A polymerase chain reaction (PCR) assay was performed in this study to amplify the major surface protein 5 (msp5) gene from the genomic DNA of Anaplasma marginale in Hyalomma asiaticum ticks by species-specific primers. Sequence analysis showed that the msp5 gene was 643 bases long and that the PCR products from the samples had an identical sequence (JX507127). Moreover, the BLAST showed that the sequence was identical to the msp5 sequences of A. marginale and most closely related to the A. marginale msp5 gene (AB704328) and the Liangdang strain of the A. marginale msp5 gene (EF546443) with similarity of 99 % (differing only by two bases). An epidemiological survey was performed in several dairy farms: a total of 68 ticks were collected from 49 cattle. As a result, 14 of the 49 (28.57 %) blood smears stained with Wright-Giemsa and 22 of the 68 (32.35 %) ticks examined by PCR assay exhibited A. marginale infection. The results of the PCR assay were mostly consistent with the results of the microscopic examination. A number of results were negative in blood smear but positive in PCR, which is important for the early diagnosis of anaplasmosis.