New method for the extraction of DNA from white blood cells for the detection of common genetic variants associated with thrombophilia.
In the last few years, FV Leiden, prothrombin 20210 and thermolabile methylene tetrahydrofolate reductase (MTHFR) 677 variants have been identified as possible risk factors associated with thrombophilia, and many PCR-based methods have been described for detecting these variations. However, the genomic DNA extraction is still a rate-limiting and time-consuming step in the PCR process. In an attempt to accelerate this procedure and make it suitable for routine laboratory, we report a single preparative technique for DNA extraction from peripheral blood samples using an anion-binding resin (GeneFizz). This method enables white blood cell lysis, DNA extraction and PCR amplification directly in the thermocycling tube on the thermocycler. The use of this new DNA extraction system coupled to a multiplex PCR allows rapid genetic screening of large cohorts of patients for thrombophilic risk factors.
Angelini
Antonio
A
Dipartimento di Medicina e Scienze dell'Invecchiamento, Sezione di Aterosclerosi e Trombosi, Facoltà di Medicina e Chirurgia, Università G. D'Annunzio, Chieti, Italy. aangelini@unich.it
Di Febbo
Concetta
C
Rullo
Agnese
A
Di Ilio
Carmine
C
Cuccurullo
Franco
F
Porreca
Ettore
E
D000837:Anion Exchange Resins; D004247:DNA / Q000302:isolation & purification*; D004252:DNA Mutational Analysis / Q000379:methods; D005165:Factor V / Q000235:genetics; D005820:Genetic Testing / Q000379:methods*; D014644:Genetic Variation; D006801:Humans; D007962:Leukocytes / Q000737:chemistry*; D008722:Methods; D042965:Methylenetetrahydrofolate Reductase (NADPH2) / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods; D019851:Thrombophilia / Q000235:genetics*
D000837:Anion Exchange Resins; C095381:factor V Leiden; D005165:Factor V; D004247:DNA; D042965:Methylenetetrahydrofolate Reductase (NADPH2)