Detection of the Inversions of Factor VIII Gene by Single-tube PCR.
- Author:
Jin Yeong HAN
1
;
Kyeong Hee KIM
;
Young Ho LEE
;
Jeong Nyeo LEE
;
Soon Yong LEE
;
Ook Hwan CHOI
;
In Joo KIM
;
Cheol Min KIM
Author Information
1. Department of Clinical Pathology, Dong-A University College of Medicine, Pusan, Korea.
- Publication Type:Original Article
- Keywords:
Hemophilia A;
Factor VIII;
Inversion;
Mutation;
Polymerase chain reaction
- MeSH:
Diagnosis;
DNA;
Factor VIII*;
Hemophilia A;
Hemorrhage;
Humans;
Incidence;
Male;
Polymerase Chain Reaction*
- From:Korean Journal of Clinical Pathology
2001;21(3):231-234
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
BACKGROUND: Hemophilia A is the most common X-linked bleeding disorder with an incidence of 1/5,000 males. Inversions within the factor VIII gene cause almost half of all cases of severe hemophilia A. However, DNA-based diagnosis has previously been carried out only by linkage analysis in Korean hemophilia A families. In this study, we aimed to establish direct inversion detection using a single-tube polymerase chain reaction (PCR) assay. METHODS: We have modified a single-tube PCR assay that combines overlapping PCR with long-distance PCR; performing PCR directly from genomic DNA with four primers P, Q, A, and B that differentiate the wild type, inversion, and the carrier detected the inversion. RESULTS: Segments PQ (12 kb) and AB (10 kb) were produced in hemizygous wild-type males. Males with hemophilia A due to the inversion showed segments PB+AQ (11 kb) along with the 10 kb segment from the nonrecombined extragenic homologue. In 20 (18.7%) patients, an inversion was found. The three segments were readily identifiable and all PCR amplifications achieved uniform reproducible results. CONCLUSIONS: The PCR was successful for the direct detection of factor VIII gene inversions. The method is simple, inexpensive, and more standardized; therefore, it may be the natural starting point for ascertaining mutations in families with severe hemophilia A.
