NGS Maser 2013/10/17
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
( CLST) ( ) Maser http://www.cell-innovation.org/
解析全体像のイメージ De novo Genome Sequencing バイオインフォ マティシャン Genome Resequencing RNA-seq ChIP-seq Bisulfite-seq プログラミング スキルや ツールマニュアル バイオDBの 配列データ や論文のMethod ノウハウ This image is modified from Nature Methods 6, S2 - S5 (2009) Photo is from morguefile http://www.morguefile.com/archive/display/187379 5
解析全体像のイメージ De novo Genome Sequencing Genome Resequencing RNA-seq ChIP-seq Bisulfite-seq 配列データ This image is modified from Nature Methods 6, S2 - S5 (2009) Photo is from morguefile http://www.morguefile.com/archive/display/187379 6
: :
(1) (SRA) [](SRR) 2013/10 http://cell-innovation.nig.ac.jp/public/contents/ sra_stat.html
[] x 10000 30 25 20 15 10 5 0 WXS WGS (2) AMPLICON RNA-Seq ChIP-Seq CLONE Bisulfite-Seq DNase-Hyperse EST / FL-cDNA MeDIP-Seq / (SRA) ( )[](SRR) ( )[%] () (NGS) (): 2013/10 80% 60% 40% 20% 100% 0% http://cell-innovation.nig.ac.jp/cgi-bin/pub_stat/pub_stat31.cgi
[] x 10000 30 25 20 15 10 5 0 WXS WGS AMPLICON RNA-Seq ChIP-Seq CLONE Bisulfite-Seq DNase-Hyperse EST / FL-cDNA MeDIP-Seq / RNA-Seq BS-seq ChIP-seq Genome Resequencing 300 CAGE De novo Genome Sequencing Metagenome
RNA-seq + de novo+ ( /) Fusion Bisulfite-seq CAGE Metagenome 16s rrna ChIP-seq ChIP-Seq QC Genome Resequencing SNV, InDel 1000 CNV/ De novo Genome Sequencing
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
(1) Maser () () Push Push http://cell-innovation.nig.ac.jp/wiki2/tiki-index.php?page=mam0000001
(2) Maser () () http://cell-innovation.nig.ac.jp/wiki2/tiki-index.php?page=mam0000002
(3) Maser () () http://cell-innovation.nig.ac.jp/wiki2/tiki-index.php?page=mam0000002
(4) Maser () () http://cell-innovation.nig.ac.jp/wiki2/tiki-index.php?page=mam0000003
(5) Maser () () http://cell-innovation.nig.ac.jp/wiki2/tiki-index.php?page=mam0000003
(6) Maser () () http://cell-innovation.nig.ac.jp/wiki2/tiki-index.php?page=mam0000003
(7) Maser () () http://cell-innovation.nig.ac.jp/wiki2/tiki-index.php?page=mam0000003
( ) Data from ENCODE project MCF-7_cell_longPolyA(SRX084666), GM12878_cell_longPolyA(SRX082565), K562_cell_longPolyA(SRX084683)
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
RNA-Seq RNA-Seq 2 () ( )
RNA-Seq De novo Trinity-bowtie- express
UCSC Genome Browser Genome Explorer Genome Explorer RNA-seq Data is from Nature. 2011 Mar 3;471(7336):68-73 ADS-iPSC(SRR094759) ADS(SRR094669) Sample A Data is from Nature. 2011 Mar 3;471(7336):68-73 ADS-iPSC(SRR094759) ADS(SRR094669) () Sample B () Sample A () () Sample B () ()
A (Fastq) TopHat-Cufflinks (TopHat) A (BAM) (Cufflinks) A (GTF) B (Fastq) (TopHat) B (BAM) (Cufflinks) B (GTF) A (GTF) (Cuffdiff) (cuffdiff output) B (GTF) (Cuffmerge) (GTF) Nat Protoc. 2012 Mar 1;7(3):562-78
TopHat-Cufflinks (1) 8 SampleA Splicing junction SampleA 2 SampleB Data is from Nature. 2011 Mar 3;471(7336):68-73 ADS-iPSC(SRR094759) ADS(SRR094669) SampleB SambleA B 3 9
Cuffdiff Cuffdiff Data is from Nature. 2011 Mar 3;471(7336):68-73 ADS-iPSC(SRR094759) ADS(SRR094669) Cuffdiff Isoform
Trinity-Bowtie-eXpress (Fasta) (Blast) (tsv) A (Fastq) (Bowtie) A (BAM) (express) B (Fastq) (Bowtie) B (BAM) A B A B
Trinity-Bowtie-eXpress Revigo GO Data from Array Express http://www.ebi.ac.uk/arrayexpress/ ERR030872 HCT20152 thyroid ERR030885 HCT20142 kidney ERR030875 HCT20149 leukocyte ERR030874 HCT20150 ovary ERR030886 HCT20143 heart + Contig ID A B GO
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
ChIP-Seq DNA (IP) DNA Input DNA IP DNA IP
