title: Next-generation carrier screening.
abstract: OBJECTIVE
  Carrier screening for recessive Mendelian disorders traditionally employs focused genotyping to interrogate limited sets of mutations most prevalent in specific ethnic groups. We sought to develop a next-generation DNA sequencing-based workflow to enable analysis of a more comprehensive set of disease-causing mutations.
  
  
  METHODS
  We utilized molecular inversion probes to capture the protein-coding regions of 15 genes from genomic DNA isolated from whole blood and sequenced those regions using the Illumina HiSeq 2000 (Illumina, San Diego, CA). To assess the quality of the resulting data, we measured both the fraction of the targeted region yielding high-quality genotype calls, and the sensitivity and specificity of those calls by comparison with conventional Sanger sequencing across hundreds of samples. Finally, to improve the overall accuracy for detecting insertions and deletions, we introduce a novel assembly-based approach that substantially increases sensitivity without reducing specificity.
  
  
  RESULTS
  We generated high-quality sequence for at least 99.8% of targeted base pairs in samples derived from blood and achieved high concordance with Sanger sequencing (sensitivity >99.9%, specificity >99.999%). Our novel algorithm is capable of detecting insertions and deletions inaccessible by current methods.
  
  
  CONCLUSIONS
  Our next-generation DNA sequencing-based approach yields the accuracy and completeness necessary for a carrier screening test.
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Umbarger
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Caleb J
authors.initials: CJ
authors.lastname: Kennedy
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Patrick
authors.initials: P
authors.lastname: Saunders
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Breton
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Niru
authors.initials: N
authors.lastname: Chennagiri
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: John
authors.initials: J
authors.lastname: Emhoff
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Valerie
authors.initials: V
authors.lastname: Greger
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Hallam
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: David
authors.initials: D
authors.lastname: Maganzini
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Cynthia
authors.initials: C
authors.lastname: Micale
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Marcia M
authors.initials: MM
authors.lastname: Nizzari
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Charles F
authors.initials: CF
authors.lastname: Towne
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA.
authors.forename: George M
authors.initials: GM
authors.lastname: Church
authors.affiliation: Good Start Genetics, Cambridge, Massachusetts, USA.
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Porreca
mesh_terms: D000818:Animals; D004252:DNA Mutational Analysis / Q000379:methods*; D005820:Genetic Testing / Q000191:economics / Q000379:methods*; D014644:Genetic Variation; D015894:Genome, Human; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D054643:INDEL Mutation; D020641:Polymorphism, Single Nucleotide; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000191:economics / Q000379:methods