EGAGenomicsIllumina HiSeq 2000Otherhttps://ega-archive.org/studies/EGAS00001000642EGAEGA study EGAS00001000642QDNAseq datasetEGArestrictedDetection of DNA copy number aberrations by shallow whole-genome sequencing (WGS) faces many challenges, including lack of completion and errors in the human reference genome, repetitive sequences, polymorphisms, variable sample quality, and biases in the sequencing procedures. Formalin-fixed paraffin-embedded (FFPE) archival material, the analysis of which is important for studies of cancer, presents particular analytical difficulties due to degradation of the DNA and frequent lack of matched reference samples. We present a robust, cost-effective WGS method for DNA copy number analysis that addresses these challenges more successfully than currently available procedures. In practice, very useful profiles can be obtained with ∼0.1× genome coverage. We improve on previous methods by first implementing a combined correction for sequence mappability and GC content, and second, by applying this procedure to sequence data from the 1000 Genomes Project in order to develop a blacklist of problematic genome regions. A small subset of these blacklisted regions was previously identified by ENCODE, but the vast majority are novel unappreciated problematic regions. Our procedures are implemented in a pipeline called QDNAseq. We have analyzed over 1000 samples, most of which were obtained from the fixed tissue archives of more than 25 institutions. We demonstrate that for most samples our sequencing and analysis procedures yield genome profiles with noise levels near the statistical limit imposed by read counting. The described procedures also provide better correction of artifacts introduced by low DNA quality than prior approaches and better copy number data than high-resolution microarrays at a substantially lower cost.DNA copy number analysis of fresh and formalin-fixed specimens by shallow whole-genome sequencing with identification and exclusion of problematic regions in the genome assembly.Scheinin Ilari I, Sie Daoud D, Bengtsson Henrik H, van de Wiel Mark A MA, Olshen Adam B AB, van Thuijl Hinke F HF, van Essen Hendrik F HF, Eijk Paul P PP, Rustenburg François F, Meijer Gerrit A GA, Reijneveld Jaap C JC, Wesseling Pieter P, Pinkel Daniel D, Albertson Donna G DG, Ylstra Bauke Bnumber, count in organism, low depth, whole genome, shallow, presence, cardinality, Genomes.biochemical pathways, Archive, Formol, human being, DNS, Procedures, (Deoxyribonucleotide)n, determination, Ass-1, Neoplasms, Addresses, number, Benign Neoplasm, Cost Comparisons, Measure, Tumor, Malignant, composed of, presence, cellular catabolism, Cost-Minimization, Deoxyribonucleic acids, Human, Intervention or Procedure, method, Techniques, low depth, Deoxyribonucleic Acid, Homo sapiens, Cost Minimization Analysis, reduced, Costs and Cost Analyses., Method, Methanal, Parafilm, method used in an experiment, AA408052, Core Genome, Studies, fold, cellular degradation, Low, Analysis, tiny, Cost Comparison, Man, Technique, thymus nucleic acid, me75, Man (Taxonomy), Genomes, Malignancy, Analyses, breakdown of chemical, interventionDescription, catabolism, Cost Measures, Cost-Minimization Analyses, Affordability, Accessory Genome, Formaldehyd, Tissue, Oxomethane, hypoplasia, Double Stranded, Measures, Interventional, Comparison, composition, Deoxyribonucleic acid, procedures, financing, Artefacts, D17Mit170, SURGICAL AND MEDICAL PROCEDURES, T1, Neoplasias, Study, ASS, Cost Analysis, funding, high frequency, Formalin, Methodological Studies, malignant neoplasm, sample, Artifact, biotransformation, s, Double-Stranded DNA, Malignancies, (Deoxyribonucleotide)m, deoxyribonucleic acids, Comparisons, DNAn, Affordabilities, shallow, Cancer, Tumors, small, close to, WGS, cellular breakdown, fees, Intervention Strategies, Noise, Malignant Neoplasm, cou, degradation, Artefact, DNAn+1, Modern, Pollution, Double-Stranded, Procedure, compositionality, Tl3, Tl2, Pangenome, Faces, (Deoxyribonucleotide)n+m, near to, read, count in organism, Lr, MT, Benign, chemical analysis, Neoplasm, sequence, FORMALIN, secretion, simple tissue, techniques, ds-DNA, desoxyribose nucleic acid, fixed, finances, breakdown of molecule, Costs and Cost Analyses, Intervention, Costs, primary cancer, frequent, biodegradation, underdeveloped, Cost, content, Cost-Minimization Analysis, salaries, Benign Neoplasms, whole genome, Cancers, Pricing, Methodological, Methodological Study, malignant tumor, human, sample population, Methylene oxide, primary structure of sequence macromolecule, Noises, Malignant Neoplasms, plan specification, cost, breakdown of substance, Pan-genome, Oxomethylene, approaches, Modern Man, ds DNA, cardinality, structure, vicinity of, Desoxyribonukleinsaeure, Bra, assay, Noise Pollution, DNA, financial management, variable, Cost Measure, Neoplasia, Cost Analyses, methodologythymus