ABRF Next Generation Sequencing Group

The goals of the ABRF-NGS Study are to evaluate the performance of NGS platforms and to identify optimal methods and best practices. The study is a coordinated effort of five ABRF Research Groups (RG’s), involving over 20 core facility laboratories. The long-term goals of the ABRF-NGS study are to help optimize the detection of genetic variation with the latest sequencing tools and to establish a community resource for self-evaluation and self-improvement that will allow users of NGS technologies to readily compare and improve their own performance data and to evaluate existing and emerging protocols and platforms.

Additionally the ABRF NGS group evaluates new emerging sequencing technologies from a neutral prospective in regards to generally accepted reference standards

Membership

The ABRF-NGS Study is a collaboration between the DNA Sequencing Research Group (DSRG), Genomics Research Group (GRG), Nucleic Acid Research Group (NARG), Biomedical ‘Omics Research Group (BORG), and Genomics Bioinformatics Research Group (GBIRG).

The ABRF-NGS leadership group consists of scientific leads, project coordinators, and the chairs of all the RG’s that are involved in the study. Chris Mason (Weill Cornell Medical College; GBIRG) and Don Baldwin (Pathonomics LLC; GRG) are the scientific leads for the study. George Grills (Cornell University; EB), Charles Nicolet (University of Southern California; DSRG) and Scott Tighe (University of Vermont; NARG) are the project coordinators. Platform working groups focus on platform-specific protocols and generate data at core laboratory sites. A bioinformatics/bio-IT working group is focused on the analytical tools and approaches for each phase of the study and on the challenges of building a web-based community database resource.

Phase 1

The ABRF-NGS Study focused on RNA sequencing (RNA-seq), using standard Microarray Quality Control Consortium (MAQC) total RNA samples combined with External RNA Controls Consortium (ERCC) synthetic spike-in RNA. This RNA profiling phase of the ABRF-NGS Study included the Illumina HiSeq 2000/2500 and MiSeq, Roche 454 GS FLX+, Life Technologies Ion Torrent PGM and Proton, and Pacific Biosciences PacBio RS platforms. The study assessed sequencing accuracy, absolute and relative expression levels, RNA splice junction detection, and differential expression detection between samples. In a similar fashion, the currently ongoing Phase 2 of the ABRF-NGS Study is collaborating with the NIST Genome in a Bottle Consortium and other sequencing community stakeholders to utilize standardized reference genomic DNA samples.

Phase 1 participants included

• Don Baldwin (Pathonomics, LLC)
• Kip Bodi (Tufts University)
• Joe Boland (NIH/NCI/SAIC-Frederick)
• William Farmerie (University of Florida)
• Yuan (Gary) Gao (Lieber Institute for Brain Development)
• George Grills (Cornell University)
• Deb Grove (Penn State University)
• Belynda Hicks (NCI/SAIC-Frederick)
• Amy Hutchinson (NIH/NCI/SAIC-Frederick)
• Nadereh Jafari (Northwestern University)
• Dewey Kim (Johns Hopkins University)
• Ryan Kim (University of California, Davis)
• Shawn Levy (HudsonAlpha Institute)
• Sheng Li (Weill Cornell Medical College)
• Dawei Lin (University of California, Davis)
• Christopher Mason (Weill Cornell Medical College)
• Charles Nicolet (University of Southern California)
• Dayna Oschwald (New York Genome Center)
• Anoja Perera (Stowers Institute for Medical Research)
• Jaroslaw Pillardy (Cornell University)
• Nalini Raghavachari (NIH/NHLBI)
• Jeffrey Rosenfeld (University of Medicine and Dentistry of New Jersey)
• Peter Schweitzer (Cornell University)
• Todd Smith (Perkin Elmer/Geospiza)
• Scott Tighe (University of Vermont)
• Agnes Viale (Memorial Sloan Kettering Cancer Center)
• Chris Wright (University of Illinois)

Phase 2

The phase II study focuses on using DNA reference standards as an evaluation tool on existing and emerging NGS technologies, and is being done in collaboration with the National Institute of Standards and Technology’s Genome in a Bottle Consortium. Each of the three projects to be conducted in ABRF-NGS Phase 2 will feature samples that are either being evaluated by the Consortium for recommendation to NIST, or are already in the NIST Reference Material production process. These projects will provide multi-lab, multi-platform performance data for whole-genome sequencing of a mother/father/child trio, exome sequencing of formalin-fixed paraffin embedded cell lines to model archival tumor specimens, and small genome sequencing over a range of GC content using a panel of bacterial species. The resulting whole-genome data will provide a major contribution toward the very deep read counts necessary to establish the consensus sequences that will accompany the human trio NIST Reference Materials and the four bacterial Reference Materials. Genome in a Bottle members will use the cell line sequencing data to help decide whether these samples should be used for development of new references at NIST. Expected completion 2016.