OBJECTIVETo establish a feasible protocol to provide genetic diagnosis and prenatal diagnosis in Chinese hemophilia patients and their relatives by direct exon sequencing.

METHODSIn our study, genetic diagnosis was performed on 5 unrelated families with informed consent, which included 3 pregnant women who asked for prenatal diagnosis. Umbilical cord blood was obtained from 2 fetuses and amniotic fluid from another fetus. After extraction of the genomic DNA, all of the exons, exon-intron boundaries and their flanks of FⅨ gene were amplified by polymerase chain reaction (PCR). PCR products were detected through direct-sequencing.

RESULTSSequence analysis indicated that the patients and carriers from 5 families have the pathogenic mutations,c.1022G>A (p.R341Q), c.484 C>T (p. R162X), c.1135C>T (p.R379X), c.799C>T (p.H267Y), c.1232G＞T (p.S411I), respectively. Except c.484 C>T (p. R162X), 4 of the 5 mutations were reported firstly in Chinese population. During prenatal diagnosis, one of the fetuses was found to be affected with c.484C>T; p.R162X. The remaining two fetuses were diagnosed as normal, the results of which were later verified by post-birth diagnosis, and factor FⅨ activities in plasma was 52.7% and 76.2%, respectively.

CONCLUSIONIn the quest of strict quality control, exon sequence on FⅨ gene was a rapid and accurate method for genetic diagnosis and prenatal diagnosis of hemophilia B.