The C57BL/6J (B6) and DBA/2J (D2) strains and two derivative populations, BXD recombinant inbred strains (BXD RIs) and B6D2F₂ were used to explore the genetic basis for variation in muscle weight at 500 days of age. Paralleling the findings in 200-day old mice (Lionikas et al. 2003), the weight of the slow-twitch soleus, the mixed gastrocnemius and the fast-twitch tibialis anterior (TA) and extensor digitorum longus (EDL) muscles was 13-22% greater (p<0.001) in B6 than in D2. The distribution of BXD RI strain means indicated that the genetic influence on muscle weight (strain effect p<0.001 for all muscles) was of polygenic origin, and the effect of the genetic factors differed between males and females (strain by sex interaction p<0.01 for soleus, EDL and p<0.05 for TA, gastrocnemius). Linkage analyses in the B6D2F₂ population identified quantitative trait loci (QTL) affecting muscle weight on Chr 1, 2, 6 and 9. Pleiotropic influences were observed for the QTL on Chr 1 (soleus, TA), 2(TA, EDL, gastrocnemius), and 9 (soleus, TA, EDL) and were not related to muscle type (fast-/slow-twitch) or function (flexor/extensor). Effect of the QTL on Chr 9 on soleus muscle was male specific. The QTL on Chr 2 and 6 were previously observed at the age of 200 days, whereas the QTL on Chr 1 and 9 are novel muscle weight QTL. In summary, muscle weight in the B6/D2 lineage is affected by a polygenic system that has variable influences at different ages, between males and females, and across muscles in a manner that is independent of muscle type.