DNA barcoding 2613851 224207099 2008-07-07T20:22:31Z Dyanega 1357080 additional reference '''DNA barcoding''' is a [[Taxonomy|taxonomic]] method that uses a short genetic marker in an organism's [[mtDNA|mitochondrial DNA]] to identify it as belonging to a particular [[species]]. It is based on a relatively simple concept: most [[eukaryote]] cells contain [[mitochondria]] and mitochondrial DNA (mtDNA) has a relatively fast [[mutation]] rate, which results in significant variance in mtDNA sequences between species and, in principle, a comparatively small variance within species. However, because all [[mtDNA]] genes are maternally inherited (except in a few [[bivalve]]s{{fact|date = May 2008}}), any occurrences of [[Hybrid (biology)|hybridization]], male-killing microoroganisms<ref name = "JohnstoneHurst">Johnstone, R.A., Hurst, G.D.D. (1996) Maternally inherited male-killing microorganisms may confound interpretation of mitochondrial DNA variability. Biol. J. Linnaean Soc. 58:453-470.</ref>, cytoplasmic incompatibility-inducing symbionts (e.g., ''[[Wolbachia]]'')<ref name = JohnstoneHurst/>, [[horizontal gene transfer]] (such as via cellular symbionts<ref name = "Hurstetal">Hurst, Gregory D. D.; Jiggins, Francis M. (2005) Problems with mitochondrial DNA as a marker in population, phylogeographic and phylogenetic studies: the effects of inherited symbionts. Proceedings of the Royal Society B, 272: 1525-1534</ref>), or other "reticulate" evolutionary phenomena in a lineage can lead to misleading results (i.e., it is possible for two different species to share mtDNA<ref>P. J. P. Croucher, G. S. Oxford, J. B. Searle (2004) Mitochondrial differentiation, introgression and phylogeny of species in the ''[[Tegenaria atrica]]'' group (Araneae: Agelenidae). Biological Journal of the Linnean Society 81: 79–89. doi:10.1111/j.1095-8312.2004.00280.x</ref>, or for one species to have more than one mtDNA sequence exhibited among different individuals)<ref name = "Whitworthetal">T.L. Whitworth, R.D. Dawson, H. Magalon, E. Baudry (2007) DNA barcoding cannot reliably identify species of the blowfly genus ''Protocalliphora'' (Diptera: Calliphoridae). Proceedings of the Royal Society B: 274: 1731-1739</ref><ref name = "Meier">Meier, R. (2008). DNA sequences in taxonomy: Opportunities and challenges. Chap. 7 in: The New Taxonomy. Wheeler, Q., ed. CRC Press, Boca Raton, FL, USA</ref>. A 648-[[base pair|bp]] region of the [[cytochrome c oxidase]] subunit I gene (COI) was initially proposed as a potential 'barcode'. ==Origin== The use of [[nucleotide]] sequence variations to investigate evolutionary relationships is not a new concept. [[Carl Woese]] used sequence differences in [[rRNA|ribosomal RNA]] (rRNA) to discover [[archaebacteria]], which in turn led to the redrawing of the [[evolutionary tree]], and molecular markers (e.g., allozymes, rDNA, and mtDNA) have been successfully used in molecular [[systematics]] for decades. DNA barcoding provides a standardised method for this process via the use of a short DNA sequence from a particular region of the genome to provide a 'barcode' for identifying species. In 2003, [[Paul D.N. Hebert|Professor Paul D.N. Hebert]] from the [[University of Guelph]], [[Ontario]], [[Canada]], proposed the compilation of a public library of DNA barcodes that would be linked to named [[specimen]]s. This library would “provide a new master key for identifying species, one whose power will rise with increased taxon coverage and with faster, cheaper sequencing”. ==Case studies== ===Identification of birds=== In an effort to find a correspondence between traditional species boundaries established by taxonomy and those inferred by DNA barcoding, Hebert and co-workers sequenced DNA barcodes of 260 of the 667 bird species that breed in [[North America]] (Hebert ''et al.'' 2004a<ref name = "HebertBirds">Hebert, P. D. N.; Stoeckle, M. Y.; Zemlak, T. S. & Francis, C. M. (2004a): Identification of Birds Through DNA Barcodes. ''[[PLoS Biology|PLoS Biol.]]'' '''2'''(10): 1657-1663. {{DOI|10.1371/journal.pbio.0020312}} [http://biology.plosjournals.org/perlserv/?request=get-pdf&file=10.1371_journal.pbio.0020312-L.pdf PDF fulltext] [http://biology.plosjournals.org/archive/1545-7885/2/10/supinfo/10.1371_journal.pbio.0020312.sg001.pdf Supporting Information]</ref>). They found that every single one of the 260 species had a different COI sequence. 