Detection by denaturing gradient gel electrophoresis of an Arg1689Cys mutation in a Chinese patient with mild hemophilia A
Ruan, C.; Gu, J.; Lavergne, J.M.; Meyer, D.
Chinese Medical Journal 110(2): 96-99
1997
ISSN/ISBN: 0366-6999 PMID: 9594277 Document Number: 483453
Objective: To detect gene defects of factor VIII (FVIII) in Chinese hemophilia A patients. Methods: 3' end of exon 14 of F VIII gene from a mild hemophilia A patient of Chinese origin was amplified by polymerase chain reaction (PCR) and identified mutations by denaturing gradient gel electrophoresis (DGGE) combining with direct sequencing. Results: An upward shift band was detected by DGGE in W381. Direct sequencing demonstrated a C to T transition resulting in substitution of Arg1689Cys within a thrombin activation site of mature FVIII protein, which created a unique a thrombin activation site of mature FVIII protein, which created a unique PstI site in amplified fragment of FVIII. Conclusions: The association of PCR and DGGE can detect a single base substitution; the Arg1689Cys mutation that inhibited activation of FVIII by thrombin is a molecular defect associated with hemophilia A in W381.