Introduction and Aim: Beta-thalassemia is a serious inherited genetic disorder and an increasing health burden globally. Beta -thalassemia is caused by genetic globin abnormalities within the hemoglobin beta (HBB) gene. This study aimed to characterize the HBB gene mutations in beta -thalassemia among southern Iraqi patients. Materials and Methods: The study included 30 beta -thalassemia patients referred to the Thi-Qar Center for Genetic Diseases, Iraq and 15 control samples from a random group of apparently healthy individuals. Genomic DNA was isolated from blood sample collected from each individual. The DNA was amplified for specific regions of the HBB gene and the amplified products sequenced. The sequences generated were analysed for mutations using sequence analysis tools. Results: Molecular analysis revealed several mutations in the HBB gene including translocation, deletion and substitution mutations in the population tested positive for the beta -thalassemia trait. Conclusion:  Thalassemia major is a serious concern in southern Iraq and therefore this study emphasizes a need for complete mutation profiling of the beta -globin gene as a strategy for screening of carriers within the population. Such examinations could be useful in pre-marital genetic counseling and for undertaking prevention and treatment measures.