHG and CHH. Furthermore, when disregarding the location of coding genes, a total of 55 genes will show differences not only in expression levels but also in DNA methylation levels, and among the majority of differential expression genes (n=50), the DNA methylation levels in the gene body region were the highest. GO analysis and KEGG pathway analysis were performed in these genes and the results showed that the genes NR1H4 and HEPHL1, ABCA4 and ABCA12, and the pathways, cell adhesion molecular, ABC transporters, ECM-receptor interaction, and neuroactive ligand-receptor interaction, may affect the color variation of the blue eggshell of Xuefeng Black-bone chickens.