GENEWATCH
 
DNA BARCODING READY FOR BREAKOUT
By Mark Stoeckle
 

from GeneWatch 26-5
Nov-Dec 2013

DNA barcoding is simple enough to be employed by high school students and versatile enough to help ferret out food fraud, rein in illicit trade in endangered timber, recognize disease-carrying mosquitos, reveal what tiny insects and big mammals eat, and speed discovery of new species on land and sea. DNA barcode similarities and differences among species help make plain the story of evolution. In its first decade DNA barcoding racked up these and many other successes. What lies ahead is expansion into routine use by consumers, educators, citizen scientists, DIY biologists, and regulatory agency enforcers.

The roots of DNA barcoding lie deep in evolutionary study and the recognition that the history of life forms is written in their DNA. The manifest story began at the University of Guelph in 2003, when Paul Hebert and colleagues made an imaginative proposal: a universal DNA identification system for all macroscopic life.1 They focused on a 650 base pair region of mitochondrial cytochrome c oxidase I (COI), previously reported recoverable from diverse animal phyla by polymerase chain reaction using a single primer pair. According to the limited data then available, this region varied little within animal species but generally differed among even closely-related species, making it usually straightforward to match sequences to species names.

Hebert and colleagues called this biological identifier a "DNA barcode" by analogy to the Uniform Product Code (UPC) for commercial goods, and presciently proposed a universal reference library. Recognizing that the value of DNA barcoding lay in standardization, it was nonetheless evident at the outset that alternative targets or amplification strategies would be needed for certain groups, including plants and fungi, as COI differed little among species in the former and often contained introns (non-coding pieces of DNA) in the latter. 

In 2003, the Alfred P. Sloan Foundation sponsored workshops at Cold Spring Harbor Laboratory which explored making this vision a reality. Participants envisioned a DNA-based standardized identification system of great use to society and science: Organisms in any life stage could be unambiguously named from bits and pieces, including morphologic "look-alike" cryptic species, by experts and non-experts, at relatively low cost, and potentially via automated devices. The biggest challenge was how to build the library, which would involve sampling multi-millions of specimens, each of which had to be identified by experts and archived in museums or herbaria so they would be available to be re-examined. New database practices were needed to link sequence records to individual specimens.

The need to organize a wide scientific community effort led to the Consortium for the Barcode of Life (CBOL), an international initiative devoted to developing DNA barcoding as a global standard for the identification of biological species, inaugurated in April 2004 with support from the Alfred P. Sloan Foundation. As of 2013, the Consortium has over 200 member organizations from over 50 countries. Participants at CBOL-sponsored workshops agreed on standard loci for animals, plants and fungi.2 The Gordon and Betty Moore Foundation provided crucial early funding that helped establish DNA barcoding as a scientific enterprise.

In parallel to CBOL, major support from Genome Canada and the Ontario Genomics Institute helped launch the International Barcode of Life (iBOL) in 2010 to speed library building and promote regional networks and taxon and ecosystem campaigns. The University of Guelph built an online database and workbench, Barcode of Life Datasystems (BOLD). As of October 2013, BOLD contains over 2.5 million barcode records from over 190,000 named animal, plant, and fungal species.

The barcoding initiative has highlighted how much we don't know about macroscopic biodiversity. Many barcoded specimens have turned out to represent new species, and many more as yet undescribed species are represented in databases, awaiting formal names. The underlying challenge is the immense number of living species: about 2 million described among an estimated 8 million existing. For animal COI barcodes, BOLD recently instituted an automated Barcode Index Number (BIN) system that assigns numbers to clusters of closely related sequences.3 In well-studied groups, BINs usually correspond to species, suggesting this algorithm can also organize records of unidentified specimens into sets representing new taxa and speed formal descriptions.

Although new sequencing technologies may supplant current Sanger standard, a decade of experience says the key elements - namely, agreed-upon standard gene region(s) and high quality sequence records from documented specimens - are irreplaceable.

Much wider use of DNA barcoding in everyday applications awaits. Potential arenas include food and herbal product testing, education, and citizen naturalists. Consumers want to know more about what they eat: where it comes from, whether it is healthy for you, and whether it is produced in a way that is good for the environment, for example. At the same time, barcoding uncovers routine mislabeling of diverse foods - in such cases, forget worrying about where it came from, it is not even the species the label says it is! These include fish (1/3 of U.S. fish products are mislabeled), ground meat (horsemeat-in-hamburger scandal in Europe), olive oil, cheese, tea, and pet food, with costs to consumers and threats to the environment.

Investigations to date suggest if a product is expensive and can't be readily identified by appearance, it is at risk of mislabeling. A recent barcoding study of herbal products, which are morphologically unidentifiable even by experts, supports this point - the majority contained contaminants, substitutions, or fillers.4 Regulatory agencies are adopting this technology, perhaps a prelude to routine use by food and herbal product distributors, enabling certification, supplemented with consumer level testing by agencies or individuals.

In education, DNA barcoding offers a relatively simple and widely applicable technology that allows students to design and carry out diverse investigations. Science is a process of discovery, but most high school laboratory exercises have pre-determined outcomes. DNA barcoding enables students to make real discoveries and contribute to reference databases. Model projects in a few dozen schools, notably in NYC, California, and Ontario, have demonstrated the potential.5-7 These could serve as templates and be expanded to be a routine component of high school biology. Another plus is that comparing DNA barcode sequences to reference databases provides students a direct look at evolution.

DNA barcoding democratizes access to knowledge about biodiversity. It can help citizen naturalists and DIY biologists make modern day voyages of bio-discovery in urban, rural, and wild environments. Just as GPS went from a cumbersome research tool to an everyday application, DNA barcoding has potential appeal to a large, diverse set of interested individuals.

For wider commercial use, such as by food distributors, certified, affordable, and speedy barcode testing services are needed. Robust and rapid protocols for DNA extraction, amplification, and sequencing are already available, indicating the hurdles, if any, are in business aspects. For consumer, educational, and DIY use, easy-to-use inexpensive kits with mail-in sequencing services would likely boost demand.

We look forward to DNA barcoding's breakout applications.

 

Mark Stoeckle, MD, is Senior Research Associate in the Program for the Human Environment at The Rockefeller University. He has been involved in the DNA Barcoding Initiative since its beginnings in 2003.




ENDNOTES

1. Hebert PDN, Cywinska A, Ball SL, et al (2003) Proc R Soc Lond B 270:313. (link)

2. More specifically, those loci are: Animals, 5' COI (2005); plants, rbcL/matK (2009); and fungi, ITS (2012).

3. Ratnasingham S, Hebert PDN (2013) PLoS ONE 8:e66213. (link)

4. Newmaster SG, Grguric M, Shanmughanandhan D, et al. (2013) BMC Medicine 11:222 (link)

5. http://www.urbanbarcodeproject.org

6. Santschi L, Hanner RH, Ratnasingham S, et al (2013) PLoS Biol 11:e1001471 (link)

7. http://malaiseprogram.ca

 
 
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