Here we report the result of three cases referred to our lab that had a combination of β-thalassemia and hemoglobin D (Hb D) traits. These individuals had no symptoms of profound anemia and hematological indices were similar to that of a β-thalassemia heterozygote. In all three cases, the Hb D level was elevated and no HbA was detected electrophoretically. The electrophoresis pattern suggested that all cases were homozygotes for Hb D. PCR followed by digestion with EcoRI and sequencing of the β-globin gene confirmed the presence of Cd 121 GAA>CAA in the heterozygous form with another β-globin mutation. In all cases, the mutations in the β-globin gene were detected by ARMS PCR technique and they were either IVSII-I or IVSI-5. Hematological studies of the family members showed that thalassemia which caused the mutations and Hb D were in the trans position.