130 species were represented by two or more specimens; in all of these species, COI sequences were either identical or were most similar to sequences of the same species. COI variations between species averaged 7.93%, whereas variation within species averaged 0.43%. In four cases there were deep intraspecific divergences, indicating possible new species. Three out of these four [[polytypic]] species are already split into two by some taxonomists. Hebert ''et al.'''s (2004a<ref name = "HebertBirds"/>) results reinforce these views and strengthen the case for DNA barcoding. Hebert ''et al.'' also proposed a standard sequence threshold to define new species, this threshold, the so-called "barcoding gap", was defined as 10 times the mean intraspecific variation for the group under study. ===Delimiting cryptic species=== The next major study into the efficacy of DNA barcoding was focused on the [[neotropical]] skipper butterfly, ''[[Astraptes|Astraptes fulgerator]]'' at the [[Area Conservacion de Guanacaste]] (ACG) in north-western [[Costa Rica]]. This species was already known as a [[cryptic species complex]], due to subtle [[morphology (biology)|morphological]] differences, as well as an unusually large variety of [[caterpillar]] food plants. However, several years would have been required for taxonomists to completely delimit species. Hebert ''et al.'' (2004b<ref name = "HebertAstraptes">Hebert, P. D. N.; Penton, E. H.; Burns, J. M.; Janzen, D. H. & Hallwachs, W. (2004b): Ten Species in One: DNA Barcoding Reveals Cryptic Species in the Neotropical Skipper Butterfly ''Astraptes fulgerator''. ''[[PNAS|Proc. Natl. Acad. Sci. USA]]'' '''101'''(41): 14812-14817. {{DOI|10.1073/pnas.0406166101}} [http://www.pnas.org/cgi/reprint/101/41/14812.pdf PDF fulltext] [http://www.pnas.org/cgi/content/full/0406166101/DC1 Supporting Information]</ref>) sequenced the COI gene of 484 specimens from the ACG. This sample included “at least 20 individuals reared from each species of food plant, extremes and intermediates of adult and caterpillar color variation, and representatives” from the three major ecosystems where ''Astraptes fulgerator'' is found. Hebert ''et al.'' (2004b<ref name = "HebertAstraptes"/>) concluded that ''Astraptes fulgerator'' consists of 10 different species in north-western Costa Rica. These results, however, were subsequently challenged by Brower (2006)<ref name = "Brower">Brower, A.V.Z. (2006) Problems with DNA barcodes for species delimitation: 'ten species' of ''Astraptes fulgerator'' reassessed (Lepidoptera: Hesperiidae). ''Systematics and Biodiversity'' 4(2): 127-132.</ref>, who pointed out numerous serious flaws in the analysis, and concluded that the original data could support no more than the possibility of three to seven cryptic [[taxa]] rather than ten cryptic species. This highlights that the results of DNA barcoding analyses can be dependent upon the choice of analytical methods used by the investigators, so the process of delimiting cryptic species using DNA barcodes can be as subjective as any other form of taxonomy. A more recent example used DNA barcoding for the identification of cryptic species included in the ongoing long-term database of tropical caterpillar life generated by Dan Janzen and Winnie Hallwachs in Costa Rica at the ACG.<ref name=caterpillar>{{cite web |url=http://janzen.sas.upenn.edu/caterpillars/database.lasso |title=Database homepage for ACG caterpillar (Lepidoptera) rearing databases |accessdate=2007-08-12}}</ref> In 2006 Smith ''et al.''