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Quality of assemblies—mouse

Explore N50 contig length, sequence coverage, assisted assembly history, and future directions in sequencing technology and applications.

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Quality of assemblies—mouse

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  1. Quality of assemblies—mouse 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 50th percentile. 7.7X sequence coverage

  2. Quality of assemblies—dog 7.5X sequence coverage

  3. Quality of assemblies—chimp 3.6X sequence Coverage Assisted Assembly

  4. History of WGA 1997 • 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 Let’s sequence the human genome with the shotgun strategy That is impossible, and a bad idea anyway Phil Green Gene Myers

  5. Some new sequencing technologies

  6. Molecular Inversion Probes

  7. Illumina Genotype Arrays

  8. Single Molecule Array for Genotyping—Solexa

  9. Nanopore Sequencing http://www.mcb.harvard.edu/branton/index.htm

  10. Pyrosequencing on a chip • Mostafa Ronaghi, Stanford Genome Technologies Center • 454 Life Sciences

  11. Polony Sequencing

  12. Short read sequencing protocol • Random, high-coverage clone library (CovG = 7 – 10x) • Low-coverage of clone by reads (CovR = 1 – 2x)

  13. Short read sequencing protocol

  14. Ordering clones into clone contigs

  15. Contig assembly

  16. Contig assembly

  17. Assembly quality Read length = 200 bp, Error rate = 1%, Net coverage = 20.0x

  18. Some future directions for sequencing • Personalized genome sequencing • Find your ~3,000,000 single nucleotide polymorphisms (SNPs) • Find your rearrangements • Goals: • Link genome with phenotype • Provide personalized diet and medicine • (???) designer babies, big-brother insurance companies • Timeline: • Inexpensive sequencing: 2010-2015 • Genotype–phenotype association: 2010-??? • Personalized drugs: 2015-???

  19. Some future directions for sequencing 2. Environmental sequencing • Find your flora: organisms living in your body • External organs: skin, mucous membranes • Gut, mouth, etc. • Normal flora: >200 species, >trillions of individuals • Flora–disease, flora–non-optimal health associations • Timeline: • Inexpensive research sequencing: today • Research & associations within next 10 years • Personalized sequencing 2015+ • Find diversity of organisms living in different environments • Hard to isolate • Assembly of all organisms at once

  20. Some future directions for sequencing • Organism sequencing • Sequence a large fraction of all organisms • Deduce ancestors • Reconstruct ancestral genomes • Synthesize ancestral genomes • Clone—Jurassic park! • Study evolution of function • Find functional elements within a genome • How those evolved in different organisms • Find how modules/machines composed of many genes evolved

  21. Multiple Sequence Alignment

  22. Genome Evolution – Macro Events • Inversions • Deletions • Duplications

  23. Synteny maps Comparison of human and mouse

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