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Abrf 2017 hadfield j


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My talk for the International Genomics session at ABRF 2017. Describing the issues caused by the uncontrolled naming of NGS methods: some examples and some suggestions about how to fix this.

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Abrf 2017 hadfield j

  1. 1. ABRF 2017 Annual Meeting A FORUM FOR ADVANCING TODAY’S CORE TECHNOLOGIES TO ENABLE TOMORROW’S INNOVATIONS March 25-28, 2017 Town and Country Resort & Convention Center San Diego, California A profusion of confusion in genomic methods Dr James Hadfield CRUK & Cambridge University
  2. 2. NGS method naming Describe the problem Present some examples Suggestions on how to fix the problem Core-facility community efforts A genomics case report
  3. 3. NGS method naming – 10 years of NGS – 400 methods – No controlled vocabulary, no convention “What does RNA-Seq mean?”
  4. 4. Fragment) DNA) Quant) DNA) Size)select) Size) select) Ligate) adapters) End9) repair) Adenylate) Tagment) Quant) DNA) Hyb) oligos) Extend) ligate) 2nd)PCR) 1st)capture) 2nd)capture) Bead)normalise) &)pool) Strand) displace) Size)select) Circularise) &)exo) Bead) purify) Shear) 5’)ligaKon) Reverse) transcribe) 3’)ligaKon) Gel)purify) Fragment) RNA) 2nd)strand)) dUTP) UNG) QC)BioAnalyser) 1st)strand) PCR) QT)qPCR) Ribozero) deplete) Oligo)dT) enrich) Library) synthesis) Ampliy) &)Index) Template) preparaKon) QC)BioAnalyser) QT)qPCR)TruSeq'stranded'mRNA' TruSeq'sRNA' TruSeq'Ribozero' TREX' Nextera'XT' Nextera' Nextera'Rapid'Exome' Nextera'Mate2Pair' TruSeq'ChIP,seq' TSCA' TruSeq'PCR,free' TruSeq'Nano' Thruplex' NGS method naming Shendure & Lieberman: The expanding scope of DNA sequencing. Nat Biotech 2012
  5. 5. Same method different names – MNase-Seq, MAINE-Seq and Nuc-seq • Methods that are essentially the same are published with different names • Suggest using MNase-Seq for all
  6. 6. Same name different methods – Nuc-seq, Nuc-seq and sNuc-Seq • Methods with the same names are distinct e.g. Nuc-seq is a nucleosome positioning method, while Nuc-seq and sNuc-Seq are RNA and DNA nucleus sequencing methods. • Suggest using MNase-Seq, snS1Nuc-Seq, snRNA-Seq
  7. 7. Homophonic names – CAP-seq, CapSeq, CaptureSeq or CAPP-seq(not pictured) • Names that sound the same and lack clarity e.g. which name refers to each of the methods for CXXC affinity purification, 5’ anchored profiling of Pol II transcripts, targeted RNA sequencing or personalized profiling of cancer patients. • Suggest using CXXCaP-Seq, 5’Cap-Seq, RNAcapture-Seq
  8. 8. Is it really a new method – ChIPmentation • New methods are modifications of an earlier method e.g. ChIPmentation is a low-input transposase-based ChIP-Seq method. In cases like this appending an additional acronym may be preferable e.g. AHT-ChIP-seq, an automated high- throughput ChIP-Seq method. • Suggest using tnChIP-Seq
  9. 9. NGS method naming – some suggestions 1. Previously published names should used e.g. ChIP-Seq; or modified e.g. HiChIP-Seq or TnChIP-Seq 2. Case is important: most acronyms are formed from the method name e.g. RRBS, but the use of lowercase characters can help pronunciation e.g. ChIP-Seq 3. Prefixes and suffixes: can be used to add information about a method e.g. “sc” and “Bis” to denote single-cells or bisulphite conversion, or to distinguish method variants e.g. miCLIP-m5A, or miCLIP-m6A 4. Hyphens: can separate the method from the technology e.g. ChIP-Seq, ChIP-qPCR & ChIP-MS or denote significant protocol modifications e.g. PAR-CLIP-Seq
  10. 10. NGS method naming – some resources – Discuss the problem (in press) – Enseqlopedia – an open methods wiki (coming soon) – Illumina methods explorer– Illumina methods explorer – closed format
  11. 11. Core-facility community efforts – NGS mapped: finding other NGS cores – Single-cell QC: sharing data to speed technology adoption
  12. 12. NGS mapped http://omicsmaps.com
  13. 13. Single-cell QC – SCQC: a website for single-cell QC data analysis – 10XQC: the first example
  14. 14. Single-cell QC – Single-cell QC
  15. 15. Single-cell QC – Single-cell QC
  16. 16. Single-cell QC – Single-cell QC
  17. 17. Single-cell QC – Single-cell QC
  18. 18. A genomic case report – 3 year old with a life-threatening immunodeficiency – Routine immunological testing was uninformative – Presentation suggested genetic component – Possible STAT1 mutation • Led to debate about clinical path • HSCT considered ineffective
  19. 19. A genomic case report – Whole genome sequencing • Illumina TruSeq PCR-free • 30x coverage of trio • Interpretation of “exome” – NFKB1A mutation • de novo het c.94A>G in NFKBIA • results in IκBα serine>glycine (p.S32G where serine usually phosphorylated) • Loss of IkBa phosphorylation prevents normal signaling via degradation • IkBa signaling is vital adaptive and innate immune response
  20. 20. A genomic case report – NFKB1A mutation detected by rapid WGS – Transplant is likely to be curative • 9/10 HLA-A-mismatched unrelated donor • one of two patients worldwide alive with this condition (accepted for publication)
  21. 21. Acknowledgements – ABRF – Charlie Nicolet – Jaques RetiefIllumina • Steve Brumpton • Kevin Maxwee Illumina – James Thaventhiran – Matt Eldridge – My lab (Paul, Hannah, Fatimah, Marta, Ros, Johanna and Tom) – Bioinformatics Core(particularly Rich and Anne) – Phil Ewels SciLifeLab – Simon Andrews Babraham – Paul Coupland – Marta Grzelak