Genomic insights into egg production in Mahuang chickens: identification of key loci and candidate genes through whole genome sequencing and validation

Genome-wide association studies provide a framework for identifying candidate genes linked to economically significant egg production traits in poultry. This study aimed to elucidate genetic variations influencing key egg production traits, specifically total number of eggs laid, number of eggs laid at 43 weeks, normal egg count, and normal egg rate in Mahuang chickens. Employing a low-coverage whole genome sequencing approach, we generated a comprehensive dataset encompassing 14 million imputed single-nucleotide polymorphisms (SNPs) from 647 individuals, complemented by high-depth sequencing (30×) of 40 chickens. To refine our association signals and address potential linkage disequilibrium (LD), we conducted conditional analyses by incorporating the most significant SNPs within each locus as fixed-effect covariates in the models. This approach allowed us to distinguish independent causal variants from those associated due to LD, leading to more accurate identification of true genetic effects. Our findings uncovered numerous novel SNPs and candidate genes, including the first reported rs7535083 in the LAMC2 gene, which introduces a missense variant (p.Gln726Arg) with potential implications for protein functionality and influence on egg production traits. Enrichment analyses identified significant candidate genes such as HEBP2, LAMC1, and LAMC2, integral to cellular proliferation pathways essential for ovarian follicle development and egg yield. Subsequent validation efforts included Kompetitive Allele Specific PCR (KASP) genotyping and quantitative real-time PCR (qRT-PCR) analysis. KASP assays successfully genotyped 9 out of 11 selected SNPs, confirming allele frequencies consistent with initial findings, particularly highlighting the rs318026917 SNP's association with the total number of eggs laid. Concurrently, qRT-PCR analysis revealed significant expression variations among candidate genes, emphasising the pivotal roles of LAMC2 and RASA3 in egg production traits. Together, these results identify specific SNPs and candidate genes associated with egg production traits in Mahuang chickens, including a novel missense variant in LAMC2, and establish validated genetic targets for marker-assisted selection and further functional investigation.