Doug Brutlag 2011 Next Generation Sequencing and Human Genome Databases Doug Brutlag Professor Emeritus of Biochemistry & Medicine Stanford University School of Medicine Genomics, Bioinformatics & Medicine

Doug Brutlag 2011 Illumina Solexa Sequencing Technology

Doug Brutlag 2011 Emulsion Based Clonal Amplification Micro-reactors Adapter carrying library DNA Anneal DNA template to capture beads Break micro-reactors Isolate DNA containing beads Single test tube generation of millions of clonally amplified sequencing templates No cloning and colony picking “Water-in-oil” emulsion + PCR Reagents + Emulsion Oil Perform emulsion PCR A B

Doug Brutlag 2011 Pacific Biosciences SMRT Sequencing

Doug Brutlag 2011 Pacific Biosciences Sequencing

Doug Brutlag 2011 Phospholinked Fluorophores

Doug Brutlag 2011 Processive Synthesis

Doug Brutlag 2011 Synthesis of Long Duplex DNA

Doug Brutlag 2011 Highly Parallel Optics System

Doug Brutlag 2011 Circular Templates Gives Redundant Sequencing and Accuracy

Doug Brutlag 2011 Circular Templates Gives Redundant Sequencing and Accuracy

Doug Brutlag 2011 Ion Torrent Sequencing

Doug Brutlag 2011 Ion Torrent Sequencing

Doug Brutlag 2011 Ion Torrent Sequencing

Doug Brutlag 2011 The Human Genome How fast is the cost going down? 2006: $ 50 million 2008: $500, : $50, : $20, : $5, :??? $1,000 Thanks to Serafim Batzoglou

Doug Brutlag 2011 Archon Genomics X-Prize

Doug Brutlag 2011 Archon Genomics X-Prize

Doug Brutlag 2011 Components of a Typical Human GeneGene IntronIntronExonExonExonPromoterTerminator TFBS

Doug Brutlag 2011 Active Genes are Transcribed into RNA PrimaryTranscript Gene IntronIntronExon Exon Exon PromoterTerminator

Doug Brutlag 2011Transcript mRNA Gene IntronIntronExon Exon Exon PromoterTerminator 5’5’5’5’ 3’3’3’3’ Splicing Splicing Transcript Yields Mature mRNA

Doug Brutlag 2011 Mature mRNA contains Coding Region and 5’ and 3’ Untranslated Regions Transcript mRNA Gene IntronIntronExon Exon Exon PromoterTerminator 5’5’5’5’ 3’3’3’3’ Splicing Coding Region 5’UTR 3’UTR 5’UTR 3’UTR

Doug Brutlag 2011 Mature mRNA contains 7-Methyl-Guanosine 5’ Cap and 3’ Poly A Tail Transcript mRNA Gene IntronIntronExon Exon Exon PromoterTerminator Splicing Coding Region 5’UTR 3’UTR 5’UTR 3’UTR 7-Me-G-Cap 3’ Poly A Tail

Doug Brutlag 2011 ESTs, Full Length cDNA UniGene & RefSeq Databases UniGeneRefSeq Transcript mRNA Gene Intron Exon PromoterTerminator 5’5’ 3’3’ 3’ ESTs 5’ ESTs Full Length cDNA Splicing

Doug Brutlag 2011 ESTs, Full Length cDNA UniGene & RefSeq Databases UniGeneRefSeq Transcript mRNA Gene Intron Exon PromoterTerminator 5’5’ 3’3’ 3’ ESTs 5’ ESTs Full Length cDNA Splicing Proteins 5’ UTR 3’ UTR Protein

Doug Brutlag 2011 GENSCAN Gene Model Hidden Markov models of gene structure

Doug Brutlag 2011 Promoters E Additional Data Genomic DNA Assembled contigs A Mapping uniSTS dbSNP Mouse ESTs Human ESTs C Expression Data Entrez Gene Mouse RefSeq Mouse UniGene Human RefSeq Human Ensembl cDNA Genome Databases nrPRO D Protein Similarity pFAM Motifs GrailEXP B Gene Prediction GenScanFGENESH FGENESH+ GeneMark F Summary Entrez Gene UCSC Browser Ensembl

Doug Brutlag 2011Inclusive Exon Prediction Entrez Gene Loci NR-Pro Genscan GrailEXP FGENESH Entrez Gene UniGene ESTs

Doug Brutlag 2011 Alternative Splicing Generates Distinct Proteins in Different Tissues Transcript mRNA-1 Gene Intron Exon PromoterTerminator 5’5’ 3’3’ Transcript mRNA-2 5’5’3’3’ Alternate Splicing Splicing

Doug Brutlag 2011 Alternative Splicing Detected in EST Libraries

Doug Brutlag 2011 NCBI Genomes

Doug Brutlag 2011 Eukaryote Genome Projects

Doug Brutlag 2011 Canis lupus familiaris Genome iew&list_uids= iew&list_uids=10726

Doug Brutlag 2011 NCBI Entrez Gene

Doug Brutlag 2011 NCBI Entrez Gene: Human Opsin

Doug Brutlag 2011 Entrez Gene: Human Opsin OPN1MW

Doug Brutlag 2011 Entrez Gene: Human Opsin OPN1MW

Doug Brutlag 2011 MapViewer: Human Opsin OPN1MW

Doug Brutlag 2011 Evidence Viewer for OPN1MW lid=2652&from= &to= lid=2652&from= &to=

Doug Brutlag 2011 OMIM Home Page

Doug Brutlag 2011 Colorblindness in OMIM

Doug Brutlag 2011 Colorblindness in OMIM

Doug Brutlag 2011 Human Genome Resources

Doug Brutlag 2011 RefSeq

Doug Brutlag 2011 RefSeq Gene

Doug Brutlag 2011 NCBI UniGene

Doug Brutlag 2011 NCBI Homologene Database

Doug Brutlag 2011 Comparative Genomics

Doug Brutlag 2011 Ensembl Home Page

Doug Brutlag 2011 EBI Genomes Home Page

Doug Brutlag 2011 Ensembl Human Genome

Doug Brutlag 2011 Ensembl Human Opsin Search x=;q=opsin x=;q=opsin

Doug Brutlag 2011 Ensembl Human Opsin Genes x=;q=opsin x=;q=opsin

Doug Brutlag 2011 Ensembl Human OPN1MW Gene ;r=X: ;r=X:

Doug Brutlag 2011 Ensembl Opsin OPN1MW Gene Location #r=X: #r=X:

Doug Brutlag 2011 Ensembl OPN1MW Transcripts #r=X: #r=X:

Doug Brutlag 2011 Ensembl OPN1MW Opsin Protein ;r=X: ;t=ENST ;r=X: ;t=ENST

Doug Brutlag 2011 Ensembl Tutorials

Doug Brutlag 2011 UCSC Genome Home Page

Doug Brutlag 2011 UCSC Genome Browser

Doug Brutlag 2011 UCSC Genome Browser

Doug Brutlag 2011 UCSC Genome Browser &hgsid= &knownGene=pack&hgFind.matches=uc004fkd.2, &hgsid= &knownGene=pack&hgFind.matches=uc004fkd.2,

Doug Brutlag 2011 UCSC Genome Browser &hgsid= &knownGene=pack&hgFind.matches=uc004fkd.2, &hgsid= &knownGene=pack&hgFind.matches=uc004fkd.2,

Doug Brutlag 2011 UCSC Proteome Browser

Doug Brutlag 2011 UCSC Proteome Browser

Doug Brutlag 2011 UCSC Help File