Measurements for Reference Material Characterization Working Group Summary Aug2012
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This document discusses plans for fully characterizing a reference genome sample. It addresses determining library preparation protocols, sequencing platforms, difficult regions to sequence, error correction and verification approaches, whether to generate new or use existing data, and other considerations like dividing the genome into easy, medium and hard to sequence regions. The goal is to develop a consensus plan to experimentally characterize a reference material.
Measurements for Reference Material Characterization Working Group Summary Aug2012
- 1. Measurements for Reference Material Characteriza4on Develop a consensus plan for experimental characteriza4on of Reference Materials Genome in a Bo;le Working Group
- 2. Determine library prepara4on protocols • Fosmid and/or a BAC library? • Need to keep some cells for plaGorm-‐specific DNA purifica4on needs (e.g., to get high-‐ molecular-‐weight DNA) • Include sequencing family members
- 3. Determine sequencing plaGorms • Illumina – 300-‐600bp inserts – 3ish kb mate-‐pair – 2x100bp reads on HiSeq • PacBio – >10kbish inserts – 1x90m “movies” – Who will do it and pay for it? – PacBio may be able to create the libraries – Chris Mason to sequence? • Life Tech – 5500 data at NIST (1,6, and 10kb mate pairs) – Ion Torrent/Proton data (who to generate?) • Complete – Standard libraries and LFR approach – Start with cells • 454? – 700-‐800bp reads? • Newer technologies – are they used for verifica4on/valida4on only? – Oxford Nanopore? – GnuBio?
- 4. “Error Correc4on” and Verifica4on (not valida4on) • ArrayCGH and SNP Chip • OpGen (or other op4cal mapping approaches) • Targeted sequencing
- 5. Difficult to sequence regions • Characterize MHC regions? – Use 454? • Other parts of the genome? • Approved CLIA-‐cer4fied specific tests? – Highly mul4plexed TaqMan/Sanger style assay • CLIA-‐cer4fied whole exome data • Do we pick very specific well-‐characterized fosmids?
- 6. Old vs New Data • What new data will we need to generate on the actual reference sample? • Can we use data that currently exists? • Or generate data now or vanilla Coriell samples?
- 7. Other Thoughts • BAC by BAC approach? • Divide the genome into the easy, medium, and hard bits – Easy = same call all the 4me on every plaGorm • How do we account for technological/ algorithmic improvements?