Direct detection of hemophilia B f9 gene mutation using multiplex PCR and conformation sensitive gel electrophoresis

Ki Young Yoo, Hee Jin Kim, Kwang Chul Lee

    Research output: Contribution to journalArticlepeer-review


    Purpose: The F9 gene is known to be the causative gene for hemophilia B, but unfortunately the detection rate for restriction fragment length polymorphism-based linkage analysis is only 55.6%. Direct DNA sequencing can detect 98% of mutations, but this alternative procedure is very costly. Here, we conducted multiplex polymerase chain reactions (PCRs) and conformation sensitive gel electrophoresis (CSGE) to perform a screened DNA sequencing for the F9 gene, and we compared the results with direct sequencing in terms of accuracy, cost, simplicity, and time consumption. Methods: A total of 27 unrelated hemophilia B patients were enrolled. Direct DNA sequencing was performed for 27 patients by a separate institute, and multiplex PCR-CSGE screened sequencing was done in our laboratory. Results of the direct DNA sequencing were used as a reference, to which the results of the multiplex PCR-CSGE screened sequencing were compared. For the patients whose mutation was not detected by the 2 methods, multiplex ligation-dependent probe amplification (MLPA) was conducted. Results: With direct sequencing, the mutations could be identified from 26 patients (96.3%), whereas for multiplex PCRCSGE screened sequencing, the mutations could be detected in 23 (85.2%). One patient's mutation was identified by MLPA. A total of 21 different mutations were found among the 27 patients. Conclusion: Multiplex PCR-CSGE screened DNA sequencing detected 88.9% of mutations and reduced costs by 55.7% compared with direct DNA sequencing. However, it was more labor-intensive and time-consuming.

    Original languageEnglish
    Pages (from-to)397-407
    Number of pages11
    JournalKorean Journal of Pediatrics
    Issue number3
    Publication statusPublished - 2010


    • DNA mutational analysis
    • Electrophoresis
    • Hemophilia B
    • Polymerase chain reaction

    ASJC Scopus subject areas

    • Pediatrics, Perinatology, and Child Health
    • Pediatrics


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