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To determine the origin of sickle cell mutation in different ethnic groups living in southern Iran, we studied the haplotype background of the βS and βA genes in subjects from the provinces of Fars, Khuzestan, Bushehr, Hormozgan, and Kerman and from the islands of Khark and Qeshm. β-globin gene cluster haplotypes were determined using the PCR-RFLP technique. Detection of -α3.7 deletion and β-thalassemia mutations were defined by PCR and reverse dot blot techniques, respectively. The framework of the β-globin gene was determined using denaturing gradient gel electrophoresis. We found that the βS mutation in southern Iran is associated with multiple mutational events. Most of the patients were from two ethnic groups: Farsi speakers (presumably Persian in origin) from Fars province and patients of Arab origin from Khuzestan province. In both ethnic groups the Arab-Indian haplotype was the most prevalent. The frequencies of the Arab-Indian and African haplotypes in sickle cell anemia patients from the provinces of Fars and Khuzestan were similar. Among βA chromosomes the Bantu A2 haplotype was the most prevalent. The decrease in α-globin production in SS patients and AS individuals appeared to be related to the reduction in mean cell volume and mean cell hemoglobin. The Arab-Indian haplotype gene flow into this region of Iran can be traced to the Sassanian Empire. It is likely that the influx of βS genes linked to the Benin and Bantu haplotypes, of African origin, must have occurred during the Arab slave trade.