Genome in a Bottle Consortium: New Sample T

hinkshop

September 2016 Workshop

New Samples

Thursday 15

th

 September 2016 1:30PM - 3:30PM

OVERVIEW

GIAB is planning to develop new whole-genome reference samples for both germline and

somatic contexts. This “thinkshop” will decide the principles for selecting, prioritizing, and

developing these new samples. A whitepaper, possibly to be developed into a ‘marker paper’

for publication in a widely read journal, will be used to guide the next phase of GIAB work.

● These samples are intended to serve as enduring scientific assets for development of

genome-sequencing measurement science.

○ They are intended to act as benchmarks and refere​ nces, or “etalons.”

Germline samples: an initial proposal is to develop ~8 new germline samples from diverse

ancestry groups from individuals in the PGP.

Cancer samples: an initial proposal is to develop tumor/normal pairing genomes from a fully

consented individual, using multiple, different cell lines derived from the same tumor.

Sample format: existing GIAB RM samples are gDNA from large batches of cells in ~50-100 kB

fragments; should new samples accommodate longer-read sequencing methods?

GERMLINE

Motivation for new samples

● The existing GIAB genomes represent limited ancestry groups.

○ Human genomes of different ancestries will pose different measurement

challenges.

○ Overrepresentation of limited ancestry groups can affect the development of

measurement science, reference genome resources, and bioinformatics tools.

Characteristics of new samples

● Similar in all ways to current GIAB samples
○ Authoritatively characterized

○ Stable and homogeneous

○ Publically available

○ Renewable

● PGP-consented (Explicit consent for public release of whole genome sequencing data

and commercial redistribution)

● Additional ancestry groups - particularly African, Hispanic, and mixed ancestries

● Available as cell lines (new samples would require fresh blood collection)

Plan for sample development and characterization

● Work with PGP and Coriell to establish appropriate cell lines and materials

○ Proposal based on PGP cell lines currently available from Coriell (single

individuals):

■ African American male

■ Puerto Rican male

■ Indian/White female

■ European/Brazilian/Portuguese female

■ Colombian male

■ PGP1 White male - has a BAC library, as well as fibroblast and iPSC lines

at coriell

■ Chinese/Filipino/Hispanic male

■ European, deeply phenotyped and characterized at Stanford

■ HuRef (not currently PGP)?, European, sequenced with Sanger and other

technologies

● GIAB community characterizes with multiple technologies

● NIST uses integration process to form high-confidence variant calls

Unaddressed needs/questions

● How does this effort relate to other efforts or other reference samples?

● Space to identify opportunity for GIAB portfolio development

SAMPLE FORMAT

Motivation for discussion

● As technologies evolve, what should GIAB samples look like?

RMs from NIST

● Authoritative samples of GIAB genomes
● Existing RMs (pilot, AJ Son, AJ Trio, Asian Son) are gDNA extracted from large batch cell

cultures

○ Batch of gDNA established as homogeneous and stable

○ Characterization is done largely from this gDNA batch

■ Some characterization (10X, BioNano, Complete Genomics LFR) done

from cell pellets

● Are these RMs needed?

● What’s the best format for dissemination?

Materials from biorepository

● Source cell lines for NIST RMs are available as cell lines, cell pellets, and extracted DNA

from Coriell in the NIGMS Repository

Commercial reference sample products

● Horizon GIAB FFPE samples

● Acrometrix Oncology Hotspot Control - AJ son with spike-ins

● Seracare - AJ son with somatic spike-ins, circulating tumor DNA, and cardiomyopathy

mutation spike-ins

SOMATIC

Motivation for new samples

(Arend Sidow, Stanford/JIMB)

● Facilitate technology development for calling somatic variants of all types in cancer

genomes

● There is limited availability of reference-grade characterized cancer genome samples

○ There are de facto references, but no formal references

● No currently available tumor-normal cell lines are explicitly consented for public release

of whole genome sequence data or commercial redistribution

● Starting to characterize cancer genome reference samples will enrich community

experience with difficult somatic variants

Experience with Tumor/Normal Pair Genomes as “A somatic

reference standard for cancer genome sequencing”

(David Craig, TGen, USC)

● Scientific Reports 6, Article number: 24607 (2016) - doi:10.1038/srep24607
Experience with somatic variation across a large tumor

(Noah Spies, JIMB)

Characteristics of new samples

● Explicit consent for public release of whole genome sequencing data

○ Is it possible to use a PGP consent?

● Homogeneous and stable DNA

○ Choose tumor types/cell lines that do not have high mutation rates (or

alternatively grow large batches of cells and extract DNA?)

● Tumor-normal pairs - ideally multiple tumor cell lines from one individual?

Plan for sample development and characterization

● Develop multiple cell lines from a single individual: tumor and normal

○ Develop 16 tumor cell lines?

■ Different cell lines from spatially distinct/distinct regions of a single tumor

■ Alternatively, create single cell line and culture it into different batches

that have different somatic mutations that occur during culture

● GIAB community characterizes with multiple technologies

● Cancer genome reference sample development is highly likely to enrich community

experience with difficult variants in difficult regions

Unaddressed needs

● How does this effort relate to other efforts or other reference samples?

● Space to identify opportunity for GIAB portfolio development