Identification of point mutations and large intragenic deletions in Fanconi anemia using next‐generation sequencing technology. Issue 6 (2nd July 2015)

• Record Type: Journal Article
• Authors: Nicchia, Elena; Greco, Chiara; De Rocco, Daniela; Pecile, Vanna; D'Eustacchio, Angela; Cappelli, Enrico; Corti, Paola; Marra, Nicoletta; Ramenghi, Ugo; Pillon, Marta; Farruggia, Piero; Dufour, Carlo; Pallavicini, Alberto; Torelli, Lucio; Savoia, Anna
• Is Part Of: Molecular genetics & genomic medicine. Volume 3:Issue 6(2015:Nov.)
• Page Start: 500
• Page End: 512
• Publication Date: 2015-07-02
• Subjects: Fanconi anemia -- next‐generation sequencing -- ion PGM system -- point mutations -- copy number variations -- diagnosis; Medical genetics -- Periodicals; Genomics -- Periodicals; 616.042
• DOI: 10.1002/mgg3.160
• Languages: English
• ISSNs: 2324-9269

Abstract

Abstract : Fanconi Anemia (FA) is a rare bone marrow failure disorder characterized by clinical and genetic heterogeneity. Since the FA molecular diagnosis process is complex and time‐consuming, we applied the next generation sequencing Ion PGM system for detection of point mutations and copy number variations, which resulted to be suitable for mutational screening in FA. Abstract: Fanconi anemia (FA) is a rare bone marrow failure disorder characterized by clinical and genetic heterogeneity with at least 17 genes involved, which make molecular diagnosis complex and time‐consuming. Since next‐generation sequencing technologies could greatly improve the genetic testing in FA, we sequenced DNA samples with known and unknown mutant alleles using the Ion PGM ™ system (IPGM). The molecular target of 74.2 kb in size covered 96% of the FA‐coding exons and their flanking regions. Quality control testing revealed high coverage. Comparing the IPGM and Sanger sequencing output of FANCA, FANCC, and FANCG we found no false‐positive and a few false‐negative variants, which led to high sensitivity (95.58%) and specificity (100%) at least for these two most frequently mutated genes. The analysis also identified novel mutant alleles, including those in rare complementation groups FANCF and FANCL . Moreover, quantitative evaluation allowed us to characterize large intragenic deletions of FANCA and FANCD2, suggesting that IPGM is suitable for identification of not only point mutations but also copy number variations.