nucleic acid, Formol, DNS, (Deoxyribonucleotide)n, determination, Genomes, DNAn+1, number, Formaldehyd, Oxomethane, Double Stranded, Deoxyribonucleic acid, whole genome, Double-Stranded, presence, Methylene oxide, Deoxyribonucleic acids, (Deoxyribonucleotide)n+m, count in organism, low depth, Formalin, Deoxyribonucleic Acid, Oxomethylene, Methanal, ds DNA, cardinality, chemical analysis, Desoxyribonukleinsaeure, FORMALIN, Double-Stranded DNA, (Deoxyribonucleotide)m, assay, DNA, deoxyribonucleic acids, DNAn, ds-DNA, desoxyribose nucleic acid, fixed, shallow, Genomes.biochemical pathways, Archive, Formol, human being, DNS, Procedures, (Deoxyribonucleotide)n, determination, Neoplasms, Addresses, number, Benign Neoplasm, Cost Comparisons, Measure, Tumor, Malignant, composed of, presence, cellular catabolism, Cost-Minimization, Deoxyribonucleic acids, Human, Intervention or Procedure, method, Techniques, low depth, Deoxyribonucleic Acid, Homo sapiens, Cost Minimization Analysis, reduced, Costs and Cost Analyses., Method, Methanal, Parafilm, method used in an experiment, Core Genome, Studies, cellular degradation, Low, Analysis, tiny, Cost Comparison, Man, Technique, thymus nucleic acid, me75, Man (Taxonomy), Genomes, Malignancy, Analyses, breakdown of chemical, interventionDescription, catabolism, Cost Measures, Cost-Minimization Analyses, Affordability, Accessory Genome, Formaldehyd, Tissue, Oxomethane, hypoplasia, Double Stranded, Measures, Interventional, Comparison, composition, Deoxyribonucleic acid, procedures, financing, Artefacts, D17Mit170, SURGICAL AND MEDICAL PROCEDURES, T1, Neoplasias, Study, Cost Analysis, funding, high frequency, Formalin, Methodological Studies, malignant neoplasm, sample, Artifact, biotransformation, s, Double-Stranded DNA, Malignancies, (Deoxyribonucleotide)m, deoxyribonucleic acids, Comparisons, DNAn, Affordabilities, shallow, Cancer, Tumors, small, close to, WGS, cellular breakdown, fees, Intervention Strategies, Noise, Malignant Neoplasm, cou, degradation, Artefact, DNAn+1, Modern, Pollution, Double-Stranded, Procedure, compositionality, Tl3, Tl2, Pangenome, Faces, (Deoxyribonucleotide)n+m, near to, read, count in organism, Lr, MT, Benign, chemical analysis, Neoplasm, sequence, FORMALIN, secretion, simple tissue, techniques, ds-DNA, desoxyribose nucleic acid, fixed, finances, breakdown of molecule, Costs and Cost Analyses, Intervention, Costs, primary cancer, frequent, biodegradation, underdeveloped, Cost, content, Cost-Minimization Analysis, salaries, Benign Neoplasms, whole genome, Cancers, Pricing, Methodological, Methodological Study, malignant tumor, human, sample population, Methylene oxide, primary structure of sequence macromolecule, Noises, Malignant Neoplasms, plan specification, cost, breakdown of substance, Pan-genome, Oxomethylene, approaches, Modern Man, ds DNA, cardinality, structure, vicinity of, Desoxyribonukleinsaeure, Bra, assay, Noise Pollution, DNA, financial management, variable, Cost Measure, Neoplasia, Cost Analyses, methodologyfalseQuantification of chromosomal copy number aberrations by shallow whole-genome sequencingDetection of DNA copy number aberrations by shallow whole-genome sequencing (WGS) faces many challenges including lack of completion and errors in the human reference genome, repetitive sequences, polymorphisms, variable sample quality, and biases in the sequencing procedures. Formalin-fixed paraffin-embedded (FFPE) archival material, the analysis of which is important for studies of cancer, presents particular analytical difficulties due to degradation of the DNA and frequent lack of matched reference samples. We present a robust, cost-effective WGS method for DNA copy number analysis that addresses these challenges more successfully than currently available procedures. In practice very useful profiles can be obtained with 0.1 fold genome coverage. We improve on previous methods by; first, implementing a combined correction for sequence mappability and GC content, and second, applying this procedure to sequence data from the 1000 Genomes Project in order to develop a blacklist of problematic genome regions. A small subset of these blacklisted regions were previously identified by ENCODE, but the vast majority are novel unappreciated problematic regions. Our procedures are implemented in a pipeline called QDNAseq. We have analyzed over 1,000 samples, most of which were obtained from the fixed tissue archives of over 25 institutions. We demonstrate that for most samples our sequencing and analysis procedures yield genome profiles with noise levels near the statistical limit imposed by read counting. The described procedures also provide better correction of artifacts introduced by low DNA quality than prior approaches, and better copy number data than high-resolution microarrays at substantially lower cost.2017-07-26 15:39:25EGAS00001000642960625236618EGAD00001000780EGAC00001000181