Accessing and visualizing genomics data Jim Noonan GENE 760

A working definition of genomics The global study of how biological information is encoded in genome sequence Genes Regulatory sequences Genetic variation How this information is read out to produce distinct biological outcomes Gene expression and regulation Cellular identity, differentiation and development Phenotypic variation among individuals and species

Genomes are vast information repositories Human 3 Gb kb = 1000 bp Mb = 1x106 bp Gb = 1x109 bp Tb = 1x1012 bp Pb = 1x1015 bp 1 Gb 10 Gb 100 Gb

Sequencing the reference human genome (1990-present; ‘finished’ 2003) Industrialization of Sanger sequencing, library construction, sample preparation, analysis, etc. $3 billion total cost 1 Gb/month at largest centers (2005) YCGA = 9.6 Tb per month (2011)

Reference genomes

Genome assembly and annotation 3 Gb >>109 sequencing reads 36 bp - 1 kb

Genome assembly Scaffold_0: 12,865,123 – 12,965-110 Assembly quality criteria: Accuracy: number of errors (Human << 1/100,000 bp) Contiguity: number of gaps (Human: est. 357) Generate reads Find overlapping reads Assemble reads into contigs contig Coverage: Average number of reads representing a particular position in the assembly Human, Mouse, Rat: > 20x Chimpanzee: ~6x Squirrel: ~2x Join contigs into scaffolds scaffold mate pair Scaffold_0: 12,865,123 – 12,965-110 Join scaffolds into “finished” sequence anchored on chromosomes AGTTGTATTATTAGAAACTGAGGGCTAAAAACTGTGCACATACACAGACACACATATTATTTTAATATAGATTTTCAATAATTGGTCTAGGATAAGGATAATATACAG Chr5: 133,876,119 – 134,876,119 7

