Next Generation Sequencing

Del Mar Photonics - Plasmonic nano-materilas for Raman spectroscopy - HeroN AFM - NanoRaman System system

Next Generation Sequencing - Day 1

TUESDAY, MARCH 17

1:00 pm Conference Registration

SEQUENCING TECH EXPO

Next-Generation Sequencing is alive, thriving, and driving discovery. As costs come down and ease increases, these new, massively parallel high-throughput sequencing platforms are infiltrating multiple aspects of traditional biological research. However, each next-generation sequencing platform best lends itself to specific sequencing goals. This Tech Expo showcases the next-generation sequencing platforms to help you make informed purchasing decisions. Sponsored Seminars Hosted by:

2:00 Chairperson’s Remarks

Kevin Davies, Ph.D., Editor-in-Chief, BioIT World

2:05 Applied Biosystems logo The SOLiD™ 3 System - Taking Next-Generation Sequencing to the Next Level

Michael Rhodes, Ph.D., Product Applications Senior Manager, Genetic Analysis, High Throughput Discovery, Applied Biosystems
The new SOLiD™ 3 System achieves new milestones in throughput in excess of 20 Gb of mate paired sequence data from a single run and 30-40 Gb of demonstrated throughput in Applied Biosystems R&D labs. Maintaining high accuracy, improvements in read length and unique mate-pair library strategies, the news system enables expanding applications from whole genome resequencing and SNP discovery to miRNA profiling. This presentation will review various applications that the new system capabilities enable and discuss an example of whole genome transcript profiling in single cells and whole genome resequencing for SNP and structural rearrangement discovery.

Helicos logo2:35 Enabling True Biology with Helicos™ Single Molecule Sequencing

Patrice M. Milos, Ph.D., VP & CSO, Helicos BioSciences Corporation

Helicos True Single Molecule Sequencing (tSMS)™ provides a unique view of genome biology through the direct sequencing of cellular nucleic acids in an unbiased manner providing both quantitation and sequence information. Using a simple DNA sample preparation which requires no ligation or PCR amplification genomic DNA is sheared, tailed with polyA and readied for hybridization to a flow cell surface containing oligo dT for initiating the sequencing by synthesis reactions. To demonstrate the fidelity and scale of the HelicosTM Genetic Analysis System three bacterial genomes were sequenced, each of distinctly different genomic contents in single flow cell channels of the available 50 channels. We have extended our research to include a variety of genomic targets including candidate gene regions, yeast and C. elegans, all with similar accuracy and coverage. The ability of our single molecule sequencing platform to provide quantitative measurements of genome biology include research efforts in small RNA measurements, assessment of copy number variation of human samples and a simple method for quantitative assessment of the transcriptome, digital gene expression – all without the requirement of ligation or amplification – a hallmark for measuring the biology of cells.

3:05 Refreshment Break, Poster and Exhibit Viewing

Illumina logo3:30 The Illumina Genome Analyzer - Transforming Systems Biology

Abizar Lakdawalla, Ph.D., Senior Product Manager, Sequencing
Applications, Illumina

The Genome Analyzer next-generation sequencing system has transformed our understanding of genome variations, epigenomics, transcriptomics, and the interaction of proteins with DNA and RNA. A comprehensive description of the Genome Analyzer system will be presented with effective approaches to address broad systems biology questions. Strategies and tools derived from sequencing multiple human genomes, large numbers of transcriptomes, and extensive ChIP-Seq samples will be described to maximize the data and sample throughput with the simple and easy-to-use Genome Analyzer workflow.

Roche 454 logo4:15 Moving Next Generation Sequencing into the Clinical Research Market:
Timothy Harkins, Ph.D., Director, 454 Sequencing Roche Applied Science, Roche, Inc
The Genome Sequencer FLX is now generating the longest reads within the next gen market with over 1 million unique sequencing reads that are 400 to 500 base-pairs in length. With a fast instrument run time of 10 hours and the ability to quickly analyze the sequencing data, projects involving 1,000’s of samples are able to be processed readily. The projects that will be presented include:
1) Sequencing HIV to detect low frequency drug resistant mutations
2) HLA sequencing – the most known polymorphic regions within the human genome
3) Using NimbleGen Sequence-Capture arrays to sequence the whole human exome
4) Detecting novel pathogens in complex environmental samples

4:45 Interactive Panel Discussion with Sequencing Leaders

Moderator; Kevin Davies, Ph.D., Editor-in-Chief, BioIT World

The term “next-generation” has become the“now generation.” As the genome unit price of these next-gen platforms continues to tumble, excitement is growing about the scientific and commercial potential of third-generation sequencing systems, from single-molecule methods to nanopores to ‘nanoball’ service models. Here, leaders from established and emerging next-gen platform providers trade insights on the latest scientific and technological advances, and answer your questions.

GenomeQuest5:30 Reception Hosted by GenomeQuest

Preparing Your Enterprise for Next-Generation Science
Ron Ranauro, President & CEO, GenomeQuest, Inc.
Those acquiring next-generation sequencing instruments are faced with a dilemma for managing the deluge of data – choose between building their own in-house system or using low-data-volume desktop tools. A sequence data management platform provides the best of both choices for managing and analyzing massive volumes of sequence data, and gets researchers back to the science, faster.

