Abstract
Objective To establish a new method for rapid detection of β-thalassemia by investigating six clinical common mutation types. Methods Fifty cases of clinical wild-type samples and 42 cases of β-thalassemia samples were collected, and β-globin gene was amplified by PCR. Uniform ligation probe (ULP)specific probes were designed for hybridization reaction to increase the reaction specificity and real-time PCR was performed to increase the sensitivity. After that, PCR products were verified by agarose electrophoresis. After examining the specificity and sensitivity, Kappa test between LDR-ULP method and reverse dot blot(RDB) method was conducted. Results Hybridization efficiency was improved 2.53 times by LDR-ULP hybridization. Each mutant type showed a significant amplification curve, whereas the wild-type had no significant curve within 40 cycles.The limit of determination of this method was 5 pg. The results of 92 cases of peripheral blood samples detected by the method of LDR-ULP and RDB were completely consistent. Conclusion In this study, a simple, inexpensive, rapid new method to detect β-thalassemia were established.(Chin J Lab Med, 2016, 39: 766-770) Key words: beta-Thalassemia; Real-time polymerase chain reaction; Ligase detection reaction; Uniform ligation probe
Published Version
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