Based on quantitative trait locus (QTL) mapping, the gene for porcine uncoupling protein 3 (UCP3) was chosen as a candidate gene for pig fat deposition and meat quality traits. In this study, a partial coding region of the UCP3 gene was sequenced and one single nucleotide polymorphism (cSNP) was found at 395 bp. The mutation was G→A and resulted in the amino acid change from glycine to arginine. This site was also recognized by restriction endonuclease SmaI. The UCP3 SmaI polymorphism was analysed among 186 individuals of Large White×Meishan F2 progeny using polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP). The genotypes of the UCP3 SmaI polymorphism were AA, AB and BB. The frequency of A and B alleles was respectively 0.56 and 0.44. Statistical analyses showed that the SmaI polymorphism in the F2 population was significantly associated with back-fat thickness at thorax–waist and buttock, as well as with intramuscular fat, drip-loss rate and water-holding capacity. The additive effect of UCP3 SmaI was clearly shown. The genotype AA reduced back-fat thickness and drip-loss rate, increased water-holding capacity, and decreased the intramuscular fat. The effect of the pig UCP3 SmaI polymorphism needs to be analysed in other populations using larger samples.