Several quantitative trait loci (QTL) affecting muscle glycogen content andrelated traits were mapped to pig chromosome 15 using a three-generationintercross between Berkshire x Yorkshire pigs. On the basis of the QTL locationthe PRKAG3 (protein kinase, AMP-activated, gamma(3)-subunit) gene was consideredto be a good candidate for the observed effects. Differences in the PRKAG3 genesequences of the founder animals of the intercross were analyzed. The RN(-)mutation previously reported was not present in the cross but three missensesubstitutions and a polymorphic short interspersed element (SINE) wereidentified. To confirm the hypothesis that at least one of these mutations wasassociated with differences in meat quality, >1800 animals from severalunrelated commercial lines were genotyped for the candidate substitutions and anassociation study was performed. The results demonstrate the presence of neweconomically important alleles of the PRKAG3 gene affecting the glycogen contentin the muscle and the resulting meat quality. Haplotype analysis was shown toresolve the effects of PRKAG3 more clearly than analysis of individualpolymorphisms. Because of their prevalence in the more common commercial breeds,the potential implications for the pig industry and consumers are considerablygreater than the original discovery of the RN(-) mutation. Furthermore, theseresults illustrate that additional alleles of genes involved in major mutationsmay play a significant role in quantitative trait variation.