Genome Analyzer 2009 11 26 2009 Illumina Inc All rights reserved 2009 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, and GenomeStudio are registered trademarks or trademarks of Illumina, Inc. All other brands and names contained herein are the property of their respective owners.
Genome Analyzer RNA ChIP 2
Genome Analyzer DNA de novo Genome Analyzer DNA RNA Small RNA 3
Genome Analyzer 800bp x 384 30 100bp x2 x135m 270 30 570 570 30 13 3000 x27 vs 160 1 52,000 1 9 4
1 Genome Analyzer 3 5 DNA (0.1-1.0 ug) 1. 2. A C T G T C G T A G T G C CT C A A C C C G G A A T T A A A C C C C G C G C G A A T T C C T G G A A T T 5 3. 1 2 3 4 5 6 7 8 9 T G C T A C G A T 4. 5. 5
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Sequencing Control Software Pipeline Pipeline Visual Controller CASAVA Genome Studio DNA RNA ChIP 3rd Party Software 7
cbot Cluster Generation System: Cluster Station & PC 1. 2. 3. 4. 5. Genome Analyzer 8
DNA 100bp DNA 100bp DNA 2 200-500bp 2-5kb 200-400bp 2 2-5kb 9
GAIIx basecall Gb/ Gb/ >Q30 /Mb 1x 35bp 2 138-168M 4.5-6 2.3-3 75-90% >99% >90% 109 GAIIx 2x 35bp 4 276-336M 6.5-11.5 2.4-2.9 75-90% >99% >90% 102 2x 50bp 5 276-336M 13.5-16.5 2.7-3.3 75-90% >99% >85% 83 2x 75bp 7.5 276-336M 20.5-25 2.7-3.3 70-85% >98.5% >80% 62 2x 100bp 9.5 276-336M 27.5-33 2.9-3.3 70% >98% >70% 58 10
3Gb x38 5Mb (x825) 7 100bp 4 1 100bp 1 1 1 1 760 66 /Mb 230 78 /Mb mrna 100bp 1 104 72 /Mb Small RNA 36bp 1 65 5,000 /Million ChIP 50bp 1 80 1.5 > 5,400 /Million 11
95Gb 12
Genome Analyzer 100bp 125bp, 150bp 150bp 250bp 13
Genome Analyzer 100 >1 33Gb 100bp GA 2009 10 400 1 15 2009 95G Genome Epigenome T i t SNP CNV DNA DNA Transcriptome microrna 14
mrna Small RNA de novo DNA ChIP de novo 35 50 75 100 200 Genome Analyzer 15
Genome de novo 10x 20x 30x 16
Epigenome MethylC-Seq Bisulphite-Seq Reduced Representation BS- Seq Methyl-Seq* Lister, Ecker 09 17
Epigenome ChIP-Seq DNA DNA RNA 18
Transcriptome mrna-seq: mrna cdna csnp Directional RNA-Seq Workflow Total RNA Small RNA: RNA sirna Small RNA Small RNA Poly-A Selection mrna Fragmentation RNA fragment clean-up RNA Adapter Ligations Perform RT-PCR Amplification Purify Library 19
Genome Analyzer RNA ChIP 20
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DNA Nature. 2009 Oct 14. ES (H1) (IMR90) DNA MethylC-Seq (+ Bisulphite PCR) mrna-seq ChIP-Seq Small RNA 22
MethylC-Seq + Bisulfate PCR 23
Non-mCG DNA TSS DNA Exon non-mcg DNA? 24
Non-mCG DNA Strand-specific RNA-Seq 25
DNA- DNA DNA ChIP-Seq 26
27 DNA
28
Nature 2008 Nov 6; 456(7218):53-9 3 BAC 6 MHC 162Kb X 153Mb 3Gb 135Gb, 2x 35 PE, 4,000M 200bp, 2kb 29
SNP 400 SNP 74% SNP (db SNP) (HumanHap550) 552,710 SNP 99.57% SNP 99.91% SNP 98.74% 0 0.43% GT>Seq 0.35% Seq>GT 0.05% 0.03% Study Human HapMap 550 SNP 552,710 99.60% 99.57% 0.43% - GT>Seq 0.35% (1,940) - Seq>GT 0.05% (258) - 0.03% (183) 30
SNP?? All SNP Heterozygous SNP Homozygous SNP 31
3.6kb 2kp 2kb 200bp 200bp 32
29 kb McCarroll et al. (2006) LOH Tuzun et al. (2005) 4 5 40 d e p 4 0 3 5 30 3 0 2 5 depth 20 2 0 1 5 10 1 0 0 5 0 0 5 0 0 1 0 0 0 1 5 0 0 2 0 0 0 2 5 0 0 3 0 0 0 3 5 0 0 4 0 0 0 4 5 0 0 5 0 0 0 w i n d o w 33
Identification of somatically acquired rearrangements in cancer using genome-wide massively parallel paired-end sequencing Nature Genetics, 2008 34
200bp 200bp 81bp 35
de novo de novo 36
37 200bp
38
SureSelect Target Enrichment System GA 3.3Mb 3Mb 120mer 55K GA 1 39
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Genome Analyzer 100 >1 33Gb 100bp GA 2009 10 400 1 15 2009 95G Genome Epigenome T i t SNP CNV DNA DNA Transcriptome microrna 41