The ABO system is the most important of all blood group systems in transfusion practice. The subgroup gives a weak reaction when treated with anti-A or anti-B. The most common subgroup found in Thai blood donors is subgroup A3, which is characterized by mixed-field agglutination when reacted with anti-A and anti-A,B, was caused by mutation in the ABO gene, especially in the exon 7. In the present study mutation in A3 were charactered in exon 7 of the ABO gene in 10 A3 phenotype Thai blood donors from the National Blood Centre, Thai Red Cross Society by PCR amplification and DNA sequencing. Mutations in exon 7 were identified in the A allele of six cases. In four cases, mutations were detected at positions 646T > A, 681G > A, 771C > T and 829G > A. One case showed a double mutations at positions 467C > Tand 745C > T and one case showed a mutation at position 467C > T. Four cases showed wild type exon 7 as A101 allele. These mutations were previously reported in BGMUT database and no novel mutation was identified These data suggest genetic heterogeneity in A3phenotype in Thai blood donors.