7:00 Close of Day

Next Generation Sequencing - Day 2

WEDNESDAY, MARCH 18

7:00 am Registration and Morning Coffee

Sponsored by
Rain Dance7:30 Breakfast Presentation
Targeted Sequencing Using Droplet-Based Microfluidics

James Brayer, Manager of Commercial Scientific Applications, RainDance Technologies, Inc.

Recent advances in DNA sequencing technologies have improved accuracy and dramatically reduced the cost of DNA sequencing. However, even with the improved efficiency of these second-generation systems, sequencing thousands of whole human genomes across various phenotypes is expensive and time consuming. In order to exploit the full potential of these new targeted sequencing techniques, a robust method for isolating biologically relevant genomic loci on the megabase scale will be required. The Sequence Enrichment application from RainDance Technologies leverages the sensitivity and specificity of PCR in a novel droplet-based format which avoids the limitations of traditional multiplex amplification or hybridization methods. The session will focus on: Key aspects of the targeted sequencing workflow that directly impact the quality of the data; An introduction to the RainDance RDT 1000 instrument and Sequence Enrichment application; A discussion of how the Sequence Enrichment process integrates into the targeted sequencing workflow for both short-read and long-read sequencing platforms; and example data from targeted sequencing of thousands of genomic loci with a focus on completeness, uniformity and specificity.

8:30 Chairperson’s Remarks

Richard Gibbs8:35 KEYNOTE PRESENTATION

Richard Gibbs, Ph.D., Director and Professor, Human Genome Sequencing Center, Baylor College of Medicine

TARGETED SAMPLE ENRICHMENT

9:15 SCODA Electrophoresis for Biomolecule Concentration

Andre Marziali, Ph.D., Director, Department of Physics and Astronomy, University of British Columbia

We have developed a novel electrophoretic concentration technology for efficiently purifying and concentrating biomolecules. The technology offers unique advantages in biomolecule concentration including exceptional rejection of PCR inhibitors, an unparalleled ability to enrich for low abundance nucleic acids, and ability to length-select nucleic fragments. Additionally, we will present recent advances in the SCODA (Synchronous Coefficient of Drag Alteration) technology, including reduction of sample processing times to less than ten minutes and applications to protein concentration.

9:45 Probing the Epigenome of Induced Pluripotent Stem Cells with Methylome Partitioning

Kun Zhang, Ph.D., Assistant Professor, Department of Bioengineering, University of California San Diego

DNA methylation is one of the primary epigenetic regulatory mechanisms that involve in normal developmental processes. We have recently developed a method for specific capture of an arbitrary subset of genomic targets using padlock probes, and for digital quantitation of DNA methylation at the single nucleotide resolution. We used this method to characterize the changes of DNA methylation during the de-differentiation of human fibroblasts into induced pluripotent stem (iPS) cells and pluripotent hybrid stem cells. We found that the phenotypic changes during the reprogramming are associated with local and specific alteration of methylation close to a small set of genes.

10:15 Targeted Extraction of Specific Non-Contiguous Loci on Mouse
Chromosome 1 forSponsored by
FEBITnewlogo Next-Generation Sequencing with HybSelectTM

Michelle Lyles, Ph.D., Vice President of Marketing and Sales, febit gmbh

The introduction of next-generation sequencers has lead to a dramatic increase in sequencing throughput. Besides whole genome de novo sequencing of small genomes, high throughput resequencing of selected regions of a large, eukaryotic genome is now available. However, these selected regions have to be separated from the remaining chromosomal DNA first due to the high complexity of the sample. Methods like PCR amplification are time-consuming and very expensive. We report an approach termed HybSelect™, which makes use of target-specific DNA extraction using Geniom® Biochips. Up to 120,000 capture probes specific for the genomic region of interest are synthesized on the Biochip and hybridized with the sample which can be analyzed via next-generation sequencing after washing and elution. Geniom® Biochips offer several advantages over regular DNA microarrays, including highly flexible probe content and a unique microfluidic architecture, enabling complete automation of the process and minimizing the required sample amount.

10:30 Coffee Break

11:00 Combination of Microfluidics and Next-Generation Sequencing for
Targeted Sequencing of Multiple Genomic Loci

Ewen Kirkness, Ph.D., Investigator, J. Craig Venter Institute

The targeted resequencing of discrete genomic loci in human populations is a growing research area where many new methodologies are currently being developed. Previously, targeting has been achieved reliably by PCR, though the procedures applied have been relatively low-throughput and expensive. In order to fully exploit the potential of new sequencing technologies for targeted resequencing, we have employed the RainDance droplet-based technology platform. This generates picoliter-volume PCR reactions, and a sequence enrichment application uses a library of PCR primers in droplets to amplify hundreds of genomic loci in a single tube. We have applied this technology to several human resequencing projects, and compared result to conventional PCR-based approaches.

