Rapid hemophilia A molecular diagnosis by a simple DNA sequencing procedure: identification of 14 novel mutations.

摘要: We here describe a simple, efficient DNA sequencing procedure for hemophilia A molecular diagnosis. In severe patients we first test the presence of factor VIII gene intron 22 inversion using recently described single-tube PCR method. moderate, mild, or inversion-negative systematically sequence promoter, all exons and splice junctions gene. Specially designed primers allow amplification 23 products under same salt conditions thermocycling parameters. The whole procedure, from blood extraction to mutation identification, can be readily done within 42 h when regular instruments in just 14 high-throughput sequencer. Thus, this is versatile cost-effective strategy with little hands-on time requirements. Since its implementation have identified mutations 45/46 patients, which are novel. Once genetic defect has been identified, accurate counseling then easily performed.