CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Quality of assemblies—mouse N50 contig length Terminology: N50 contig length If we sort contigs from largest to smallest, and start Covering the genome in that order, N50 is the length Of the contig that just covers the 50 th percentile. 7.7X sequence coverage

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Quality of assemblies—dog 7.5X sequence coverage

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Quality of assemblies—chimp 3.6X sequence Coverage Assisted Assembly

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 History of WGA 1982: -virus, 48,502 bp 1995: h-influenzae, 1 Mbp 2000: fly, 100 Mbp 2001 – present  human (3Gbp), mouse (2.5Gbp), rat *, chicken, dog, chimpanzee, several fungal genomes Gene Myers Let’s sequence the human genome with the shotgun strategy That is impossible, and a bad idea anyway Phil Green 1997

$399 Personal Genome Service $2,500 Health Compass service $985 deCODEme (November 2007) (April 2008) $350,000 Whole-genome sequencing (November 2007) Genetic Information Nondiscrimination Act (May 2008)

Whole-genome sequencing Comparative genomics Genome resequencing Structural variation analysis Polymorphism discovery Metagenomics Environmental sequencing Gene expression profiling Applications Genotyping Population genetics Migration studies Ancestry inference Relationship inference Genetic screening Drug targeting Forensics

CS273a Lecture 9, Aut08, Batzoglou Sequencing applications Demand for more sequencing Sequencing technology improvement Increase in sequencing data output New sequencing applications

CS273a Lecture 9, Aut08, Batzoglou Sequencing technology Sanger sequencing $10.00 $1.00 $0.10 $0.01 Cost per finished bp: Read length:15 – 200 bp500 – 1,000 bp Throughput: “grad-student years”2 ∙ 10 6 bp/day Fred Sanger

CS273a Lecture 9, Aut08, Batzoglou Sequencing technology Sanger sequencing 3 ∙ 10 9 bp 1x coverage 10x coverage 2 ∙ 10 6 bp/day = 40 years × 3 ∙ 10 9 bp 10x coverage × 3 ∙ 10 9 bp × $0.001/bp = $30 million

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Pyrosequencing on a chip Mostafa Ronaghi, Stanford Genome Technologies Center 454 Life Sciences

CS273a Lecture 9, Aut08, Batzoglou Sequencing technology Next-generation sequencing Read length:250 bp Throughput:300 Mb/day Cost: ~ 10,000 bp/$ De novo:yes Genome Sequencer / FLX “short reads”

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Single Molecule Array for Genotyping—Solexa

CS273a Lecture 9, Aut08, Batzoglou Sequencing technology Next-generation sequencing Read length: ~ 35 bp Throughput:300 – 500 Mb/day Cost: ~ 100,000 bp/$ De novo:yes Genome AnalyzerSOLiD Analyzer “microreads”

CS273a Lecture 9, Aut08, Batzoglou Sequencing technology Next-generation sequencing Read length: ~ bp Throughput:3 Gb/day Cost: ~ 3,000,000 bp/$ De novo:yes Genome AnalyzerSOLiD Analyzer reads

CS273a Lecture 9, Aut08, Batzoglou Illumina Projections

CS273a Lecture 9, Aut08, Batzoglou Complete Genomics  $5,000 this summer  Quality?...  1,000 genomes in 2009  20,000 genomes in 2010

CS273a Lecture 9, Aut08, Batzoglou Pacific Biosciences

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall : $10 million 2008: $100, : $10,000 ? $1,000 ??? $100 So, how fast is cost going down?

CS273a Lecture 9, Aut08, Batzoglou Molecular Inversion Probes

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Illumina Genotype Arrays

CS273a Lecture 9, Aut08, Batzoglou Sequencing technology Next-generation sequencing Read length:1 bp Throughput:1 – 2 Mb/day Cost:5,000 bp/$ De novo:no Infinium AssayGeneChip Array genotypes “SNP chips”

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Nanopore Sequencing

CS273a Lecture 9, Aut08, Batzoglou Sequencing technology Next-generation sequencing

CS273a Lecture 9, Aut08, Batzoglou Sequencing technology TechnologyRead length (bp) Throughput (Mb/day) Cost (bp/$) De novo Sanger1, ,000 Solexa / ABI ,000 SNP chip125,000 ApplicationSanger454Solexa/ABI SNP chip Bacterial sequencing$ Mammalian sequencing$$$$$ not likely today Mammalian resequencing$$$$$$ Metagenomics$$$ Genotyping$$$

CS273a Lecture 9, Aut08, Batzoglou Multiple Sequence Alignment

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Evolution at the DNA level …ACGGTGCAGTTACCA… …AC----CAGTCCACCA… Mutation SEQUENCE EDITS REARRANGEMENTS Deletion Inversion Translocation Duplication

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Evolutionary Rates OK X X Still OK? next generation

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008 Orthology, Paralogy, Inparalogs, Outparalogs

CS273a Lecture 9, Aut08, Batzoglou CS273a Lecture 9, Fall 2008