11:30 FEATURED PRESENTATION
New Approaches in DNA Sequencing

Ronald W. Davis, Ph.D., Professor, Biochemistry & Genetics, Stanford Genome Technology Center

12:00 pm Close of Morning Session

Sponsored by
Applied Biosystems logo12:15 Luncheon Presentation
Sample Enrichment Strategies for Sequencing with the SOLiD™ System

Michael Rhodes, Ph.D., Product Applications Senior Manager, Genetic Analysis, High Throughput Discovery, Applied Biosystems

Targeted resequencing strategies have been a key focus for translational genomics. There are multiple techniques for accomplishing this and various commercial alternatives. Is there a right one? This discussion will outline the different pathways to enrich samples for use on next generation sequencers, in particular the SOLiD ™ 3 System. The SOLiD ™ 3 System is capable of providing 10X human genome coverage in a single run. At what point is whole genome sequencing a more appropriate approach?"

EXPANDING THE ENVELOPE OF APPLICATIONS

2:00 Chairperson’s Remarks

2:05 Capturing and Sequencing the Protein Coding Genome

Jay Shendure, Ph.D., Assistant Professor, Genome Sciences, University of Washington

Next-generation sequencing technologies have reduced the cost of DNA sequencing by several orders of magnitude. While the routine resequencing of full human genomes continues to be prohibitively expensive for studies involving large numbers of individuals, the cost of resequencing the ~1% of the human genome that is protein-coding, may soon be on par with that of dense genotyping arrays. We are exploring several strategies for massively multiplex capture of discontiguous genomic subsequences, a prerequisite for efficient PCG resequencing. These include an approach based on oligonucleotide libraries derived from programmable microarrays, which then serve as Molecular Inversion Probes, while another approach involves using dense microarrays for capture-by-hybridization. By using HapMap samples and coupling both capture methods to high-throughput sequencing on the Illumina platform, we have observed high sensitivity and specificity for variant discovery at well-covered positions. Ongoing work is primarily directed at optimizing for capture uniformity and efficiency.

2:30 Sequencing Out the Stem Cell Epigenome

R. David Hawkins, Ph.D., Ludwig Institute for Cancer Research, University of California San Diego School of Medicine

We are using ChIP-Seq to identify the location of various histone modifications in the human embryonic stem cell epigenome. We are able to identify distinct patterns, some of which correlate with genomic regulatory elements. An extensive investigation into these patterns should shed light on their role in gene transcription.

2:55 Shading Light into the Composition of Natural Micro- bial Communities using Multiplex Pyrosequencing

Lutz Krause, Ph.D., Bioinformatics, Bioanalytical Science Department, Nestlé Research Center, Switzerland

The recently published multiplex pyrosequencing allows the simultaneous sequencing of hundreds of microbial 16S DNA samples at a low cost. A validation study will be presented, demonstrating that the reconstructed community composition of the analyzed samples strongly depends on the used PCR primers and amplified regions. Furthermore, the power of multiplex pyrosequencing to investigate the taxonomic composition of entire microbial communities is high-lighted on a real-world example.

3:25 Refreshment Break

NEXT-NEXT GEN

This session showcases the next-next or “third” generation of sequencing technologies.

4:00 Complete Genomics: Revolutionizing Human Genome Sequencing
Radoje Drmanac, Ph.D., Chief Scientific Officer, Complete Genomics
Researchers need to compare large numbers of human genomes to be able to characterize rare genetic variants in order to discover the missing links between our genes and diseases. To facilitate that discovery process and provide affordable complete genetic diagnostics, We have developed a new sequencing platform that capitalizes on its advances in both DNA arrays and sequencing assays. Its platform employs the first sub-micron arrays, which are populated with DNA nano-balls™, and uses a non-sequential read technology, referred to as combinatorial probe-anchor ligation or cPAL™, that reduces both reagent consumption and imaging time. Another unique feature of its platform is its long fragment read technology (LFR™) that generates separate sequences for homologous pairs of parental chromosomes. This capability is critical to sequencing diploid human genomes, as it allows heterozygote phasing over large intervals (potentially entire chromosomes), even in areas with high recombination rates. This approach can be used to resolve extensive rearrangements in cancer genomes and determine full-length sequences of alternatively spliced transcripts. We will share its latest genome sequencing data results and provide an update on its technology progress.

4:20 Single Molecule Mega-Genomic Analysis in Nanochannel Array

Michael Boyce-Jacino, Ph.D., President and CEO, BioNanomatrix, Inc.

The BioNanomatrix technology analyzes individual long strands of native genomic DNA in a massively parallel format, avoiding the fragmentation and complex data re-assembly required by other approaches. Highly standardized, semi-conductor fabrication based manufacturing methods promise highly reliable, inexpensive methods for ultra-small sample genome analysis and diagnostics. The affordability, speed and simplicity of the technology are expected to make the routine use of genetic information feasible in broad-ranging applications in sequencing, SVs analysis, molecular diagnostics and personalized medicine.

4:40 The PinPoint Sequencer: High Throughput, Low Cost DNA Sequencing

Jerzy Olejnik, Ph.D., Vice President, Process R&D, Intelligent Bio-Systems, Inc.

