Abstract: Toll-like receptors 4 performed two functions: recognition of pathogen ligands and signaling to initiate innate and adaptive immune responses. In this experiment, a 243bp segment of the exon3 of TLR4 gene of Chinese Holstein, Sanhe cattle and Chinese Simmental was amplified by Created Restriction Site PCR. The genetic polymorphisms of three populations were detected by digestion with restriction endomuclease HinfⅠ. After sequencing, a polymorphic site in amplified production was identified of having either a C and a T at position 27bp, which induced that Thr changed into Ile. Statistical results of χ2 test indicated that the polymorphism sites locus in Sanhe cattle did not fit Hardy-Weinberg equilibrium(P<0.05). Meanwhile the effect of polymorphism of TLR4 gene on somatic cell score was analyzed,The results showed that: the somatic cell score of individual with genotype AA were lowest significantly than that of other genotype（P<0.05）. In a word, the allele A might play an important role in mastitis resistance in bovine.