<ref name=Smith2006>{{cite journal |author=Smith MA, Woodley NE, Janzen DH, Hallwachs W, Hebert PD |title=DNA barcodes reveal cryptic host-specificity within the presumed polyphagous members of a genus of parasitoid flies (Diptera: Tachinidae) |journal=Proc. Natl. Acad. Sci. U.S.A. |volume=103 |issue=10 |pages=3657–62 |year=2006 |url=http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=1383497 |pmid=16505365 |doi=10.1073/pnas.0511318103}}</ref> examined whether a COI DNA barcode could function as a tool for identification and discovery for the 20 morphospecies of ''[[Belvosia]]'' [http://www.itis.gov/servlet/SingleRpt/SingleRpt?search_topic=TSN&search_value=650659] [[parasitoid]] flies ([[Tachinidae]]) that have been reared from [[caterpillar]]s in ACG. Barcoding not only discriminated among all 17 highly host-specific morphospecies of ACG ''Belvosia'', but it also suggested that the species count could be as high as 32 by indicating that each of the three generalist species might actually be arrays of highly host-specific cryptic species. In 2007 Smith ''et al.'' expanded on these results by barcoding 2,134 flies belonging to what appeared to be the 16 most generalist of the ACG tachinid morphospecies.<ref name=Smith2007>{{cite journal |author=Smith MA, Wood DM, Janzen DH, Hallwachs W, Hebert PD |title=DNA barcodes affirm that 16 species of apparently generalist tropical parasitoid flies (Diptera, Tachinidae) are not all generalists |journal=Proc. Natl. Acad. Sci. U.S.A. |volume=104 |issue=12 |pages=4967–72 |year=2007 |url=http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=1821123 |pmid=17360352 |doi=10.1073/pnas.0700050104}}</ref> They encountered 73 mitochondrial lineages separated by an average of 4% sequence divergence and, as these lineages are supported by collateral ecological information, and, where tested, by independent nuclear markers (28S and ITS1), the authors therefore viewed these lineages as provisional species. Each of the 16 initially apparent generalist species were categorized into one of four patterns: (i) a single generalist species, (ii) a pair of morphologically cryptic generalist species, (iii) a complex of specialist species plus a generalist, or (iv) a complex of specialists with no remaining generalist. In sum, there remained 9 generalist species classified among the 73 mitochondrial lineages analyzed. However, also in 2007, Whitworth ''et al.'' reported that flies in the related family [[Calliphoridae]] could not be discriminated by barcoding.<ref name = Whitworthetal/> They investigated the performance of barcoding in the fly genus ''[[Protocalliphora]]'', known to be infected with the [[endosymbiotic]] bacteria ''[[Wolbachia]]''. Assignment of unknown individuals to species was impossible for 60% of the species, and if the technique had been applied, as in the previous study, to identify new species, it would have underestimated the species number in the genus by 75%. They attributed the failure of barcoding to the non-monophyly of many of the species at the mitochondrial level; in one case, individuals from four different species had identical barcodes. The authors went on to state: {{quote|The pattern of ''Wolbachia'' infection strongly suggests that the lack of within-species monophyly results from introgressive hybridization associated with ''Wolbachia'' infection. Given that ''Wolbachia'' is known to infect between 15 and 75% of insect species, we conclude that identification at the species level based on mitochondrial sequence might not be possible for many insects.<ref name = Whitworthetal/>}} Marine biologists have also considered the value of the technique in identifying cryptic and polymorphic species and have suggested that the technique may be helpful when associations with voucher specimens are maintained<ref>Schander, C. & Willassen E. 2005. What can Biological Barcoding do for Marine Biology? ''Marine Biology Research'' '''1'''(1): 79-83. <!