Intelligent Bio-Systems’ next-generation sequencing platform is based on proprietary sequencing by synthesis (SBS) chemistry invented at Columbia University. The system utilizes single-stranded, clonally amplified DNA fragments as templates and incorporates specially labeled nucleotide analogs into a growing complementary second strand one base at a time. The instrument design leverages inherently rapid extension and cleavage times that result in higher throughput and lower cost than other currently available systems. The high performance of the PinPoint instrument is achieved by combining fast chemistry cycles, a high speed imaging system and an efficient, ordered-array-based chip.

5:00 Solid-State Nanopores as Single-Molecule Hybridization Detectors for Genomic Sequencing

John Oliver, Ph.D., Vice President Research, NABsys, Inc.

NABsys is developing a new sequencing platform that uses solid-state nanopores as single molecule detectors. Genomic DNA will be randomly cleaved to generate long fragments. The fragments will be hybridized with oligonucleotide probes. The hybridized strands are translocated through a nanopore and the relative positions of the probes are determined. The data for each probe is used to assemble the fragments into probe map. The sequence of the genomic DNA is reconstructed from a collection of probe maps. The method promises to be low cost, and to retain long range information.

5:20 Commercialization of Lightning Terminators™, LaserGen’s Next-Generation Reversible Terminator Chemistry

Michael L. Metzker, Ph.D., President and CEO, LaserGen, Inc.
The demand for DNA sequence information has never been greater, yet current Sanger technology is expensive, time consuming, and labor intensive to meet the ongoing demand. To overcome this challenge, we have developed a novel sequencing platform called cyclic reversible termination (CRT), which differs significantly from Sanger sequencing and pyrosequencing. CRT comprises three steps: incorporation, imaging, and cleavage. At the heart of the CRT method is the reversible terminator (RT). Several groups have created a variety of modified nucleotides, whose function directly affects the performance of the sequencing platform. To enable these nucleotides, a variety of mutations are introduced into the DNA polymerase. This improves incorporation, but at the expense of specificity. We have constructed a microfluidic device that integrates all cycle steps of the CRT method, including reagent delivery and temperature control, UV cleavage, and fluorescence imaging. The advantages of the CRT method are demonstrated by the sequencing of E. coli1655 using a mate-pair, emulsion PCR protocol, the genome of which is useful in providing a benchmark for comparing sequencing metrics such as read-length, accuracy, and coverage with other next-generation platforms.

5:40 Question & Answer Session with Next-Next Gen Speakers

Hosted by
Roche 454 logo6:00 Workshop
(Cocktails & Hor’dourves Served)

Next Generation Sequencing - Day 3

THURSDAY, MARCH 19

Sponsored by
Fluidigm7:30 am Breakfast Presentation

Implementation of Fluidigm’s SlingShot Absolute Quantitation Product Offers the Dual Advantage of Replacing the Costly and Time-Consuming Titration Step as Well as Recovery of Sub-Optimal DNA Samples for 454 Next Generation Sequencing

Joseph Boland, Ph.D., M.S., Dedicated Scientific Operations Leader, SAIC-Frederick, Inc., The National Cancer Institute

We have successfully implemented Fluidigm’s SlingShot technology into our 454 sequencing process at two key junctions: sub-optimal sample quantitation and replacement of the library titration. This implementation has allowed us to process previously failed samples (Roche recommends samples be between 3ug and 5ug) and replace the costly titration step saving us processing time (up to 2 days) plus the cost of a sequencing reaction.
Our initial experiment utilizing SlingShot consisted of 12 samples ranging in amounts from 990ng to 2.5ug. We processed these 12 samples using the approved Roche 454 protocol through library prep. After completion of the library prep, each sample was quantitated using the SlingShot product to accurately assess the amount of sample to go forward into bulk emulsion prep. Due to limitations of the existing quantitation methods available at the core genotyping facility (Ribogreen and Agilent), these samples would have never made it to the sequencer. We will present data highlighting the quality coverage attained (each sample had at least 20X coverage) from these samples using the quantitation obtained through SlingShot. We will also discuss future experimentation as well as future updates we would like to see with this platform.

8:15 Successful Sequencing Discussion Groups
Grab a cup of coffee and join a facilitated discussion group focused around specific themes. This unique session allows conference participants to exchange ideas, experiences, and develop future collaborations around a focused topic. Topics include:

Table 1: Keeping Up with a Next-Gen Sequencer: Alternatives to Owning a Giant Computer Cluster
Host: Martin Gollery, CEO, Tahoe Informatics
Discussion focus topics include:

* FPGA acceleration- the current range of choices
* Supercomputing on a graphics card
* Tesla workstations
* Cloud computing

Table 2: Getting Most of Your Illumina Genome Analyzer II
Host: Iwanka Kozarewa, Ph.D., Sequencing Technology Development, Wellcome Trust Sanger Institute
The discussion will be focused on key steps in sample preparation procedure using Illumina and sequencing flow as well as on some of the most promising applications of next-generation sequencing. Discussion topics include:

* Quantification
* Multiplexing
* Targeted sequencing
* Bioinformatics tools for read alignment, SNP calling and de novo assembly

Table 3: Next Generation Storage Networking
Host: Jacob Farmer, CTO, Cambridge Computer
Topics include:

* Fundamentals of storage virtualization: the storage I/O path
* Shortcomings of conventional SAN and NAS architectures
* In-band and out-of-band virtualization architectures
* The latest storage interfaces: SATA (serial ATA), SAS (serial attached SCSI), 4Gb Fibre Channel, Infiniband, iSCSI
* Content-Addressable Storage (CAS)
* Information Life Cycle Management (ILM) and Hierarchical Storage Management (HSM)
* The convergence of SAN and NAS
* High-performance file sharing, SAN-enabled file systems and parallel file systems
* Wide-area file systems (WAFS)

Table 4: Deep Sequencing of Small RNAs in hESC: Exploring Epigenetic Networks in Development
Host: Ron Hart, Rutgers University, and Loyal Goff, CSAIL & Broad, MIT
Discussion to include:

* Preparing small RNA libraries
* Genomic alignment in colorspace
* Bioinformatic analysis of predicted microRNAs
* Protein coding and Non-coding RNAs in hESC

Table 5: Pyrosequencing to Pipeline Development
Host: Alla Lapidus, Ph.D., Group Leader, Joint Genome Research Institute
Topics include:

* Sanger vs. pyrosequence vs. ultra short reads
* QC approaches
* Metagenomic assemblies: assemble or not?
* New tools for metagenomic assembly
* Ability to annotate short and VERY short reads
* Tremendous amount of genes in an unassembled pyrosequenced data set
* Pipeline development

Table 6: Complete Genomics: Rapid, Accurate, Affordable Human Genome Sequencing
Host: Radoje Drmanac, Ph.D., Co-founder and Chief Scientific Officer, Complete Genomics
Discussion topics include:

* The promise: How Complete Genomics will deliver
* Outside of the box: Sequencing as a service
* Medically-relevant human genome sequencing: Elucidating the genetic basis of disease

Table 7: Clinical Utilities of the Next-Generation Sequencing Technology: Issues Before Implementation in Routine Laboratories
Host: Bob Chou, Ph.D., Scientist, Research and Development/Molecular Sequencing and Genetics, ARUP Laboratories
Discussion topics include:

* Data management and analysis
* Guidelines for QC/QA of sequencing data
* Optimization of pre-NGS sample processing steps
* DNA sample requirements for NGS sequencing

Table 8: Using magnetic beads in Sample Prep Applications for Sequencing
Host: Patrick J. Finn Ph.D., Director of Research and Development, Agencourt Bioscience
Clean sample preparation is important to any successful application. Using high performance paramagnetic bead-based technologies immobilizes nucleic acids onto magnetic microparticles. This technology has set an industry standard in sample preparation for Sanger sequencing where products, such as the the Agencourt® CleanSEQ® and Agencourt AMPure® product lines, have been essential reagents for labs engaged in high throughput Sanger sequencing. With the emergence of second generation sequencing systems such as Roche’s 454 FLX, Illumina’s GAII and Life Technologies SOLiD system, the this technology is re-emerging as a key reagent for library sample prep upstream of each sequencing system.

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DATA MANAGEMENT AND ANALYSIS

9:00 Chairperson’s Remarks
Michael Rhodes, Ph.D., Product Applications Senior Manager, Applied Biosystems

9:05 Gene Wiki: Community Intelligence Applied to Gene Annotation

Andrew Su, Ph.D., Group Leader, Computational Discovery, Genomics Institute of the Novartis Research Foundation

There are ~25,000 genes in the human genome, and evidence shows we’ve only scratched the surface in terms of describing their function. It is becoming increasingly clear that the research community cannot solely rely on curators at annotation authorities to annotate genes in databases. Therefore, following broader trends in information management, we present two efforts to harness the power of community intelligence. First, the”Gene Wiki” enables the scientific community to collaboratively summarize gene function in real time. Second, BioGPS offers a gene annotation portal that the bioinformatics community and data providers can collaboratively develop and extend.

9:35 Assembling Short and Error-Prone DNA Reads: Does the Read Length Matter?

Mark Chaisson, Ph.D., Postdoctoral Scholar, Computer Science Engineering, University of California, San Diego

Current sequencing technologies attempt to maximize read length without sacrificing base calling accuracy. Since fragment assembly with inaccurate reads is difficult, the common practice is to trim the inaccurate tails of the reads. We present EULER-USR tool for assembling short reads that bypasses the problem of limited accuracy in the tails of reads. An important and counterintuitive implication of this result is that one may extend sequencing reactions “past their prime”’ to where the error rate grows above what is normally acceptable for fragment assembly. We compare EULER-USR with other short read assemblers and illustrate its applications for assembling mate-paired Illumina reads. We further use EULER-USR to study the question “Does the read length matter?” and to compute the “read length barrier”. We show that increasing the read length beyond the read length barrier does not improve the quality of assembly.

