title: Genotyping of fanconi anemia patients by whole exome sequencing: advantages and challenges.
abstract: Fanconi anemia (FA) is a rare genomic instability syndrome. Disease-causing are biallelic mutations in any one of at least 15 genes encoding members of the FA/BRCA pathway of DNA-interstrand crosslink repair. Patients are diagnosed based upon phenotypical manifestations and the diagnosis of FA is confirmed by the hypersensitivity of cells to DNA interstrand crosslinking agents. Customary molecular diagnostics has become increasingly cumbersome, time-consuming and expensive the more FA genes have been identified. We performed Whole Exome Sequencing (WES) in four FA patients in order to investigate the potential of this method for FA genotyping. In search of an optimal WES methodology we explored different enrichment and sequencing techniques. In each case we were able to identify the pathogenic mutations so that WES provided both, complementation group assignment and mutation detection in a single approach. The mutations included homozygous and heterozygous single base pair substitutions and a two-base-pair duplication in FANCJ, -D1, or -D2. Different WES strategies had no critical influence on the individual outcome. However, database errors and in particular pseudogenes impose obstacles that may prevent correct data perception and interpretation, and thus cause pitfalls. With these difficulties in mind, our results show that WES is a valuable tool for the molecular diagnosis of FA and a sufficiently safe technique, capable of engaging increasingly in competition with classical genetic approaches.
authors.affiliation: Department of Human Genetics, University of Wuerzburg, Wuerzburg, Germany. kerstin.knies@uni-wuerzburg.de
authors.forename: Kerstin
authors.initials: K
authors.lastname: Knies
authors.forename: Beatrice
authors.initials: B
authors.lastname: Schuster
authors.forename: Najim
authors.initials: N
authors.lastname: Ameziane
authors.forename: Martin
authors.initials: M
authors.lastname: Rooimans
authors.forename: Thomas
authors.initials: T
authors.lastname: Bettecken
authors.forename: Johan
authors.initials: J
authors.lastname: de Winter
authors.forename: Detlev
authors.initials: D
authors.lastname: Schindler
mesh_terms: D001483:Base Sequence; D002453:Cell Cycle; D059472:Exome; D005199:Fanconi Anemia / Q000175:diagnosis / Q000235:genetics*; D052236:Fanconi Anemia Complementation Group D2 Protein / Q000235:genetics; D005260:Female; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008297:Male; D009154:Mutation; D010375:Pedigree; D015203:Reproducibility of Results
chemical_list: D052236:Fanconi Anemia Complementation Group D2 Protein