-- {{DOI|10.1080/17451000510018962}} DOI seems incorrectly formed -->[http://www.bolinfonet.org/pdf/schander&willassen_2005.pdf PDF]</ref>, though cases of "shared barcodes" (e.g., non-unique) have been documented in [[cichlid]] fishes and [[cowries]]<ref name =Meier/>. ===Identifying flowering plants=== [[W. John Kress|Kress]] ''et al.'' (2005<ref name = "Kress">Kress, W. J.; Wurdack, K. J.; Zimmer, E. A.; Weigt, L. A. & Janzen, D. H. (2005): Use of DNA Barcodes to Identify Flowering Plants. ''[[PNAS|Proc. Nat. Acad. Sci. USA]]'' '''102'''(23): 8369-8374. {{DOI|10.1073/pnas.0503123102}} [http://www.pnas.org/cgi/reprint/102/23/8369.pdf PDF fulltext] [http://www.pnas.org/cgi/content/full/0503123102/DC1 Supporting Information]</ref>) suggest that the use of the COI sequence “is not appropriate for most species of plants because of a much slower rate of cytochrome c oxidase I gene evolution in higher plants than in animals”. A series of experiments was then conducted to find a more suitable region of the [[genome]] for use in the DNA barcoding of [[flowering plant]]s. Three criteria were set for the appropriate genetic [[locus (genetics)|loci]]: #Significant species-level genetic variability and divergence #An appropriately short sequence length so as to facilitate DNA extraction and amplification, and #The presence of conserved flanking sites for developing universal primers. At the conclusion of these experiments, Kress ''et al.'' (2005<ref name = "Kress"/>) proposed the nuclear [[internal transcribed spacer]] region and the plastid trnH-psbA intergenic spacer as a potential DNA barcode for flowering plants. These results suggest that DNA barcoding, rather than being a 'master key' may be a 'master keyring', with different kingdoms of life requiring different keys. ===Cataloguing ancient life=== Lambert ''et al.'' (2005<ref>Lambert, D. M.; Baker, A,; Huynen, L.; Haddrath, O.; Hebert, P. D. N. & Millar, C. D. (2005): Is a Large-Scale DNA-Based Inventory of Ancient Life Possible? ''Journal of Heredity'' '''96'''(3): 279-284. {{DOI|10.1093/jhered/esi035}} [http://jhered.oxfordjournals.org/cgi/reprint/96/3/279.pdf PDF fulltext]</ref>) examined the possibility of using DNA barcoding to assess the past diversity of the earth's [[biota]]. The COI gene of a group of extinct [[ratite]] birds, the [[moa]], were sequenced using 26 [[subfossil]] moa bones. As with Hebert's results, each species sequenced had a unique barcode and intraspecific COI sequence variance ranged from 0 to 1.24%. To determine new species, a standard sequence threshold of 2.7% COI sequence difference was set. This value is 10 times the average intraspecies difference of North American birds, which is inconsistent with Hebert's recommendation that the threshold value be based on the group under study. Using this value, the group detected six moa species. In addition, a further standard sequence threshold of 1.24% was also used. This value resulted in 10 moa species which corresponded with the previously known species with one exception. This exception suggested a possible complex of species which was previously unidentified. Given the slow rate of growth and reproduction of moa, it is probable that the interspecies variation is rather low. On the other hand, there is no set value of molecular difference at which populations can be assumed to have irrevocably started to undergo [[speciation]]. It is safe to say, however, that the 2.7% COI sequence difference initially used was far too high. ==Criticisms== DNA barcoding has met with spirited reaction from scientists, especially [[systematics|systematists]], ranging from enthusiastic endorsement to vociferous opposition. For example, many stress the fact that DNA barcoding does not provide reliable information above the species level, while others indicate that it is inapplicable at the species level, but may still have merit for higher-level groups<ref name = Whitworthetal/>. Others resent what they see as a gross oversimplification of the science of taxonomy. And, more practically, some suggest that recently diverged species might not be distinguishable on the basis of their COI sequences<ref>Kevin, C.R. Kerr, Mark Y. Stoeckle, Carla J. Dove, Lee A. Weigt, Charles M. Francis & Paul D. N. Hebert. 2006. Comprehensive DNA barcode coverage of North American birds. Molecular Ecology Notes. (OnlineEarly Articles). doi:10.1111/j.1471-8286.2006.01670.x [http://www.blackwell-synergy.com/doi/full/10.1111/j.1471-8286.2006.01670.x Full text]</ref>. Due to various phenomena, Funk & Omland (2003<ref>Funk, D.J., Omland, K.C. (2003) Species-level paraphyly and polyphyly: frequency, causes, and consequences, with insights from animal mitochondrial DNA. Annual Review of Ecology and Systematics 34: 397-423.