Illumina Custom Capabilities: Amplicons, Enrichment and iselect
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1 Illumina Custom Capabilities: Amplicons, Enrichment and iselect Daniel Peiffer, PhD Sr. Product Manager DNA Sequencing Applications Tuesday, March 22, Illumina, Inc. All rights reserved. Illumina, illuminadx, Solexa, Making Sense Out of Life, Oligator, Sentrix, GoldenGate, GoldenGate Indexing, DASL, BeadArray, Array of Arrays, Infinium, BeadXpress, VeraCode, IntelliHyb, iselect, CSPro, GenomeStudio, Genetic Energy, HiSeq, and HiScan are registered trademarks or trademarks of Illumina, Inc. All other brands and names contained herein are the property of their respective owners.
2 Illumina s Custom Toolset: Your Choice. Leading seq & array technologies coupled with an expert knowledgebase in custom designs Custom Genotyping Arrays Custom Targeted Resequencing Choice of multi-sample arrays offering range of (1 to 1M+) custom variants Leverage proven GoldenGate and Infinium Assays Add on content to previously designed panels Offers high-throughput processing with unique multiplexed sample enrichments Target large amounts of content with TruSeq Custom Enrichment (1 to 10Mb) Multiplex large numbers of samples and targets with TruSeq Custom Amplicon 2
3 TruSeq Sample Prep Solutions Integrated sequencing workflow from sample to analyzed data TruSeq DNA Simple, scalable and cost effective TruSeq RNA Optimized, gel-fee, low input TruSeq Custom & Exome Enrichment Lowest cost and most scalable targeted resequencing TruSeq Small RNA Hi throughput mirna discovery & profiling TruSeq Chemistry Clustering & Sequencing 3
4 Epicentre Nextera Technology for Library Prep Single Tube, Rapid Library Prep SIMPLE, FAST LIBRARY PREP IN LESS THAN 2 HOURS Transposon-mediated library preparation Fragments genomic DNA and ligates adapters in a single step (5mins) PCR to add barcodes 19bp recognition sequence 5 mins Transposase Adapter sequences Low input requirements (50ng) barcode Gels not necessary for range of applications Adey et al Genome Biology (2010) 4
5 Illumina TruSeq Targeted Resequencing A broad suite of customizable tools for discovery or validation experiments Option Amount of sequence Best for Availability TruSeq Exome Enrichment ~62Mb Study of Mendelian disease; casecontrol exome studies, exome-wide linkage Now! TruSeq Custom Enrichment ~1Mb to ~10Mb GWAS follow-up; validation of variants, variant discovery Mid-2011 TruSeq Custom Amplicon ~100kb Amplicon sequencing; highthroughput CE experiments, ultra deep seq, variant disc, screening 2H2011 5
6 Science Magazines Top Breakthroughs of 2010 Published on December 16, 2010 Quantum motion machine Synthetic Biology Neanderthal Genome HIV Prophylaxis Exome Sequencing/Rare Disease Genes Molecular Dynamics Simulations Quantum Simulator Next-Generation Genomics RNA Reprogramming The Return of the Rat 6
7 Success of exome approach is evident in growing publication # Major focus has been on study of Mendelian Disease Dramatic increase in number of exome publications over the past 3 years 2008: : : 66 Huge increase in number of variants 7
8 TruSeq Exome Enrichment Pre-enrichment pooling and comprehensive coverage for the most-cost effective exome MOST COMPREHENSIVE EXOME SOLUTION High coverage uniformity & on-target enrichment Lowest DNA input PLATE-BASED PROCESSING FOR UP TO 96 SAMPLES Simple & scalable workflow PRE-ENRICHMENT POOLING OF UP TO 6 SAMPLES Reduces hands-on time Decreases costs Gel-free protocol coming soon! INTEGRATED WITH TRUSEQ DNA SAMPLE PREP KITS Optimized workflow Internal QC controls 8
9 TruSeq Exome Enrichment Most up-to-date and comprehensive exome available with >340k validated probes CCDS coding exons (31Mb): 97% RefSeq (refgene) coding exons (33Mb): 96% RefSeq (refgene) exons plus (68Mb): 88% Encode/Gencode coding exons* (26Mb): 93% Predicted microrna targets** (9Mb): 78% Only empirically tested probes included Product details: Genes targeted: 20,794 Exons targeted: 201,121 Total region size: 62Mb # of probes: 340,427 Rec library size: bp * Manual V4 ** mirbase 15 9
10 Coming soon! TruSeq Custom Enrichment Kits Premier seq technology coupled with an expert knowledgebase in custom oligo designs TARGET 1 MB 10 MB OF DNA PER SAMPLE Highest enrichment efficiency and coverage uniformity LEVERAGES ILMN S CORE COMPETENCY IN OLIGO PRODUCTION Personalized and easy to use design tool High coverage of targeted regions PRE-ENRICHMENT SAMPLE POOLING Up to 12 samples per enrichment reaction Reduce hands-on time; increase throughput INTEGRATED WITH TRUSEQ DNA SAMPLE PREP KITS Fully optimized workflow Most cost-effective and scalable TRS Solution Early access program has begun! First custom order expected summer 2011! 10
11 Highest Coverage Uniformity High coverage uniformity for 6-plex sample pooling 11
