title: Pitfalls in molecular diagnosis in a family with severe factor VII (FVII) deficiency--misdiagnosis by direct sequence analysis using a PCR product.
abstract: Molecular diagnostic tests are becoming a routine analysis in many laboratories. These modern analyses are widely used in clinical medicine, forensic, genetic and prenatal diagnosis and also in preimplantation genetic diagnosis. The accuracy of analysis is highly dependent on the success achieved in minimising genotyping errors. The pitfalls in molecular diagnostic tests can be due to a simple technique such as the polymerase chain reaction (PCR) used universally. This technique is routinely used for its apparent accuracy, but it is also a well-known source of errors. We report an error introduced during PCR reaction that leads to a wrong sequence result and consequently to a 'false' molecular result in a next prenatal diagnosis in a family with severe factor VII (FVII) deficiency. This error was verified using an unsuitable primer design in a rich repetitive sequence of the FVII gene that leads to a false annealing and then to a wrong molecular diagnosis. It is essential to link closely molecular data with clinical and phenotype analysis in order to avoid false-negative or false-positive results, which is of great importance to diagnosis and molecular prevention.
authors.affiliation: A Bianchi Bonomi Hemophilia and Thrombosis Centre, Fondazione Luigi Villa, IRCCS Maggiore Hospital Milan and University of Milan, Milan, Italy.
authors.forename: Isabella
authors.initials: I
authors.lastname: Garagiola
authors.forename: Roberta
authors.initials: R
authors.lastname: Palla
authors.forename: Flora
authors.initials: F
authors.lastname: Peyvandi
mesh_terms: D000328:Adult; D002648:Child; D017931:DNA Primers; D003937:Diagnosis, Differential; D005167:Factor VII / Q000235:genetics*; D005168:Factor VII Deficiency / Q000175:diagnosis* / Q000235:genetics*; D005189:False Positive Reactions; D005260:Female; D005817:Genetic Counseling; D006801:Humans; D008297:Male; D009154:Mutation / Q000235:genetics; D010375:Pedigree; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D011296:Prenatal Diagnosis
chemical_list: D017931:DNA Primers; D005167:Factor VII