title: Confirmation of the value of a modified long-distance polymerase chain reaction in the detection of inversion intron 22 in severe hemophilia a: a technical note.
abstract: Hemophilia A is one of the most common bleeding disorders in man. Approximately half the families with a severe disease have an inversion of the factor VIII gene. The inversion may be detected with a long polymerase chain reaction. This is a simple and reproducible method that may yield satisfactory results in approximately 24 hours. The long-distance polymerase chain reaction amplify three very large amplicons with a very GC-rich region of several kilobases, and is detected with a Expand Long Template DNA polymerase (Roche, Mannheim, Germany). Recently this polymerase has been changed with a new chemical composition. The object of the present method is to standardize the technique using the new Expand Long polymerase. For the new protocol, the cycling conditions, the concentration of nucleotide primers, and the buffer are changed. The need for a rapid response is determined in the case of hemophilia A patients, not only by the desire to reach a proper classification, but also by the urgency to inform the carrier status of the mother or of a female relative.
authors.affiliation: Department of Medical and Surgical Sciences, 2nd Chair of Internal Medicine, University of Padua Medical School, Padua, Italy. annamaria.lombardi@unipd.it
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Lombardi
authors.forename: Laura
authors.initials: L
authors.lastname: Cabrio
authors.forename: Ezio
authors.initials: E
authors.lastname: Zanon
authors.forename: Donatella
authors.initials: D
authors.lastname: Pellati
authors.forename: Silvia
authors.initials: S
authors.lastname: Vettore
authors.forename: Antonio
authors.initials: A
authors.lastname: Girolami
mesh_terms: D006467:Hemophilia A / Q000235:genetics* / Q000473:pathology*; D006801:Humans; D007438:Introns / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*