10:05 Estimation of Evolutionary Parameters, and Design of Association Studies Using Resequencing Data

Paul Marjoram, Ph.D., Assistant Professor, Preventative Medicine, Keck School of Medicine, University of Southern California

We will demonstrate how data from next-generation sequencing technologies can be used to estimate evolutionary parameters (mutation and recombination rates), despite the fact that the data does not necessarily provide complete sequence information. We will also present two refinements of our methods: one that is more robust to sequencing errors and another that can be used when no reference genome is available. Finally, we will discuss the implications of these technologies on the design of association studies, and how the power of such studies varies with differing levels of coverage.

10:35 Refreshment Break and Poster Viewing

11:00 Poster Awards Sponsored by Oxford Nanopore

11:10 Next-Generation Sequencing of Humans

Pauline Ng, Ph.D., Senior Scientist, Genomic Medicine, J. Craig Venter Institute

We are now in an exciting era where individual human genomes and large genomic regions from human populations can be sequenced. This has been made possible by next-generation sequencing technologies such as Roche 454, ABI SOLiD, and Illumina GA. We have applied NGS technologies to an individual human genome and to population-based sequencing. We discuss the performance of these technologies and how this has improved our understanding of what it will take to achieve a $1,000 genome.

11:30 End-User Presentation: Next-Gen Sequence Data Analysis Reveals Grapevine Virus
Maher Al Rwahnih, Ph.D., Department of Plant Pathology/FPS, University of California
The informatics infrastructure needed for aligning and annotating next-generation reads against reference sequence data presents a daunting challenge for researchers. Plant pathologists at the University of California, Davis are facing a novel grapevine disease that they suspected may be caused by a previously uncharacterized RNA virus. They sampled and sequenced the overall RNA content of the cells of both diseased and apparently healthy grapevines to generate about 400,000 reads with an average length of about 200 nucleotides. Pairwise comparisons would create tens of trillions of alignments, taking weeks on a typical computer, creating an intractable situation.
This case study will demonstrate how these UC Davis researchers overcame computational and analysis limitations to annotate, assemble, and contig the next-gen reads and ultimately identify a new virus, using On-Demand Informatics that minimized cost and delivered timely results.

12:00 Close of Morning Session

12:15 Luncheon Presentation Sponsored by Agilent Technologies
Targeted Resequencing by Agilent in-Solution Genome Partitioning in Combination with the Illumina Genome Analyzer Sequencing Platform
Lira Mamanova, Ph.D., Sequencing Technology Development Group, Wellcome Trust Sanger Institute
Next-generation DNA sequencing technologies have greatly increased the throughput of DNA sequencing and have drastically reduced the cost. In spite of this, however, it is still not feasible to perform whole genome sequencing of large numbers of samples, and so it is desirable to be able to isolate specific regions of interest prior to sequencing. Several approaches to sequence capture are available to enrich the specific genomic fragments for targeted resequencing. Here we present an evaluation of the Agilent SureSelect™ Target Enrichment System, where specific regions of a genome are selected by hybridization to biotinylated RNA-oligonucleotides in solution, in combination with the Illumina Genome Analyzer sequencing platform.

Utilizing Agilent’s SureSelect Technology for Resequencing Projects on the Illumina GA Platform
Ryan Tewhey, Division of Biological Sciences, University of California, San Diego Scripps Genomic Medicine, The Scripps Research Institute
While next generation sequencing platforms have greatly increased our ability to analyze millions of bases, the need still exists to enrich for sequences of interest. We have applied Agilent’s SureSelect enrichment technology, a solution based hybridization method, to both DNA isolated from cell lines and clinical blood samples. We have evaluated the technology’s ability to both design probes and subsequently uniformly capture over 3.8 Mb of targeted sequence. The ultimate goal of individual sequencing is the discovery of variants, therefore we have also evaluated the ability of the SureSelect technology to efficiently capture both alleles in comparison to known genotypes.

Sponsored by
Isilon Systems12:45 Luncheon PresentationThe Next-Next-Generation of
Bioinformatics -Maximizing the Value of Unprecedented Data Growth

Ram Appalaraju, Ph.D., Vice President of Product Marketing and Development, Isilon Systems

The data growth and performance needs of DNA sequencing are changing rapidly and unexpectedly, driven by a recent flurry of next-generation technologies. With research possibilities veritably limitless, a critical challenge remains – where to store this deluge of data and how best to maximize its potential for creating future biomedical breakthroughs. The solution? Scale-out, file-based storage architectures have emerged as an ideal solution for powering bioinformatics research, delivering the performance, scalability and ease of use necessary to allow researchers to focus on their science and not their storage. This session will focus on: Advances in DNA sequencing and other bioinformatics-related technologies driving unprecedented data growth; The implications of this data growth for researchers and scientists, as well as its potential to transform modern medicine; Scale-out, file-based storage architectures as the most compelling solution for maximizing the potential of bioinformatics data.

