ABRF 2017 Annual MeetingA FORUM FOR ADVANCING TODAY’S CORE TECHNOLOGIES

TO ENABLE TOMORROW’S INNOVATIONS

March 25-28, 2017

Town and Country Resort & Convention CenterSan Diego, California

A profusion of confusion in genomic methods

Dr James HadfieldCRUK & Cambridge University

ABRF 2017 Annual MeetingA FORUM FOR ADVANCING TODAY’S CORE TECHNOLOGIES

TO ENABLE TOMORROW’S INNOVATIONS

March 25-28, 2017

Town and Country Resort & Convention CenterSan Diego, California

NGS method namingDescribe the problemPresent some examplesSuggestions on how to fix the problem

Core-facility community effortsA genomics case report

NGS method naming

– 10 years of NGS– 400 methods– No controlled vocabulary, no convention

“What does RNA-Seq mean?”

NGS method namingShendure & Lieberman: The expanding scope of DNA sequencing. Nat Biotech 2012

http://enseqlopedia.com/2013/09/finding-your-way-around-ngs-sample-prep

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

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

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

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

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

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

Core-facility community efforts

– NGS mapped: finding other NGS cores– Single-cell QC: sharing data to speed technology adoption

NGS mappedhttp://omicsmaps.comhttp://enseqlopedia.com/ngs-mapped

Single-cell QC

– SCQC: a website for single-cell QC data analysis– 10XQC: the first example

Single-cell QC

– Single-cell QC

Single-cell QC

– Single-cell QC

Single-cell QC

– Single-cell QC

Single-cell QC

– Single-cell QC

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

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 response3 week TAT in 2014

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)

Acknowledgements

– ABRF – Charlie Nicolet

– Jaques Retief Illumina

• 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