ATATCATGCTTGTCATTCGTTTATCAGAGGCCAAATGTTTTTCTTTGTAAACGTGTGTAAAACATTCTCAGAATTTTAAACAATAACAAATCAGGGCTGAATGTGGCCAACATGCAAAGAGGAAATCTCCCATCTGTCCAAATCAAACAGTTGTATTATTAGAAACTGAGGGCTAAAAACTGTGCACATACACAGACACACATATTATTTTAATATAGATTTTCAATAATTGGTCTAGGATAAGGATAATATACAGAGAACATGCCAAAAGTTTAAGCAAGAAGAAAACAAAGACTGTTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTAGAAAGACAATGAAACAGAGCCATGTGACCAATGAGAGAGATGAGGGTGGCAGCAGCCTGTTTTAGATAAGGTACCTGATTGGTGGGATTGGAAGACCTCTCTGAGATTAGTGTCTTCAGATATGCCTTAATGATATGAAAGAACCATTCATGGGAAGGCCTAGCATTAAAAACCGTCTAGGCAGAATGAGCAGCAAGTGCAAGGGTCCTGGATAGGAATGAGCTGGATATACTCAAGGAAGAAAGAGAAACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTATCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCAACATGCAAAGAGGAAATCTCCATATCATGCTTGTCATTCGTTTATCAGAGGCCAAATGTTTTTCTTTGTAAACGTGTGTAAAACATTCTCAGAATTTTAAACAATAACAAATCAGGGCTGAATGTGGCCAACATGCAAAGAGGAAATCTCCCATCTGTCCAAATCAAACAGTTGTATTATTAGAAACTGAGGGCTAAAAACTGTGCACATACACAGACACACATATTATTTTAATATAGATTTTCAATAATTGGTCTAGGATAAGGATAATATACAGAGAACATGCCAAAAGTTTAAGCAAGAAGAAAACAAAGACTGTTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTAGAAAGACAATGAAACAGAGCCATGTGACCAATGAGAGAGATGAGGGTGGCAGCAGCCTGTTTTAGATAAGGTACCTGATTGGTGGGATTGGAAGACCTCTCTGAGATTAGTGTCTTCAGATATGCCTTAATGATATGAAAGAACCATTCATGGGAAGGCCTAGCATTAAAAACCGTCTAGGCAGAATGAGCAGCAAGTGCAAGGGTCCTGGATAGGAATGAGCTGGATATACTCAAGGAAGAAAGAGAAACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTATCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCAACATGCAAAGAGGAAATCTCCATATCATGCTTGTCATTCGTTTATCAGAGGCCAAATGTTTTTCTTTGTAAACGTGTGTAAAACATTCTCAGAATTTTAAACAATAACAAATCAGGGCTGAATGTGGCCAACATGCAAAGAGGAAATCTCCCATCTGTCCAAATCAAACAGTTGTATTATTAGAAACTGAGGGCTAAAAACTGTGCACATACACAGACACACATATTATTTTAATATAGATTTTCAATAATTGGTCTAGGATAAGGATAATATACAGAGAACATGCCAAAAGTTTAAGCAAGAAGAAAACAAAGACTGTTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTAGAAAGACAATGAAACAGAGCCATGTGACCAATGAGAGAGATGAGGGTGGCAGCAGCCTGTTTTAGATAAGGTACCTGATTGGTGGGATTGGAAGACCTCTCTGAGATTAGTGTCTTCAGATATGCCTTAATGATATGAAAGAACCATTCATGGGAAGGCCTAGCATTAAAAACCGTCTAGGCAGAATGAGCAGCAAGTGCAAGGGTCCTGGATAGGAATGAGCTGGATATACTCAAGGAAGAAAGAGAAACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTATCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCAACATGCAAAGAGGAAATCTCCATATCATGCTTGTCATTCGTTTATCAGAGGCCAAATGTTTTTCTTTGTAAACGTGTGTAAAACATTCTCAGAATTTTAAACAATAACAAATCAGGGCTGAATGTGGCCAACATGCAAAGAGGAAATCTCCCATCTGTCCAAATCAAACAGTTGTATTATTAGAAACTGAGGGCTAAAAACTGTGCACATACACAGACACACATATTATTTTAATATAGATTTTCAATAATTGGTCTAGGATAAGGATAATATACAGAGAACATGCCAAAAGTTTAAGCAAGAAGAAAACAAAGACTGTTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTAGAAAGACAATGAAACAGAGCCATGTGACCAATGAGAGAGATGAGGGTGGCAGCAGCCTGTTTTAGATAAGGTACCTGATTGGTGGGATTGGAAGACCTCTCTGAGATTAGTGTCTTCAGATATGCCTTAATGATATGAAAGAACCATTCATGGGAAGGCCTAGCATTAAAAACCGTCTAGGCAGAATGAGCAGCAAGTGCAAGGGTCCTGGATAGGAATGAGCTGGATATACTCAAGGAAGAAAGAGAAACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTATCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCAACATGCAAAGAGGAAATCTCC

Genome annotation Genes: Genetic variation: Sequence conservation Coding, noncoding, miRNA, etc. Isoforms Expression ~3 billion bp ACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTAGAAAGACAATGAAACAGAGCCATGTGACCAATGAGAGAGATGAGGGTGGCAGCAGCCTGTTTTAGATAAGGTACCTGATTGGTGGGATTGGAAGACCTCTCTGAGATTAGTGTCTTCAGATATGCCTTAATGATATGAAAGAACCATTCATGGGAAGGCCTAGCATTAAAAACCGTCTAGGCAGAATGAGCAGCAAGTGCAAGGGTCCTGGATAGGAATGAGCTGGATATACTCAAGGAAGAAAGAGAAACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACTAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTACTATGGAAAAATGAAAATAGATTTTAAAACATGTTAATTCACGTTACTTTTTGTTAAATTTACTTTTCTTCTTTCACTTCTTACCTGTCAATGTTATTAATATTTTTAGGAACAATAAATCACATTAATTCCTTATCTCATGTGAAATTTCATATTTATGATTGATACCTTTAAATGTCATTTGTTGAAGGAAGATTATTCATTTTTTCATTCAATAAATATTTTTTAGAATAATAAGTCCCAGGCACAAGACCAGTATTATGTTCTAGGCATTGGGGATACCATGTTCACAAGACAGACTATGATTTACAGGATCAGATGTGGACTCTCAAATTCGACTGAGAATAAAACAGACACAAACAAGTAAATAAAGTTAATTTCAAGTTGTAATTGATGCTATCCCAGGCACAAGACCA…. Genetic variation: SNPs and CNVs Sequence conservation Regulatory sequences: Promoters Enhancers Insulators Epigenetics: DNA methylation Chromatin

