The application of high-resolution melting curve analysis in the detection of F9 gene mutations in haemophilia B patients
10.3760/cma.j.issn.1009-9158.2012.07.009
- VernacularTitle:高分辨率熔解曲线分析在血友病B F9基因突变检测中的应用
- Author:
Tingting YU
;
Jing DAI
;
Qihua FU
;
Xuefeng WANG
- Publication Type:Journal Article
- Keywords:
Hemophilia B;
Factor Ⅸ;
Mutation;
Polymerase chain reaction
- From:
Chinese Journal of Laboratory Medicine
2012;35(7):620-624
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo establish an effective method for F9 gene mutation detection by using high resolution melting ( HRM ) curve analysis.Methods Peripheral blood samples of 55 hemophilia B (HB) patients were collected from Shanghai Ruijin Hospital during January 2005 to June 2010.Genomic DNA was extracted from the peripheral blood.PCR amplification combined with sequencing was used to identify the F9 gene mutations in 40 patients.HRM assay was established on the 21 DNA samples with known mutations in exonl to exon7 of F9 gene.Mutation scanning of exonl to exon7 by HRM combined with direct sequencing of exon8 was used in the molecular diagnosis of 15 HB patients with unknown F9 gene mutations.ResultsF9 gene mutation was detected in each of the 40 HB patients by direct sequencing.By HRM,the different melting curve patterns were identified in 19 out of 21 cases.The detection rate was about 90％.Through mutation scanning of exonl to exon7 by HRM combined with direct sequencing of exon8,F9 gene mutations were detected in all the 15 HB patients with unknown F9 gene mutations.Thirty-four F9 gene mutations had been identified in the 55 HB patients.ConclusionsA new strategy of HB genetic diagnosis,scanning mutations of exonl to exon7 combined with DNA sequencing of exon8 of F9 gene,is established in this study.The new strategy is efficient and reliable.
