Conference Program - ABRF 2011 - Association of Biomolecular ...

156 Comparison of Custom TargetEnrichment Methods: Agilent vs.NimblegenK. Bodi 1 , P.S. Adams 2 , D. Bintzler 3 , A. Perera 4 ,K. Dewar 5 , D.S. Grove 6 , J. Kieleczawa 7 , R.H. Lyons 8 ,T. Neubert 9 , A. C. Noll 1 , S. Singh 10 , R. Steen 11 ,M. Zianni 121Tufts University, Boston, MA, United States;2Trudeau Institute, Saranac Lake, NY, UnitedStates; 3 DNA Analysis, Inc., Cincinnati, OH, UnitedStates; 4 Stowers Institute, Kansas City, MO, UnitedStates; 5 McGill University, Montreal, QC, Canada;6Pennsylvania State University, University Park,PA, United States; 7 Pfizer Research, Cambridge,MA, United States; 8 University of Michigan, AnnArbor, MI, United States; 9 New York University, NewYork, NY, United States; 10 University of Minnesota,Minneapolis, MN, United States; 11 Harvard MedicalSchool, Cambridge, MA, United States; 12 OhioState University, Columbus, OH, United StatesOver the last four years, we witnessed the tremendous advances inNext Generation Sequencing (NGS) that have dramatically decreasedthe cost of whole genome sequencing. However, the cost of sequencinglarger genomes is still significant. In addition and depending on thegoal of study, whole genome sequencing creates a large amount ofadditional/auxiliary data that complicates data analysis. There areseveral commercial methods available for isolating subsets of genomesthat greatly enhance the efficiency of NGS by allowing researchersto focus on their regions of interest. For the 2009-11 DSRG study,we compared products from two leading companies; Agilent andNimblegen, that offer custom enrichment methods. Both companiesobtained the same genomic DNA stock and performed DNA captureon the same specified regions. Following capture, the Illumina GenomeAnalyzer IIx system was used, in two different laboratories, to generatethe sequence data. We present our data comparing in terms of cost,quality, reproducibility and most importantly completeness and depthof coverage. Acknowledgements: We would like to thank Agilent,Illumina and Nimblegen for all their support in making this studypossible.157 A Methodology Study forMetagenomics Using Next GenerationSequencersD. Grove 1 , I. Albert 1 , D. Bintzler 2 , K. Bodi 3 ,M. Bruns 1 , K. Dewar 4 , G. Gloor 5 , T. Johnson 6 ,J. Kieleczawa 7 , R.H. Lyons 8 , T. Neubert 9 ,A.G. Perera 10 , S. Singh 6 , R. Steen 11 , M. Zianni 121Penn State University, University Park, PA, UnitedStates; 2 DNA Analysis, Inc., Cincinnati, OH, UnitedStates; 3 Tufts University, Boston, MA, UnitedStates, 4 McGill University, Montreal, QC, Canada;5University of Western Ontario, London, ON,Canada; 6 University of Minnesota, Minneapolis,MN, United States; 7 Pfizer Research, Cambridge,MA, United States; 8 University of Michigan, AnnArbor, MI, United States; 9 New York University, NewYork, NY, United States; 10 Stowers Institute, KansasCity, MO, United States; 11 Harvard Medical School,Cambridge, MA, United States; 12 Ohio StateUniversity, Columbus, OH, United StatesMetagenomics is one of several genomics applications, which hasbenefited immensely from the high throughput and cost efficacy ofNext Generation sequencers. And although hundreds of studies onmetagenome analysis have been published over the past few years,the methodology for conducting them is still very much evolving.In this DSRG study we will evaluate the influence of various samplepreparation methods, specifically DNA extraction and amplificationapproaches, on data output along with a comparative analysis of NextGeneration sequencing platforms. We will study the effect of thesedifferent experimental and technical strategies on determination ofsample biodiversity.158 What Your Blood Has to Say:Amplifying Blood RNA for theAffymetrix GeneChip® PlatformN. Supunpong Hernandez, T. Barta, C. Willis,R. Conrad, P. Whitley, K. BramlettLife Technologies, Austin, TX, United StatesPoster AbstractsGene Expression measurements from human blood RNA havebecome an increasingly important research area of focus. A reliableand consistent workflow to obtain RNA measurements from bloodwould serve to open the clinical arena to gene expression studies aspotential diagnostic indicators. The challenge lies in isolating highquality RNA from human whole blood at room temperature withoutcompromising gene expression profiles. To address this challenge, wereport a workflow using newly developed MagMax TM RNA isolation kitsfor Tempus TM stabilized blood and PAXgene® stabilized blood. Thesetwo stabilization methods are currently the most widely used methodsfor blood collection and stabilization in clinics in the United States.High quality RNA generated from these two RNA isolation processeswas amplified using the Ambion MessageAmp TM Premier RNAAmplification Kit and hybridized to Affymetrix GeneChip® microarrays.This platform and experimental tools are used, in combination, todemonstrate high quality gene expression profiles from human bloodRNA. The reported experimental design includes human whole bloodobtained from two donors collected and stabilized in either Tempus TMPAXgene® tubes. RNA was isolated using new MagMax TM RNA isolation76 • ABRF 2011 — Technologies to Enable Personalized Medicine