INTRODUCTION TO DNA BARCODING Let’s Talk Science Fish Market Survey Action Project

1. INTRODUCTION TO DNA BARCODING Let’s Talk Science Fish Market Survey Action Project

2. What is the elementary unit for a physicist? A metre The distance traveled by light in absolute vacuum in 1/299,792,458 of a second

3. What is the elementary unit for a chemist? A mole One mole contains Avogadro's number (approximately 6.022×1023) entities.

4. What is the elementary unit for a biologist? • A species • There are 23 – 36 definitions for what a species is

5. “... are groups of actually or potentially interbreeding natural populations which are reproductively isolated from other such groups” Ernst Mayr (1942) “... is a single lineage of ancestor-descendent populations which maintains its identity from other such lineages and which has its own evolutionary tendencies and historical fate” Ed Wiley (1978), modified from George Simpson (1961) “... is the most inclusive population of individuals having the potential for phenotypic cohesion through intrinsic cohesion mechanisms” Alan Templeton (1989) “... is the smallest diagnosable cluster of individual organisms within which there is a parental pattern of ancestry and descent. ” Joel Cracraft (1983) “… are segments of population-level lineages.” Kevin De Queiroz (1998) “... is what a good taxonomist says it is.” Anonymous

7. Who tells us what a species is? Taxonomists How? Taxonomic Keys Identification by morphological traits

8. EL EL Worker ant head measurements HL HL HW HW

9. What about unknown species? Traditional methods have identified about 1.7 million species Estimates of the total number of species are between 10 million-100 million

10. DNA Barcode: A short standardized sequence enabling species discrimination in a large block of life

11. The Barcode Region

12. DNA-based identification system

13. Known species:>150,000 >250,000 >300,000 > 30,000 flies, mosquitoes flowering plants beetles crabs, lobsters Why DNA-based identification? …because there are many, many species!

14. And many species have a variety of life forms Life cycle of Monarch butterfly

15. Biodiversity Institute of Ontario & Canadian Centre for DNA Barcoding World’s largest facility for the production and analysis of DNA barcodes

16. The Process of DNA Barcoding Collection Data Specimen Publicly-Accessible DNA Barcode, Data and Analysis Photograph Tissue Sample

17. A small portion of the specimen is removed for DNA extraction. This is incubated overnight in a chemical cocktail that breaks down cellular structure and proteins.

18. Extractions are conducted using a dedicated Biomek FX robotic system. 94 specimens can be extracted in approximately 15 minutes.

19. The gene region of choice is amplified using the polymerase chain reaction (PCR). This is carried out using specific enzymes, primers and temperatures created in these thermocyclers.

20. The success of a PCR reaction is determined by loading PCR product onto an agarose gel containing ethidium bromide. This compound binds to DNA and causes it to fluoresce under UV light.

21. A successful reaction will display this band – a failure will lack a band. High-success plates proceed to sequencing.

22. The DNA barcode is sequenced using two ABI 3730xl DNA sequencers.

23. www.barcodinglife.org

24. Current Status and Goals • International Barcode of Life (iBOL) • Goal: Barcode all life on Earth! • Currently 5.9 million DNA barcode sequences for ~545,000 species • Goal for 2020 is 1 000 000 species