Computational protocol: A systematic approach to the reporting of medically relevant findings from whole genome sequencing

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Protocol publication

[…] Lossless BAM files are converted to FASTQ format to obtain sequence read data and reads are realigned to the NCBI reference sequence (GRCh37) using the Burrows-Wheeler Aligner 0.6.1-r104 []. Variant calls are made using the Genomic Analysis Tool Kit (GATK) version 2.3-9-gdcdccbb [] for all positions with ≥8X coverage. Variant annotation is derived from ALAMUT HT version 1.1.2, Variant Effect Predictor version 2.6 and the LMM’s GeneInsight laboratory database. Annotated variants are subsequently filtered to identify: (1) variants with a minor allele frequency (MAF) <5% in European American (EA) or African American (AA) chromosomes from the NHLBI Exome Sequencing Project (ESP; classified as disease causing (DM) or possible disease causing mutations (DM?) in the Human Gene Mutation Database [] or as Pathogenic or Likely pathogenic by the LMM; (2) nonsense, frameshift, and canonical splice-site (+/−1,2) variants with a MAF <1% in EA or AA chromosomes from the NHLBI ESP from a list of 4,631 disease-associated genes curated by expert review of many sources of gene-disease relationships (Online Mendelian Inheritance in Man (OMIM),ClinVar, etc.; and (3) pharmacogenomic variants for metformin (C11orf65 rs11212617), clopidogrel (CYP2C19 rs12248560, rs4244285, rs4986893, rs28399504, rs41291556, rs72552267, rs72558186, rs56337013), warfarin (CYP2C9 rs1057910, rs1799853, rs7900194, rs9332131, rs28371685, rs28371686 and VKORC1 rs9923231), simvastatin (SLCO1B1 rs4149056), and digoxin (ABCB1 rs1045642) metabolism. The disease-associated gene list is iteratively refined as gene-disease association information is curated. In addition, blood group antigens are predicted through a parallel pipeline as noted below. [...] Variant information, including classifications, interpretations, and associated references are stored in an internal laboratory knowledge base using the GeneInsight software suite []. Approved variant classifications with supporting evidence descriptions are submitted to the ClinVar database to support community knowledge sharing []. GRs are drafted and finalized using GeneInsight Lab and electronically delivered to physicians participating in the MedSeq Project via GeneInsight Clinic. Changes to variant classification in GeneInsight Lab resulting from new data are automatically communicated to participating physicians via an email notification that links them to a patient report update within the GeneInsight Clinic application for subsequent decision support []. […]

Pipeline specifications

Software tools BWA, GATK, VEP, GeneInsight Suite
Databases ClinVar OMIM HGMD
Applications Miscellaneous, WES analysis
Organisms Homo sapiens