The paired box 7 (Pax7) gene encoding for the transcription factor can regulate the conversion of stem cells into myogenic cells and participate in the development and regeneration of skeletal muscle. The aims of this study were to detect variations of the Pax7 gene by DNA pool sequencing and aCRS-RFLP methods in 1441 cattle from five breeds and to investigate their associations with growth traits in Nanyang and Chinese red steppe cattle. Altogether, three novel single nucleotide polymorphisms (SNPs) were identified in the last intron of the Pax7 gene: NC_007300: ss1 (g. G103688A), ss2 (g. T103735C), and ss3 (g. A103764T). Genotypes and the referring haplotype frequencies showed a high similarity trend among five breeds, and the G, T, and A allele frequencies of the three loci were always superior when separate or in combination. Association analysis of the single SNPs and haplotype combinations revealed that the T allele of ss2 and ss3 loci and the haplotype H(2)H(2) (GG-TT-TT) showed significant effects on growth traits such as body height, body mass, and chest girth in cattle at early stages (6 and 12 months old) (P < 0.05). The results showed that Pax7 gene variations and their corresponding genotypes may be considered as molecular markers for economic traits in cattle breeding.