Density of biological information in the human genome Chr5: 133,876,119 – 134,876,119 Genes Transcription TF binding Histone mods Mouse orthology SNPs Repeats

Annotation depth varies by species Human, Mouse (Fly, Worm, Yeast): Chromosome assemblies Dense gene and regulatory maps, variation, etc. Other models (Dog, Chicken, Zebrafish): Chromosome assemblies Partial gene maps; variation; little regulatory data Low coverage vertebrate genomes: Scaffold assemblies Few annotated genes Used for comparative purposes

Portals to access and interpret genomes UCSC Genome Browser (genome.ucsc.edu): Visualization, data recovery, simple analysis (also http://genome-preview.ucsc.edu/) ENSEMBL (ensembl.org): Visualization, data recovery, simple analysis Integrative Genomics Viewer (broadinstitute.orgsoftware/igv/): Local genome viewer (visualize local and remote data) Galaxy (main.g2.bx.psu.edu): Complex data analysis and workflows

UCSC Genome Browser genome.ucsc.edu Wiki Page: genomewiki.ucsc.edu

Read the User Guide

Human genome main page (Feb 2009 assembly) There are multiple assemblies for many genomes! Different genome assemblies have different coordinate systems and may have different annotations: chr2:236,438,403-236,438,948 in March 2006 (hg18) is chr2:236,773,664-236,774,209 in Feb 2009 (hg19)

Categories of data: displayed as tracks Genome Viewer Categories of data: displayed as tracks Discrete intervals (genes) or continuous (transcription) Category: Genes and Gene Prediction Hyperlinks and tabs for individual tracks Go to track description page Hide or show data in genome viewer Some tracks include multiple datasets (‘subtracks’) Go to track description page to select Different assemblies have different annotations!

Sample Genome Viewer image: PITX1 Base position Gene model (discrete) Transcription (continuous) TF binding SNPs Repeats

Which gene annotation to use?

Gene description page and links to other resources

‘Layered’ tracks: Transcription Display options Subtracks

Integrating different types of annotation data

Integrating different types of annotation data Proximal enhancer Promoter

Common Genome Browser file formats BED format For interval data (e.g., exons) Tab-delimited format: chr start stop identifier BED coordinates are ‘zero-based, half-open’: the start position is 0-based, the end position is 1-based Position coordinates on the browser are 1-based. This leads to confusion if you are not careful. chr16 80372593 80373755 is shown in the browser as chr16:80372594-80373755 BEDTools: utilities for comparing genomic features you will use on your problem sets WIG format For continuous data (e.g., the Transcriptome track mentioned earlier) WIG files are very large! BigWig is an alternative format you will learn about in discussion.

The Table Browser (under Tools) Select datasets Compare datasets Download data

Integrating your own experimental data Proximal enhancer Promoter Mapping binding sites for a transcription factor of interest

Custom tracks and sessions Display and share your own data on the browser Custom tracks can be intersected, etc. in the Table Browser

Track Hubs (under My Data)

Integrating Track Hub data with your own experimental data

Genome Browser utilities: BLAT (under Tools) Rapidly find sequence locations in an assembly DNA sequences >24 bp and 95% identical to target genome

Assembly quality and annotation vary across genomes Assembly not anchored to chromosomes Poor gene annotation Assembly quality metrics Whole-genome alignment to mouse

Genome Browser utilities: LiftOver (under Tools) Convert coordinates from one assembly to another (e.g., hg18 to hg19) Identify orthologous positions between genomes (e.g., human to mouse)

Galaxy main.g2.bx.psu.edu

Wrap-up Problem Set #1: Learn how access and manipulate genomic datasets Next lecture: High-throughput sequencing technologies