CASE STUDIES

2:00 Chairperson’s Remarks

Kevin Davies, Ph.D., Editor-in-Chief, BioIT World

2:05 Case Study One: Metagenomics as a New Challenge for Next-Generation Sequencing Technologies

Ludmilla Chistoserdova, Research Scientist, Chemical Engineering, University of Washington
Alla Lapidus, Ph.D., Group Leader, Joint Genome Research Institute

Sequencing DNA of entire microbial communities, known as metagenomics, along with the downstream meta-approaches, metatranscriptomics and metaproteomics, present incredible opportunities for analyzing the genomes of uncultivated microbes. The two major challenges in metagenomics, compared to analyzing individual genomes, are the significantly increased complexity of the DNA complement and unequal abundance of DNA of different organisms. Thus the success of metagenomics will rely on the significantly increased sequence coverage that next-generation sequencing technologies offer, but will ultimately depend on the availability of tools allowing reliable assembly, organismal binning and annotation. We use a highly complex microbial community inhabiting Lake Washington sediment, as a model, to test the feasibility of the currently available sequencing and analysis tools.

2:50 Case Study Two: Whole Transcriptome Sequencing of Cancer Biopsies for Concurrent Analysis of Expression,
Splicing and Mutation

Trevor Pugh, B.Sc., Genome Sciences Centre, BC Cancer Agency
Ryan Morin, B.Sc., M.Sc., Genome Sciences Centre, BC Cancer Agency

Clinicians routinely obtain tumor biopsies for diagnostic purposes, making them an appealing source of DNA and RNA. The amount of RNA that can be obtained from some biopsies is often too small to enable the construction of libraries for next-generation sequencing. We have tested multiple RNA amplification strategies to enable robust production of RNA-seq libraries from biopsies. The paired reads are aligned to the human reference genome supplemented with a set of exon-exon junction sequences. The number of unambiguous reads deriving from individual genes, exons, and exon-exon junctions are counted, providing a digital measure of gene expression and splicing. This provides an opportunity to compare the expression of genes and their inclusion of individual exons between tumor and normal samples. In addition to gene expression changes, we are also able to identify SNPs and somatic mutations in expressed genes. The ability to confidently identify mutations is affected by individual gene expression, sampling depth, and the amount of “end bias” in a given library. Using this approach, we have identified novel somatic mutations in a lobular breast cancer, as well as candidate mutations in multiple biopsies from lymphomas and lung adenocarcinomas.

3:35 Case Study Three: Novel MicroRNA Gene Prediction in Human Enbryonic Stem Cells and Neural Precursors

Ronald Hart, Ph.D., Professor, Keck Center for Collaborative Neuroscience, Rutgers University

Loyal A. Goff, Ph.D., Postdoctoral Associate, Computer Science and Artificial Intelligence Laboratory, The Broad Institute @ MIT

4:20 Case Study Four: Tools to Analyze Metagenomics: A Cystic Fibrosis Case Study

Rob Edwards, Ph.D., Department of Biology, San Diego State University
Dana Hall, Ph.D., Department of Biology, San Diego State University

5:05 Close of Meeting

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http://www.chidb.com/2008/seq/day2.asp

Wednesday, April 23

7:30-8:15am Conference Registration and Morning Coffee

TECH EXPO

Explore available next-generation screening platforms as presented by sequencing leaders. An unparalleled opportunity to compare and contrast these next-generation sequencing platforms to best suit your research needs.

8:15 Chairperson’s Opening Remarks
Sponsored Seminars hosted by:
8:30 True Single Molecule Sequencing: Current Research and Our Path to the $1000 Genome

Patrice M. Milos, Ph.D., Vice President and Chief Scientific Officer, Helicos
Helicos has developed a novel genetic analysis platform to efficiently and accurately determine the direct sequence of individual DNA molecules. Simplicity in sample preparation, development of novel surfaces, chemistry to enable incorporation of single nucleotides into DNA strands and finally the visualization of fluorophore addition to monitor real-time sequencing by synthesis has been achieved. This platform will provide the opportunity for researchers to interrogate the genome on a new scale and provides a path to sequence individual genomes at a cost to make integration of genome knowledge and healthcare possible. To demonstrate the current power of the HelicosTM platform we will discuss the use of the True Single Molecule Sequencing Technology (tSMSTM) for candidate gene resequencing, RNA measurements and miRNA analyses.
9:15 Illuminating the Genome

Abizar Lakdawalla, Ph.D., Senior Product Manager, Sequencing Applications, Illumina
The Illumina Genome Analyzer, based on the Solexa massively parallel sequencing-by-synthesis technology, is being used for a broad set of functional genomics applications including chromosomal re-arrangements, to single nucleotide variations, variation in DNA methylation, whole transcriptome analysis, small RNA analysis, digital gene expression, DNA-protein, and DNA-RNA interaction analysis. Details on the current state of the technology as well as a summary of chromosomal resequencing studies, whole genome epigenetic changes, tissue-specific mRNA splice variatns and 5’-UTRs, microRNAs and DNA-protein interactions studies will be presented.
10:00 454/Roche

