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DNA Sequencing Technology and its Applications in Evolution Research

DNA Sequencing Technology and its Applications in Evolution Research. Julie Urban, Ph.D. Assistant Director, Genomics & Microbiology Laboratory NC Museum of Natural Sciences. Planthopper Evolution. 1953. Planthopper Evolution. 15 Feb, 2001. Planthopper Evolution. (2001). X. X.

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DNA Sequencing Technology and its Applications in Evolution Research

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  1. DNA Sequencing Technology and its Applications in Evolution Research Julie Urban, Ph.D.Assistant Director, Genomics & Microbiology LaboratoryNC Museum of Natural Sciences

  2. Planthopper Evolution

  3. 1953 Planthopper Evolution

  4. 15 Feb, 2001

  5. Planthopper Evolution (2001)

  6. X X Planthopper Evolution (2008) (2001)

  7. Today MiSeq ($126,000) Ion Proton ($149,000)human genome for < $5,000, ~1 day (?) Ion Torrent ($90,000)

  8. Planthopper Evolution

  9. Planthopper Evolution

  10. Planthopper Evolution

  11. Capillary sequencing Single molecule methods Sequencing by synthesis

  12. Capillary (Sanger) Sequencing Planthopper Evolution

  13. Capillary (Sanger) Sequencing

  14. Fluorescence Important in (most) Next Gen Sequencing

  15. Steps Common to all Next Generation Sequencing Systems Planthopper Evolution 1. Make Library: -- fragment DNA into unique fragments-- add custom adaptors (“linkers”) 2. Amplify Library -- make many identical copies (“clones”) of each unique fragment -- copies are made on a solid surface 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment All sequence DNA in “massive parallel” fashion

  16. Roche/454 Sequencing Planthopper Evolution 1. Make Library: -- fragment DNA into unique fragments-- add custom adaptors (“linkers”)

  17. Roche/454 Sequencing Planthopper Evolution 2. Amplify Library -- make many identical copies (“clones”) of each unique fragment -- copies are made on a solid surface

  18. Roche/454 Sequencing Planthopper Evolution 2. Amplify Library -- make many identical copies (“clones”) of each unique fragment -- copies are made on a solid surface

  19. Roche/454 Sequencing Planthopper Evolution 2. Amplify Library -- make many identical copies (“clones”) of each unique fragment -- copies are made on a solid surface

  20. Roche/454 Sequencing Planthopper Evolution 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment

  21. Illumina Sequencing Planthopper Evolution 1. Make Library: -- fragment DNA into unique fragments-- add custom adaptors (“linkers”)

  22. Illumina Sequencing Planthopper Evolution 1. Make Library: -- fragment DNA into unique fragments-- add custom adaptors (“linkers”)

  23. Illumina Sequencing Planthopper Evolution 2. Amplify Library -- make many identical copies (“clones”) of each unique fragment -- copies are made on a solid surface

  24. Illumina Sequencing Planthopper Evolution 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment

  25. Illumina Sequencing Planthopper Evolution 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment

  26. Life Tech/ABI Ion Torrent & Proton Sequencing Planthopper Evolution 1. Make Library: -- fragment DNA into unique fragments-- add custom adaptors (“linkers”)

  27. Life Tech/ABI Ion Torrent & Proton Sequencing Planthopper Evolution 2. Amplify Library -- make many identical copies (“clones”) of each unique fragment -- copies are made on a solid surface

  28. Life Tech/ABI Ion Torrent & Proton Sequencing Planthopper Evolution 2. Amplify Library -- make many identical copies (“clones”) of each unique fragment -- copies are made on a solid surface

  29. Life Tech/ABI Ion Torrent & Proton Sequencing Planthopper Evolution 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment

  30. Life Tech/ABI Ion Torrent & Proton Sequencing Planthopper Evolution 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment

  31. Life Tech/ABI Ion Torrent & Proton Sequencing Planthopper Evolution 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment

  32. Life Tech/ABI Ion Torrent & Proton Sequencing Planthopper Evolution 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment

  33. Life Tech/ABI Ion Torrent & Proton Sequencing Planthopper Evolution 3. Sequencing by Synthesis -- fragments are copied again (sequencing reaction) -- incorporation of each base “fires” a signal -- system detects signal coming from all clones “firing” at once for that unique fragment

  34. Single Molecule Systems: Pacific Biosciences Planthopper Evolution

  35. Single Molecule Systems: Oxford Nanopore Planthopper Evolution

  36. Capillary sequencing Single molecule methods Sequencing by synthesis

  37. Applications of Next Generation Sequencing Planthopper Evolution Metagenomics

  38. Applications of Next Generation Sequencing Planthopper Evolution Microbiome Human Microbiome Project http://commonfund.nih.gov/hmp/

  39. Applications of Next Generation Sequencing Planthopper Evolution Microbiome

  40. Applications of Next Generation Sequencing Planthopper Evolution Bacterial Genome Sequencing Bacteriome = insect organ made of bacteriocytes Planthoppers

  41. Applications of Next Generation Sequencing Planthopper Evolution Ancient (=degraded) DNA Sequencing

  42. Planthopper Evolution

  43. Planthopper Evolution Annual Review of Genomics and Human Genetics, 2008. 9: 387-402. Youtube: Current Topics in Genome Analysis 2012; NHGRI Lecture Series. -- Elaine Mardis -- Jonathan Eisen -- Julie Segre (Microbiome)

  44. Collaborators and Funding Holly Menninger NCSU Rob Dunn NCSU Jason Cryan NCMNS Julie Horvatn NCMNS Greg Pahel NCMNS Megan Ehlers NCSU undergrad Megan Thoemmes NCSU undergrad Dan Fergus NCMNS

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