FALL 2011

PacBio Customers Present Data at ICHG/ASHG Annual Meeting

Customers from the Ontario Institute for Cancer Research (OICR), UC Davis, the Broad Institute, GATC Biotech and Cold Spring Harbor Laboratory presented data (primarily human) generated using the PacBio RS at the International Congress of Human Genetics and American Society for Human Genetics Annual Meeting in Montreal October 11-15.

TECHNOLOGY WORKSHOP

The Pacific Biosciences technology workshop "Single-Molecule Sequencing: Insights Into Clinical Utility," featured presentations by Drs. Andrew Brown of OICR and Ryan Poplin of the Broad Institute.

Using the PacBio RS platform, OICR has developed methods for the rapid characterization of a validated set of known oncogenes to uncover potential actionable mutations that can guide therapy. The trial involves patients with advanced metastatic cancers who are potential candidates for early phase clinical trials of targeted agents. For the pilot study, OICR is focusing on 19 oncogenes that overlap the Oncocarta V.1 panel (Sequenom, Inc.) to allow for cross technology comparisons. However, to fully take advantage of the sequencing yield of the PacBio RS, they are expanding to a larger list of ~200 genes.

Andrew Brown, Ph.D. of OICR discussed their work to optimize methods for amplicon sequencing. He described methods for DNA quantitation, best amplicon size, and CCS analysis pipeline. This combination allowed OICR to generate up to 150 Mb per SMRT® cell. In a recent patient study, they demonstrated the ability to go from patient consent to a final genomics report in less than three weeks, with 50% of the cases resulting in actionable information. The clinical resequencing is being done in collaboration with a diagnostic CAP/CLIA molecular diagnostic laboratory to provide validation of all results prior to utilization.

Ryan Poplin, Ph.D. of the Broad Institute explained how to utilize GATK with PacBio data. Dr. Poplin stated that PacBio's random error profile combined with the known quality score resulted in highly accurate variant calls and fewer false positives than other methods. He showed that with GATK, the quality scores can be recalibrated, resulting in PacBio data being best-in-class for variant validation. He shared examples of PacBio overturning a few Illumina variant calls and confirmed through further investigation that PacBio made the right call. One remaining challenge, easily solved through algorithm development according to Dr. Poplin, is that there is some reference bias due to the PacBio indel errors. This workflow will be turned into a pipeline in a future release of GATK along with a future paper on this work.

POSTERS

The following technical posters were also presented at the meeting. Click on the link to access a PDF of the poster.

• Single Molecule Targeted Resequencing to Individualize Cancer Therapy
  A. Brown, et al., Ontario Institute for Cancer Research
• Content Sequencing of Expanded CGG Repeats in the FMR1 Gene
  E. Loomis, et al., UC Davis
• Combining Short (Illumina) and Long (PacBio) NGS Reads to Improve de novo Genome Assemblies of Rice and Saccharomyces Cerevisiae Genomes
  M. Schatz, et al., Cold Spring Harbor Laboratory