12 High On-Target Enrichment >80% enrichment rates for 6-plex sample pooling Shown are the percentage of reads mapping to target from total reads. Data is from HiSeq /- 150bp includes flanking up- & down-stream regions of target 12
13 TruSeq Custom Enrichment Probes are optimally spaced to efficiently enrich targets 13
14 TruSeq Custom Enrichment - early access High success rates designing against exons 14
15 Coming soon! TruSeq Custom Amplicon Sequencing Highly multiplexed sample and target amplicon resequencing FULLY CUSTOMIZED TARGET PROBES AND CAPTURE Based upon GoldenGate Technology INTERACTIVE PROBE DESIGN AND ORDERING Personalized and easy to use design tool Rapid probe turnaround RAPID & ECONOMICAL Up to 384 amplicons per sample Plate based processing; 96 samples per plate Assay time <8 hours No additional hardware requirements UP TO 10X MORE COST EFFECTIVE THAN CE* *Based upon example study of 96 samples and 384 targets 15
16 TruSeq Amplicon 96-well plate CE TruSeq Custom Amplicon Comparison to CE Sequencing Example: TruSeq Custom Amplicon with 96 Samples x 96 Targets on MiSeq Each well contains DNA + oligos for 1 target 2 rounds of PCR & cleanup 96 plates 3730xl 5 days ~$3/target Each well contains DNA + oligos for 96 targets and indices for sample pooling Assay biochemistry 1 plate MiSeq 1 day <$0.50/target 16
17 Custom Genotyping from Illumina: iselect 2010 Illumina, Inc. All rights reserved. Illumina, illuminadx, Solexa, Making Sense Out of Life, Oligator, Sentrix, GoldenGate, GoldenGate Indexing, DASL, BeadArray, Array of Arrays, Infinium, BeadXpress, VeraCode, IntelliHyb, iselect, CSPro, GenomeStudio, Genetic Energy, HiSeq, and HiScan are registered trademarks or trademarks of Illumina, Inc. All other brands and names contained herein are the property of their respective owners.
18 Applications of Custom Genotyping Genome-wide selection Dog Pine Corn Mouse Potato Genetic prediction SNP discovery Post-GWAS variant confirmation Post-sequencing target validation Targeted disease panels Targeted rare variant panels Chicken Soy Bean Human Atlantic Cod Atlantic Salmon Citrus Lettuce Grape Vine Barley Cotton Rapeseed Spruce False Brome Tomato Sweet Sorghum P. falciparum Cattle Cherry Zebra Finch Zebra Fish Horse Fine mapping Cat Rat Cacao Citru s Swine Sheep And more! Poplar Peach Turkey Rhesus Rice Wheat Honey Bee Rye grass Canola Armadillo 18
19 Illumina Custom Genotyping Driving the Cycle of Discovery and Validation VeraCode GoldenGate Indexing GoldenGate iselecthd 3K-90K iselecthd 90K-250K iselecthd 250K-1M Semi-Custom OEx+ The power of the GoldenGate assay on a platform and at a price accessible to all labs. Enhancement of the GoldenGate assay to deliver the highest throughput at industry leading value. Optimized design of customizable content using the most trusted assay & the highest quality data. Validating genomic discoveries with the most robust, powerful data and flexible content design. Enabling more genomic discoveries with the most robust, powerful data and flexible content design. Enabling more genomic discoveries with the most robust, powerful data and flexible content design. Customizable GWAS array with industry proven quality at an exceptional price Plex Plex 96 3,072 Plex 3,072 1,000, k Plex 19
20 Features of iselect Custom Genotyping Add-On Content Now add markers after initial design is complete Take advantage of new data for variant confirmation, fine mapping and target validation Enabled by internal operational improvements Increased Plexity Former maximum: 200,000 markers New maximum: 1,000,000 markers Enabled by addition of 4- sample chip Illumina s Proven Quality Powered by the proven Infinium assay 2011 Promotional Pricing Improved pricing for projects up to 50,000 samples and 500,000 attempted beadtypes, good for the full year of
21 iselect BeadChip Formats and Supported Marker Densities Multisample arrays offering 3k to 1M attempted beadtypes BeadChip Format 24-sample 12-sample 4-sample Previous plex range 3,072 60,800 60, ,000 n/a New plex range 3,072 90,000 90, , ,001 1M Add-on content range* Up to limit of 90,000 Up to limit of 250,000 Up to limit of 1M *Add-on content range allowed = (maximum attempted beadtypes on BeadChip) MINUS (existing beadtypes in original pool) 21
22 Add-on Content for iselect is now available Beadpool 1 (original) merge Beadpool 2 (add-on) iselect+ HumanOmniExpress+ Beadpool 1+2 (iselect+ or OmniExpress+) Gives the flexibility to add additional content onto existing beadpools (fueled by next gen sequencing, GWAS, exome sequencing, etc) enables fine mapping, target validation, and variant confirmation. Add-on turnaround time is weeks 22
23 Join us for an upcoming webinar this Thursday! 23
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