Objective To investigate the carriage of the thalassemia gene in sperm donors in Chongqing，China by analyzing the thalassemia screening and gene mutation types of sperm donors in Chongqing Human Sperm Bank（CHSB），and to provide a reference for thalassemia screening in sperm donors in Chongqing. Methods Blood samples were collected from 488 sperm donors in CHSB（donors with all tests qualified during the quarantine） from December 2021 to August 2024. A complete blood count analysis was performed to measure the mean corpuscular volume（MCV） and mean corpuscular hemoglobin（MCH）. Polymerase chain reaction（PCR）/agarose gel electrophoresis was used to detect deletional α-thalassemia mutations. PCR and DNA reverse dot blot hybridization were used to detect 3 common non-deletional α-thalassemia mutations and 17 common β-thalassemia mutations. The test results were then analyzed. Results Among the 488 sperm donors aged 25.64±4.95 years，15（3.07%，all Han nationality） tested positive for thalassemia. This carriage rate was lower than that of the thalassemia population in Chongqing（9.24%）. Among the 15 thalassemia carriers，13 had deletional α-gene mutations（-α3.7 genotype in 11 cases［73.33%］， -α4.2 genotype in 1 case［6.67%］，and -SEA genotype in 1 case ［6.67%］），and 2 had non-deletional α-gene mutations（both with the WSM heterozygous mutation）. There was only 1 case of abnormal MCV and MCH2 indicators among thalassemia carriers（screening positive rate was 6.67%）. Conclusion Thalassemia screening in sperm donors in CHSB reveals a low thalassemia carriage rate，reflecting geographic （predominantly Sichuan and Chongqing origins） and demographic （healthy men of childbearing potential） characteristics. Given the significant impact of thalassemia on the offspring conceived through sperm donation，it is recommended to conduct thalassemia gene screening among sperm donors in human sperm banks.