Rapid genotyping of known mutations and polymorphisms in β-globin gene based on the DHPLC profile patterns of homoduplexes and heteroduplexes

Background β-thalassemia represents a great heterogeneity as over 200 mutations have been identified for the β-globin gene responsible for this disease. A rapid genotyping test with high accuracy, selectivity, and reproducibility suitable for the determination of known mutations is needed for prenatal screening and post-natal diagnosis of this disease in clinical setting. Design and methods We have performed the validation of a DHPLC assay for direct genotyping of known causative mutations in β-globin gene using the chromatographic pattern-based strategy under partially-denaturing conditions. Results DHPLC assay was established based on the analysis of 795 DNA samples from a group of various genotypes for the 20 mutations and 8 polymorphisms in β-globin gene then validated on 319 tests in a blind study. The results obtained with this assay were in concordance with the results obtained by DNA sequence analysis. Conclusion This simple method can meet the requirements of direct genotyping of known β-thalassemia mutations and/or polymorphisms in the clinical setting for Chinese and in general as a model for other populations.