Hb E [β26(B8)Glu→Lys]/β-thalassemia (β-thal) is a worldwide inherited disorder. We determined the phenotype of 65 unrelated Hb E/β-thal subjects and 70 healthy individuals in the Guangxi Province of the Republic of China (ROC). Single nucleotide polymorphism (SNP) rs35959442 in HBS1L analysis was performed using the polymerase chain reaction (PCR)/restriction enzyme method. The data suggested that the frequency of the rs35959442 polymorphism was relatively high in patients with Hb E/β-thal in Guangxi Province, ROC, when associated with Hb F augmentation.