MYH₃ is a major contractile protein which converts chemical energy into mechanical energy through the ATP hydrolysis. MYH₃ is mainly expressed in the skeletal muscle in different stages especially embryonic period, and it has a role in the development of skeletal muscle and heart. In this study, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was applied to analyze the genetic variations of the MYH₃ gene and verify the effect on growth and carcass traits in a total of 365 Qinchuan cattles. The PCR product was digested with some restriction enzyme and demonstrated the polymorphism in the population, the single nucleotide polymorphisms (SNPs) at nucleotides g. +1215T>C, g. +3377C>T, and g. +28625C>T were in linkage disequilibrium with each other. The result of haplotype analysis showed that nineteen different haplotypes were identified among the five SNPs. The statistical analyses indicated that the five SNPs were significant association with growth and carcass traits (P < 0.05, N = 365); whereas the five SNPs were no significant association between 18 combined genotypes of MYH₃ gene and growth and carcass traits. Taken together, our results provide the evidence that polymorphisms in MYH₃ are associated with growth and carcass traits in Qinchuan cattle, and may be used as a possible candidate for marker-assisted selection and management in beef cattle breeding program.