Rapid detection of beta-thalassemia by LDR-ULP combined with real-time PCR
10.3760/cma.j.issn.1009-9158.2016.10.009
- VernacularTitle:LDR-ULP 结合荧光定量 PCR 快速检测β-地中海贫血的应用研究
- Author:
Huan XU
;
Cheng YANG
;
Fake LI
;
Jie LUO
;
Wenbin JIANG
;
Fengling ZHANG
;
Chao WANG
;
Baosong YAN
;
Kai CHANG
;
Ming CHEN
- Publication Type:Journal Article
- Keywords:
beta-Thalassemia;
Real-time polymerase chain reaction;
Ligase detection reaction;
Uniform ligation probe
- From:
Chinese Journal of Laboratory Medicine
2016;39(10):766-770
- CountryChina
- Language:Chinese
-
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 ofβ-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.
