(W14-1) Tools for Next Generation Sequencing Data Analysis
Tufts University School of Medicine, Tufts University Core Facility, Boston, MA, United States
As NGS technology continues to improve, the amount of data generated per run grows exponentially. Unfortunately, the primary bottleneck in NGS studies is still bioinformatics analysis. There are a variety of workflows for NGS analyses that use open-source tools. This includes peak calling analyses for ChIP-Seq (MACS, GeneTrack indexer, Peak predictor), RNA-Seq (Tophat, Cufflinks), and finding small insertions, deletions, and SNPs using SAMtools.
Research Group Presentations
(R5b) Microarray Research Group Projects, 2010-11
D. Baldwin1, N.G. Reyero-Vinas2, N. Jafari3
1Penn Molecular Profiling Facility, University of Pennsylvania, Philadelphia, PA, United States; 2Jackson State University, Jackson, MS, United States; 3Northwestern University, Evanston, IL, United States
Members of the MARG will discuss our research projects: Comparison of microarray and deep sequencing platforms for microRNA profiling, Performance of a synthetic human microRNA reference panel, Participation in the SEQC Sequencing Quality Control consortium, and RNA-Seq profiling of environmental samples exposed to the Gulf oil spill.
120 Genome Technology Center at the NYU Langone Medical Center: New Support for Clinical and Translational Science
J. Zavadil, S. Mische
New York University Langone Medical Center, New York, NY, United States
To significantly enhance support for clinical and translational research within the framework of its CTSI, the NYU Langone Medical Center consolidated the Microarray and DNA Sequencing Cores into a new Genome Technology Center, a shared resource overseen by the Office for Collaborative Science.
128 NYULMC Office of Collaborative Science Cores – Enabling Personalized Medicine through Translational Research
C. Curchoe, T. Winner, J. Salcedo, S. Mische, D. Levy
New York University, Langone Medical Center, New York, NY, United States
The New York University Langone Medical Center (NYULMC) has committed $15 million to ensure that researchers have access to cutting edge enabling technologies. The Genome Technology Centers sequencing and microarray units are dedicated to RNA-seq of cancer patient transciptomes, array expression profiling and SNP genotyping, enabling translation research as well as the preparation of nucleic acids for molecular profiling, mutational analysis and microRNA screens.
132 Transcriptome Library Preparation Workflow to Enable Low Input RNA Amounts
C. San Jose Hinahon
Life Technologies, Austin, TX, United States
The SOLiDTM Total RNA-Seq (STaR-Seq) kit from Life Technologies provides a complete workflow for generating directional, random primed, whole transcriptome libraries from total RNA and fractionated RNA.
133 RNA-Seq Analysis with NextGENe Software
J. McGuigan, Y. You, C. Liu
SoftGenetics, LLC, State College, PA, United States
RNA sequencing is a powerful tool for interrogating the entire transcriptome at once. It allows identification of novel isoformsincluding gene fusions and alternative splicing- and expression level analysis across several orders of magnitude. NextGENe’s new RNA-seq application can be used to analyze data from any of the three main 2nd generation sequencing systems including Roche GS FLXTM and FLX Titanium, Illumina Genome Analyzers, and Applied Biosystems SOLiDTM Systems.
138 Use of Synthetic Transcript Pools to Evaluate RNA-Seq Performance and Analytical Methods
P. Whitley, L. Qu, A. Lemire, J. Brockman, S. Heater, J. Schageman, J. Gu, K. Lea, C. San Jose, N. Hernandez, D. Batten, K. Bramlett, D. Ilsley, C. Mueller, R. Setterquist
Life Technologies, Ambion R&D, Austin, TX, United States
RNA-Seq and related Massively Parallel Sequencing methods are rapidly becoming standardized platforms for transcriptome studies . Although RNA-Seq is now being used routinely, there has not been a thorough investigation of fundamental performance metrics using standardized reagents and analytics.
150 ScriptSeq RNA-Seq Library Preparation Method: A Simplified Work-Flow for Directional NGS RNASeq Library Preparation with Whole- Transcript Representation
A. Khanna1, R. Sooknanan2, J. Hitchen2, A. Radek2
1Epicentre BIOtechnologies, Madison, WI, United States; 2RiboTherapeutics Inc., Saint Laurent, QC, Canada
RNA sequencing is an emerging revolutionary tool for wholetranscriptome analysis that provides information about the structure of transcripts and their expression levels. Current methods for making sequencer-specific di-tagged DNA fragment libraries for RNA-Seq typically comprise preparing rRNA-depleted RNAand either (i) RNA fragmentation, 5’ and 3’ adaptor-ligation, size selection, cDNA synthesis, and multiple clean-up steps; or (ii) cDNA synthesis followed by cDNA fragmentation, end-polishing, 5’ and 3’ adaptor-ligation, size selection and multiple clean-up steps.
163 Transcriptome Analysis Using Next- Generation Sequencing Technology
K. Bramlett, K. Lea, L. Qu, P. Whitley, J. Schageman, J. Gu
Life Technologies, Carlsbad, CA, United States
High throughput RNA sequencing (RNA-Seq) is becoming increasingly utilized as the technology of choice to detect and quantify known and novel transcripts. Multiple next-generation sequencing (NGS) platforms are available that enable transcriptome profiling through RNA-Seq workflows.
168 MicroRNA Analysis Using RNA Extracted from Matched Formalin- Fixed Paraffin-Embedded (FFPE) and Fresh Frozen Samples on SOLiDTM System
K. Lea, J. Gu, E. Zeringer, S. Heater, J. Schageman, C. Mueller, K. Bramlett
Life Technologies, Carlsbad, CA, United States
Archived formalin-fixed paraffin-embedded (FFPE) specimens represent excellent resources for biomarker discovery, but it has been a major challenge to study gene expression in these samples due to mRNA degradation and modification during fixation and processing. MicroRNAs (miRNAs) regulate gene expression at post-transcriptional level and are considered as important regulators of cancer progression.