10:45 Networking Coffee Break, Poster and Exhibit Viewing

11:30 Interactive Panel Discussion: The $1000 Genome Threshold
Moderator: Kevin Davies, Ph.D., Editor-in-Chief, BioIT World
The past 30 months have witnessed remarkable advances in next-generation sequencing throughput, accuracy, and results. More than 200 instruments have now been deployed in organizations around the world, and the range of questions and applications being addressed continues to astound. Together with rapid advances in consumer genomics, we are fast approaching the Holy Grail of the $1000 human genome sequence. This panel discussion will review the most recent technical advances and scientific applications of next-generation sequencing, as well as consider the practical, medical and ethical issues surrounding personalized genomics.
12:30pm Luncheon Technology Workshop hosted by

Your Sequencing Machine Just Completed its Run. Now what?
Powerful “information throughput” enabling timely upstream analysis
Ron Ranauro, President and CEO, GenomeQuest, Inc.
With next-generation sequencing machines churning out data at a rate of 200 million to 1 billion bases per run, researchers are left with the daunting task of making sense of the data. In this workshop we will discuss a breakthrough technology that delivers compute power, algorithms, data management, and results analysis to manage and mine huge volumes of data, directly to your desktop.

DATA ANALYSIS – WHAT DOES IT TAKE?

2:00 Chairperson’s Remarks

Kick-Off Keynote Presentation
2:10 Biomedical Analysis and Applications of Large-Scale DNA Sequence Data
Nicholas J. Schork, Ph.D., Director of Research, Scripps Genomic Medicine and Professor, Molecular and Experimental Medicine, The Scripps Research Institute
This talk will focus on the analysis and potential applications of data generated by next-generation sequencing technologies, with a focus on two main themes: the analysis of individual DNA sequences in human association studies (including cancer and host-pathogen interaction studies) and an assessment of the unique nature of the human diplome.

3:00 SolexaTools: An Open Source Sequencing Framework
Brian O’Connor, Ph.D., Post-Doctorate, Human Genetics, University of California – Los Angeles
The SolexaTools project was started to meet the computation infrastructure needs of scientists using the Solexa massively parallel sequencing platform. This open source project provides two key features for the community. First, a LIMS system for tracking sequencing runs (SolexaLIMS) and, second, a pipeline framework for organizing the analysis of data (SolexaPipeline). These tools allow scientists to automate most of the processing and report generation required by Solexa data. The software is freely available and has been written to allow for customization and community driven development. Both are important given the inherent flexibility of this new sequencing technology.

3:30 Genomics of System-Specific Model Species
Matthew Hudson, Ph.D., Assistant Professor, Crop Sciences, University of Illinois
Next-generation sequencing can be used to produce low-cost genomic resources for “system specific” eukaryotic models: plants and animals which are specifically suited to a particular scientific question. While complete genome sequencing using short reads alone is not yet feasible for higher eukaryotes, current technology can provide an aid to whole-genome sequencing for smaller genomes, and a route to gene-space sequencing of larger genomes. New and adapted sequence analysis methods are necessary to facilitate these genomics projects. Results will be presented on data analysis methods and biological insights from partial genome and transcriptome sequencing using short-read technology.

4:00 Networking Refreshment Break, Poster and Exhibit Viewing

NEXT-NEXT GENERATION SEQUENCING

4:30 SMRT (Single Molecule Real-Time) DNA Sequencing - a Transformative Method for High-Throughput DNA Sequencing
Stephen Turner, Ph.D., Chief Scientific Officer, Research and Development, Pacific Biosciences
SMRT (Single Molecule Real-Time) DNA sequencing is a novel, high-throughput method for sequencing DNA. Though the majority of DNA sequence data collected to-date has been acquired through the use of DNA polymerase enzymes, the methods used squander the inherent power of the enzyme as a sequencing engine. Viewed as a sequencing engine, DNA polymerases can ‘read’ up to 1000 bases per second per molecule, do so over DNA lengths of 100,000 bases or more, replicate with high fidelity and consume only one molecule per base ‘sequenced.’ To harness this power, Pacific Biosciences has developed a method of eavesdropping on template-directed synthesis by DNA polymerase in real-time. We show proof-of-concept data that indicates this will be a high-throughput sequencing technology with long readlengths, limited ultimately by the processivity of the enzyme and fast cycle times dictated by the incorporation rate of the enzyme.

4:55 Base Pair Discrimination via Transmission Electron Microscope
William Glover, President, R&D, ZS Genetics
ZS Genetics (“ZSG”) is developing a Third Generation Sequencing platform to read DNA base pairs directly with a specialized Transmission Electron Microscope. Nucleotides modified with single heavy atoms provide a signal sufficient for discrimination by a sub-angstrom-resolution TEM. dsDNA, initially in the 8 kb to 12 kb range, will be linearized on a thin-membrane substrate. Both strands will be labeled and independently resolved, providing built-in error checking. Speeds for ZSG’s Beta system at the end of 2008 will be comparable to next-generation sequencing technologies and increase thereafter via automation and customization of TEM components.

5:20 Speaker to be Determined

5:45 Networking Reception, Poster and Exhibit Viewing

6:45 End of Day