Comparison of real-time polymerase chain reaction SYBR Green 1 with high resolution melting analysis and TaqMan MGB probes for detection of alpha-thalassemia-1 South-East Asian type on dried blood spots

摘要

alpha-Thalassemia-l South-East Asian (SEA) type is the main alpha-thalassemia abnormality in South-East Asia (1, 2). The gap-polymerase chain reaction (PCR) analysis currently used to diagnose a-thalassemia-1 SEA type requires post-PCR processing steps and hazardous chemicals (3-5). In an effort to develop a more straightforward diagnostic test, quantitative real-time PCR with the specific probe has been used for detection of alpha-thalasse-mia-1 SEA type (6, 7). However, using the specific probe is relatively expensive. As a cost-effective method, realtime PCR with SYBR Green 1 followed by melting curve analysis (8, 9) and real-time gap-PCR with SYBR Green 1 followed by high resolution melting (HRM) analysis (10) have been developed and used to enhance the speed of detection of a-thalassemia-1 SEA type. In low-resource settings, real-time PCR diagnostic protocols are considered too costly and complex. Dried blood spot (DBS) has been applied for many molecular investigations as it carries less of a biohazard risk than liquid samples and is easier to ship (11-15). Therefore, reliable detection of a-thalassemia-1 SEA type on DBS by realtime PCR has a potential to control a serious genetically transmitted disease in low-resource settings.