EGAGenomicsIain McNeishhttps://ega-archive.org/dacs/EGAC00001000388EGAEGA DAC EGAC00001000388EGAi.mcneish@imperial.ac.uk<h4>Background</h4>Next-generation sequencing (NGS) of tumour samples is a critical component of personalised cancer treatment, but it requires high-quality DNA samples. Routine neutral-buffered formalin (NBF) fixation has detrimental effects on nucleic acids, causing low yields, as well as fragmentation and DNA base changes, leading to significant artefacts.<h4>Patients and methods</h4>We have carried out a detailed comparison of DNA quality from matched samples isolated from high-grade serous ovarian cancers from 16 patients fixed in methanol and NBF. These experiments use tumour fragments and mock biopsies to simulate routine practice, ensuring that results are applicable to standard clinical biopsies.<h4>Results</h4>Using matched snap-frozen tissue as gold standard comparator, we show that methanol-based fixation has significant benefits over NBF, with greater DNA yield, longer fragment size and more accurate copy-number calling using shallow whole-genome sequencing (WGS). These data also provide a new approach to understand and quantify artefactual effects of fixation using non-negative matrix factorisation to analyse mutational spectra from targeted and WGS data.<h4>Conclusion</h4>We strongly recommend the adoption of methanol fixation for sample collection strategies in new clinical trials. This approach is immediately available, is logistically simple and can offer cheaper and more reliable mutation calling than traditional NBF fixation.Methanol-based fixation is superior to buffered formalin for next-generation sequencing of DNA from clinical cancer samples.Piskorz A M AM, Ennis D D, Macintyre G G, Goranova T E TE, Eldridge M M, Segui-Gracia N N, Valganon M M, Hoyle A A, Orange C C, Moore L L, Jimenez-Linan M M, Millan D D, McNeish I A IA, Brenton J D JDHigh-Throughput RNA, Formol, High Throughput DNA Sequencing, DNS, Malignant Neoplasm, Alcohol, Nucleotide Sequencing, (Deoxyribonucleotide)n, Massively-Parallel, DNAn+1, Neoplasms, Benign Neoplasm, High-Throughput DNA Sequencing, Methyl alcohol, Methyl, CH3OH, High-Throughput DNA, Double-Stranded, Tumor, Methylalkohol, Malignant, Illumina, Next-Generation Sequencing, Deoxyribonucleic acids, (Deoxyribonucleotide)n+m, High-Throughput RNA Sequencing, Ion Torrent Sequencing, MT, Benign, Deoxyribonucleic Acid, Deep, Methanal, DNA Sequencing, Massively-Parallel Sequencing, Next-Generation, Neoplasm, Wood Alcohol, FORMALIN, Sodium, MeOH, Ion Torrent, Massively Parallel Sequencing, ds-DNA, desoxyribose nucleic acid, Carbinol, wood alcohol, Next Generation Sequencing, Deep Sequencing, carbinol, thymus nucleic acid, primary cancer, Ion Proton Sequencing, Malignancy, Illumina Sequencing, Formaldehyd, Oxomethane, Double Stranded, Benign Neoplasms, Deoxyribonucleic acid, Pyrosequencing, Cancers, malignant tumor, Methylene oxide, Sequencing, High Throughput Sequencing, Neoplasias, Wood, Formalin, High Throughput RNA Sequencing, Oxomethylene, High-Throughput Sequencing, wood naphtha, malignant neoplasm, spirit of wood, wood spirit, ds DNA, Desoxyribonukleinsaeure, High-Throughput, RNA Sequencing, High Throughput Nucleotide Sequencing, Double-Stranded DNA, Malignancies, (Deoxyribonucleotide)m, Sodium Methoxide, DNA, deoxyribonucleic acids, DNAn, Ion Proton, Methyl Alcohol, Neoplasia, Methoxide, High-Throughput Nucleotide, Cancer, Tumors, Malignant Neoplasms.fbwd4, l(2)04454, DmelCG1772, dac, CIB1, shsf3, shfm3, Decapo., E(Sev-CycE)2B, p21[dacapo], Fbw4, FBW4, dactylin, E130112M23Rik, Dach, CDKN2B, cdi4, SHFM3, Dac, DAC, p27[Dap], dactylyn, dacapo/cyclin-dependent kinase interactor 4, AI182278, p21, CG1772, FBWD4, Dap, Cdi4, CDI4, CES5A1, p27, P15, fbw4, SHSF3Formol, dalpha-SNAP, Nucleotide Sequencing, Massively-Parallel, SEC9, Tumor, bA416N4.2, Techniques, High-Throughput RNA Sequencing, Adoptions, sampling, Method, Methanal, Mutations., dJ1068F16.2, alpha-SNAP, Deep Sequencing, wood alcohol, treatment, thymus nucleic acid, me75, Biopsies, Genomes, Illumina Sequencing, bases, Formaldehyd, Tissue, Oxomethane, Basen, procedures, Artefacts, D17Mit170, T1, SNAP-25, High Throughput RNA Sequencing, acidos nucleicos, Methodological Studies, malignant neoplasm, disease management, Therapies, Malignancies, Double-Stranded DNA, Ovarian Cancers, deoxyribonucleic acids, Sodium Methoxide, DNAn, SIMPLE, 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High Throughput Nucleotide Sequencing, Treatment, nucleobases, Ion Proton, Neoplasia, High-Throughput Nucleotide, methodology, Methoxide, snapfalseDAC of the BRITROC projectData Access Committee EGAC00001000388EGAC00001000388960626681675EGAS00001007292EGAS00001001433EGAS00001002557EGAD00001004174EGAD00001001938EGAD00001011049EGAD00001011058EGAD00001004189EGAD00001004173EGAD00001004172EGAD00001001937