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DNA Barcoding

DNA Barcoding. Amy Driskell Laboratories of Analytical Biology National Museum of Natural History Smithsonian Institution, Wash. DC. Outline. Barcoding in general Uses & Examples Barcoding Bocas Algae Data, Analysis, etc. What is a DNA barcode?.

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DNA Barcoding

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  1. DNA Barcoding Amy Driskell Laboratories of Analytical Biology National Museum of Natural History Smithsonian Institution, Wash. DC

  2. Outline • Barcoding in general • Uses & Examples • Barcoding Bocas Algae • Data, Analysis, etc.

  3. What is a DNA barcode? • A fingerprint for identification of everything • A short, easily and universally amplifiable, and reasonably variable piece of DNA

  4. RequirementsFor “Barcode” label in GenBank, for “Reference” in BOLD • Sequence is from a vouchered specimen - Re-identify • Voucher meta-information required: GPS coordinates, photographs (in situ, in some cases), collector and identifier data - Re-collect • DNA sequence is high quality (minimum length, minimum coverage, minimum “quality scores” - Compare • Raw data (chromatograms) publicly available - Re-analyze

  5. Organizations • Consortium for the Barcode of Life (CBOL) (www.barcoding.si.edu) • Barcode of Life Database (BOLD) (www.barcodinglife.org) • International Barcode of Life (iBOL) (www.ibolprojec.org) • FishBOL, All Birds Barcoding Initiative, MarBOL, etc. etc.

  6. Barcode “Markers” • Animals: Mitochondrial cytochrome oxidase subunit I (CO1/COX1) ~ 650 bp • BUT, not variable in cnidarians, 16S rDNA historically more informative in many groups (e.g. frogs, some crustaceans). Second marker possible. • Fungi: Nuclear internal transcribed spacer (ITS) • Non-coding gene and difficult to align. But long history in fungal studies, large existing databases, CO1 contains introns.

  7. Barcode “Markers” • Red & Brown Algae: CO1 • Not nearly as “universally” amplifiable as in animals, requires many different primers • Green Plants: Not decided • SI botanists promote rBCL and the trnH-psbB spacer • Others prefer for rBCL and matK • All are chloroplast genes • Green algae? Still not clear

  8. DNA Barcoding is an imperfect science • Evolutionary History: • Hybridization (plastid genomes) • Pseudogenes • Heteroplasmy or multiple copies (nuclear markers) • Lineage sorting, recent speciation • Slow rates of sequence divergence • Practical Difficulties • Lack of universality • Co-amplification • Incomplete sampling • Lack of taxonomic experts

  9. Barcode of Life Database (BOLD)

  10. Today’s Statistics BOLD Total: 570,000 Species: 62,000 “Reference”: 234,000 “Ref” species: 13,774 GenBank “Barcode” keyword: 37,000

  11. Algal Barcodes in BOLD

  12. SI’s Barcoding Philosophy • Collaborate - taxonomic experts, students • Train - students, interns, other researchers • Assist - lab setup, protocol development, collecting • Archive - a “lending library” of high quality, well-vouchered DNAs Explore biodiversity in collaboration with taxonomists and phylogeneticists.

  13. Biologically Interesting Uses Biodiversity Exploration • Discovery of sibling species • Quick assessment of local diversity • “DNA assisted alpha taxonomy” • Ecological & Behavioral Studies • Habitat assessment • Stomach content analysis

  14. L.A.B. Examples • LAB & Bocas taxonomic workshops • CeDAMAR Antarctic deep sea surveys • Caribbean Fish • Central American Frogs • Biocode Moorea

  15. Practical, Fundable Uses • Method of identification, particularly useful to non-experts • Import/export and pest control • Fisheries management • Water and habitat quality assessment • Partial or fragmentary sample analysis • Associating different life stages

  16. L.A.B. Examples • USAF/FAA “Bird Strike” Project • Larval Fish ID • US Food & Drug Agency • US Enviromental Protection Agency

  17. Barcoding Bocas Algae • Collections 2007, 2008, 2009 • ~1000 specimens • Red Algae: ~250 specimens sequenced • Lab protocol and primer development

  18. The Process of Barcoding • Extraction: methods, machines, timing

  19. The Process of Barcoding 2. Amplification - Polymerase Chain Reaction -Primers

  20. The Process of Barcoding 3. Sequencing

  21. The Process of Barcoding • Submission and organization of metadata • Quality control, data processing • Accuracy checking

  22. Nov. 9-13, 2009 Website: dnabarcodes2009.org

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