ChIP ChIP QC ENCODE QC (Fastq) (Bowtie) (BAM) ChIP-QC (phantompeakqualtools) (MACS) (html) (BED) (html)
(Fastq) ChIP (Bowtie) (BAM) (MACS) (BED) (html) IP IP This image is modified from the Nat Rev Genet. 2009 Oct;10(10):669-80 Data of Genome Browser view on the right figures ChIP-Seq data from ENCODE project FOXA1_GSM1010826(exp=SL2666,input=SL2665), GSM1010725(input=SL2665) Input Input MACS2
(Fastq) (Bowtie) (BAM) (MACS) (BED) (html) (GADEM) DB (JASPAR) (MotIV)
ChIP-Seq Bowtie MACS Motif finding (Fastq) (Fastq) (Csfasta +qual) Bowtie1/2 BWA TMAP Bowtie (colorspace) (BAM) MACS PeakSeq SISSRs ZINBA SICER (BED) Motif finding
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
BS-seq(Bisulfite-seq) (Cm) Bisulfite() (C) (T) DNA m m m m Bisulfite m m m m 75% 25%
A:iPS(ADS-iPSC) B: (ADS) CpG (GenomeExplorer) 1 A B G C Data is from Nature. 2011 Mar 3;471(7336):68-73 ADS(SRX026833) ADS-iPSC(SRX026835) http://goo.gl/vm3zv1
Bisulfite-Seq CG/CHG/CHH (50x) PBATBMap BMap A (Fastq) B (Fastq) (BMap) or (Bismark) (BMap) or (Bismark) A () B () ( ) ( ) A B () i.e)cpg,, bin, () (FET)
CpGID
GenomeExplorer C CG A B
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
Genome Resequencing SNV, INDEL (CNV) (SV) Nature Genetics 43, 491-498 (2011) doi:10.1038/ng.806
Genome Resequencing BWA, GATK and snpeff + GE (Fastq) (BWA) (BAM) (GATK) (VCF) (snpeff + ) (TSV) SNV, INDEL 1000 PolyPhen2, PROVEAN
BWA, GATK and snpeff + GE
BWA, GATK and snpeff + GE (Fastq) (BWA) (BAM) (GATK) (VCF) (snpeff + ) (TSV) b / ID2 c / ID2 d / ID4 e / ID4 f / ID5 g / ID5 h / ID5 i / ID5 k / ID6 l / ID6 a / ID1 ID4 / ID3 j / ID6
(Fastq) (BWA) (BAM) (GATK) (VCF) (snpeff + ) (TSV) (Ins.) (Del.) (Ins.) (Del.)
(Fastq) (BWA) (BAM) (GATK) (VCF) (snpeff + ) (TSV) 1000 (%) PROVEAN (deleterious = ) PolyPhen2 (damaging = ) 0/0 0/1 1/1
(Fastq) (BWA) (BAM) (GATK) (VCF) (snpeff + ) (TXT) a / ID1 b / ID2 c / ID2 m / ID3 d / ID4 e / ID4 f / ID5 g / ID5 h / ID5 i / ID5 j / ID6 k / ID6 l / ID6
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
De novo Genome Sequencing ncrna
De novo Genome Sequencing annotation after assembling (Fastq) (Fasta) (Blast) (TSV) (JPEG) Illumina SOAPdenovo, Ray 454, IonPGM Newbler ( ) PacBio Sprai 100Mbase
annotation after assembling (Fastq) (Fasta) (Blast) (TSV) (JPEG)
K-mer (Fastq) (PNG) (Fasta) (ERX026224) (Blast) (TSV) (JPEG) K-mer 10 1,000 100,000 10,000,000 800 10 50 100 500 1000 5000 10000 K-mer
(Fastq) (Fasta) (Blast) (TSV) (JPEG) Augustus Uniprot, NCBI NT Blast ( GC% ) Uniprot GO NCBI NT ()
(Fastq) (SRX026594) GC% (Fasta) (Blast) (TSV) (JPEG) () GC log10( )
PSMC (Fastq) (Fasta) (Blast) (TSV) (JPEG) ( ) Effective population size (x10 4 ) 1.8 1.6 1.4 1.2 1 0.8 0.6 0.4 0.2 0 10 4 10 5 10 6 10 7 1~6 Heng Li, et al. Nature 475, 493 496 (28 July 2011)
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
Metagenome DNA NGS 16S, 18S rrna Nature. 2012 Sep 13;489(7415):250-6.
Metagenome blastn for NT database (Fastq) (Blast) Blast (TXT) (Megan) (TSV) 16S rdna SILVA Whole metagenome NCBI NT MEGAN (MEtaGenome ANalyzer) Daniel Huson
Metagenome (Fastq) (Blast) Blast (TXT) (Megan) Megan (TSV) SRR061688 SRR061718 SRR061704
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
CAGE 5 (TSS) mrna G 5
CAGE nant-icage IDR paraclu ver3.1 (Fastq) (BAM) (HTML) TSS (HTML) FANTOMCAGE IlluminaCAGE
(Fastq) ( ) ( ) (BAM) (HTML) TSS (HTML) A549 Gm12878 ( ) ( ) ( ) ( )
(Fastq) CAGE (BAM) (HTML) TSS (HTML) ()
(Fastq) (BAM) (HTML) TSS (HTML)
(Fastq) (BAM) (HTML) TSS (HTML)
CAGE
Maser RNA-seq Genome Resequencing De novo Genome Sequencing Metagenome ChIP-seq CAGE BS-seq
Maser
NGS