Genome Analyzer. RNA ChIP

Transcription

1 Genome Analyzer 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.

2 Genome Analyzer RNA ChIP 2

3 Genome Analyzer DNA de novo Genome Analyzer DNA RNA Small RNA 3

4 Genome Analyzer 800bp x bp x2 x135m x27 vs ,

5 1 Genome Analyzer 3 5 DNA ( ug) 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 T G C T A C G A T

6 6

7 Sequencing Control Software Pipeline Pipeline Visual Controller CASAVA Genome Studio DNA RNA ChIP 3rd Party Software 7

8 cbot Cluster Generation System: Cluster Station & PC Genome Analyzer 8

9 DNA 100bp DNA 100bp DNA bp 2-5kb bp 2 2-5kb 9

10 GAIIx basecall Gb/ Gb/ >Q30 /Mb 1x 35bp M % >99% >90% 109 GAIIx 2x 35bp M % >99% >90% 102 2x 50bp M % >99% >85% 83 2x 75bp M % >98.5% >80% 62 2x 100bp M % >98% >70% 58 10

11 3Gb x38 5Mb (x825) 7 100bp bp /Mb /Mb mrna 100bp /Mb Small RNA 36bp ,000 /Million ChIP 50bp > 5,400 /Million 11

12 95Gb 12

13 Genome Analyzer 100bp 125bp, 150bp 150bp 250bp 13

14 Genome Analyzer 100 >1 33Gb 100bp GA G Genome Epigenome T i t SNP CNV DNA DNA Transcriptome microrna 14

15 mrna Small RNA de novo DNA ChIP de novo Genome Analyzer 15

16 Genome de novo 10x 20x 30x 16

17 Epigenome MethylC-Seq Bisulphite-Seq Reduced Representation BS- Seq Methyl-Seq* Lister, Ecker 09 17

18 Epigenome ChIP-Seq DNA DNA RNA 18

19 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

20 Genome Analyzer RNA ChIP 20

21 21

22 DNA Nature Oct 14. ES (H1) (IMR90) DNA MethylC-Seq (+ Bisulphite PCR) mrna-seq ChIP-Seq Small RNA 22

23 MethylC-Seq + Bisulfate PCR 23

24 Non-mCG DNA TSS DNA Exon non-mcg DNA? 24

25 Non-mCG DNA Strand-specific RNA-Seq 25

26 DNA- DNA DNA ChIP-Seq 26

27 27 DNA

28 28

29 Nature 2008 Nov 6; 456(7218): BAC 6 MHC 162Kb X 153Mb 3Gb 135Gb, 2x 35 PE, 4,000M 200bp, 2kb 29

30 SNP 400 SNP 74% SNP (db SNP) (HumanHap550) 552,710 SNP 99.57% SNP 99.91% SNP 98.74% % GT>Seq 0.35% Seq>GT 0.05% 0.03% Study Human HapMap 550 SNP 552, % 99.57% 0.43% - GT>Seq 0.35% (1,940) - Seq>GT 0.05% (258) % (183) 30

31 SNP?? All SNP Heterozygous SNP Homozygous SNP 31

32 3.6kb 2kp 2kb 200bp 200bp 32

33 29 kb McCarroll et al. (2006) LOH Tuzun et al. (2005) d e p depth w i n d o w 33

34 Identification of somatically acquired rearrangements in cancer using genome-wide massively parallel paired-end sequencing Nature Genetics,

35 200bp 200bp 81bp 35

36 de novo de novo 36

37 37 200bp

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39 SureSelect Target Enrichment System GA 3.3Mb 3Mb 120mer 55K GA 1 39

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41 Genome Analyzer 100 >1 33Gb 100bp GA G Genome Epigenome T i t SNP CNV DNA DNA Transcriptome microrna 41