CCAAT/enhancer-binding protein alpha (C/EBPα) is an essential transcriptional factor regulating the differentiation of adipocytes. We report a novel single nucleotide polymorphism (C271A) of the C/EBPα gene in six indigenous Chinese cattle breeds using PCR-SSCP and DNA sequencing methods. Allele frequencies were investigated and evaluated by the χ(2) test in 817 individuals; all populations were found to be in Hardy-Weinberg equilibrium. Gene heterozygosity, effective allele numbers and polymorphism information content of the C/EBPα locus varied from 0.50 to 0.54, 1.84 to 1.99 and 0.35 to 0.37, respectively. We also evaluated a potential association of the C/EBPα SNP with ultrasound traits in 555 individuals; individuals of the AA genotype had greater ultrasound backfat thickness than did genotype CC (0.36 versus 0.34 cm, P < 0.01); genotypes AA and CA had higher ultrasound marbling scores than did genotype CC (3.53, 3.52 versus 3.37, P < 0.05). Analysis based on meat quality data in another 204 Qinchuan cattle showed that animals with genotype AA had bigger loin eye areas than did genotype CA (87.10 versus 79.08 cm(2), P < 0.05). These results indicate that the C271A SNP of the C/EBPα gene could be used as a molecular marker for selecting beef cattle with superior carcass traits.