Objective The chicken chr3:100005430 T > G locus had been proposed as a potential lethal mutation. This study aimed to investigate its population genetic structure across diverse local chicken breeds and to evaluate its effects on egg production and quality traits, thereby assessing its potential as a genetic marker for molecular breeding in local chickens.

Method A total of 1 380 blood samples from 25 Chinese local chicken breeds and one introduced breed, along with 225 dead-embryo samples from Wuhua Yellow chickens, were collected. Genotyping was performed using PCR-RFLP. In Wuhua Yellow chickens (n = 672), egg-production traits including age at first egg, egg number at 33, 43 and 52 weeks, average clutch length, and average pause length were recorded. Egg quality traits at 43 weeks were measured, covering egg weight, egg shape index, eggshell strength, eggshell thickness, yolk color, Haugh unit, and yolk ratio. Chi-square and Fisher's exact tests were used to analyze genotype frequency distribution and Hardy?Weinberg equilibrium; independent-samples t-test and Mann?Whitney U test were applied to compare trait differences between TT and TG genotypes; a linear model was employed to estimate the allele substitution effect and phenotypic variance explained.

Result Three genotypes (TT, TG, and GG) were identified in the 1 380 samples, with frequencies of 57.32%, 41.81%, and 0.87%, respectively. The GG genotype was detected for the first time in live individuals of several breeds including Wuhua Yellow chicken, Guangxi Yellow chicken, and Luxi gamecock. Moreover, two GG genotypes were identified among the 225 dead-embryo samples. In Wuhua Yellow chickens, no significant differences (P > 0.05) were observed between TT and TG genotypes for any egg production or egg quality traits. The 52-week egg number was 146.32 (±41.02) for TT and 142.71 (±45.77) for TG genotypes. Similarly, egg quality parameters such as eggshell strength (TT: 3.23 ± 0.70 kg/cm²; TG: 3.21 ± 0.53 kg/cm²) did not differ significantly. Genetic effect analysis demonstrated that the phenotypic variance explained by this locus for all traits was below 0.81%, with only minor allele substitution effects.

Conclusion The chr3:100005430 T > G locus is not a lethal mutation. It has no significant influence on egg production performance or egg quality in Wuhua Yellow chickens, indicating limited direct value as a molecular marker for breeding selection.