The enzyme β,β-carotene-15,15'-mono-oxygenase 1 (BCMO1) is responsible for the symmetrical cleavage of β-carotene into retinal. We identified a polymorphism in the promoter of the BCMO1 gene, inducing differences in BCMO1 mRNA levels (high in adenines (AA) and low in guanines (GG)) and colour in chicken breast muscle. The present study was designed to test whether this polymorphism could affect the response to dietary β-carotene. Dietary β-carotene supplementation did not change the effects of the genotypes on breast muscle properties: BCMO1 mRNA levels were lower and xanthophyll contents higher in GG than in AA chickens. Lower vitamin E levels in the plasma and duodenum, plasma cholesterol levels and body weight were also observed in GG than in AA chickens. In both genotypes, dietary β-carotene increased vitamin A storage in the liver; however, it reduced numerous parameters such as SCARB1 (scavenger receptor class B type I) in the duodenum, BCMO1 in the liver, vitamin E levels in the plasma and tissues, xanthophyll contents in the pectoralis major muscle and carcass adiposity. However, several diet × genotype interactions were observed. In the GG genotype, dietary β-carotene increased ISX (intestine-specific homeobox) and decreased BCMO1 mRNA levels in the duodenum, decreased xanthophyll concentrations in the duodenum, liver and plasma, and decreased colour index and HDL-cholesterol concentration in the plasma. Retinol accumulation following dietary β-carotene supplementation was observed in the duodenum of AA chickens only. Therefore, the negative feedback control on β-carotene conversion through ISX appears as functional in the duodenum of GG but not of AA chickens. This could result in a higher availability of β-carotene in the duodenum of GG chickens, reducing the uptake of xanthophylls, liposoluble vitamins and cholesterol.