</ref>) found that some 23% of animal species are [[polyphyletic]] if their mtDNA data are accurate, indicating that using an mtDNA barcode to assign a species name to an animal will be ambiguous or erroneous some 23% of the time (see also Meyer & Paulay, 2005<ref>Meyer, C.P., Paulay, G. (2005) DNA barcoding: error rates based on comprehensive sampling. ''[[PLoS Biology|PLoS Biol.]]'' '''3''' e422</ref>). Studies with insects suggest an equal or even greater error rate, due to the frequent lack of correlation between the mitochondrial genome and the nuclear genome or the lack of a barcoding gap (e.g., Hurst and Jiggins, 2005<ref name = Hurstetal/>, Whitworth ''et al.'', 2007<ref name = Whitworthetal/>, Wiemers & Fiedler, 2007<ref>Wiemers, M. & Fiedler, K. (2007) Does the DNA barcoding gap exist? – a case study in blue butterflies (Lepidoptera: Lycaenidae). ''[http://www.frontiersinzoology.com/content/4/1/8 Front. Zool. 4:8]''</ref>). Problems with mtDNA arising from male-killing microoroganisms and cytoplasmic incompatibility-inducing symbionts (e.g., ''[[Wolbachia]]'')<ref name = JohnstoneHurst/> are also particularly common among insects. Given that insects represent over 75% of all known organisms[http://www.environment.gov.au/biodiversity/abrs/publications/other/species-numbers/02-exec-summary.html#allspecies], this suggests that while mtDNA barcoding may work for [[vertebrate]]s, it may not be effective for the majority of known organisms. Moritz and Cicero (2004<ref>Moritz, C. & Cicero, C. (2004): DNA Barcoding: Promise and Pitfalls. ''[[PLoS Biology|PLoS Biol.]]'' '''2'''(10): 1529-1531. {{DOI|10.1371/journal.pbio.0020354}} [http://biology.plosjournals.org/perlserv/?request=get-pdf&file=10.1371_journal.pbio.0020354-L.pdf PDF fulltext]</ref>) have questioned the efficacy of DNA barcoding by suggesting that other avian data is inconsistent with Hebert ''et al.'''s interpretation, namely, Johnson and Cicero's (2004<ref name = "JohnsonCicero">Johnson, N. K.; Cicero, C. (2004): New mitochondrial DNA data affirm the importance of Pleistocene speciation in North American birds''. ''[[Evolution]]'' '''58''': 1122-1130.</ref>) finding that 74% of sister species comparisons fall below the 2.7% threshold suggested by Hebert ''et al.'' These criticisms are somewhat misleading considering that, of the 39 species comparisons reported by Johnson and Cicero, only 8 actually use COI data to arrive at their conclusions. Johnson and Cicero (2004<ref name = "JohnsonCicero"/>) have also claimed to have detected bird species with identical DNA barcodes, however, these 'barcodes' refer to an unpublished 723-bp sequence of ND6 which has never been suggested as a likely candidate for DNA barcoding. The DNA barcoding debate resembles the [[phenetics]] debate of decades gone by. It remains to be seen whether what is now touted as a revolution in taxonomy will eventually go the same way as phenetic approaches, of which was claimed exactly the same decades ago, but which were all but rejected when they failed to live up to overblown expectations.<ref>Will, K. W., B. D. Mishler & Q. D. Wheeler 2005. The Perils of DNA Barcoding and the Need for Integrative Taxonomy Syst. Biol. 54(5):844–851 [http://www.erin.utoronto.ca/~w3bio/bio443/seminar_papers/perils_of_dna_barcoding.pdf]</ref> Controversy surrounding DNA barcoding stems not so much from the method itself, but rather from extravagant claims that it will supersede or radically transform traditional taxonomy. Other critics fear a "[[big science]]" initiative like barcoding will make funding even more scarce for already underfunded disciplines like [[taxonomy]], but barcoders respond that they compete for funding not with fields like taxonomy, but instead with other big science fields, such as [[medicine]] and [[genomics]].<ref>Gregory, R. T. 2005. DNA barcoding does not compete with taxonomy. Nature 434:1067 (28 April 2005) [http://www.bolinfonet.org/pdf/DNA_barcoding_does_not_compete_with_taxonomy.pdf]</ref> The current trend appears to be that DNA barcoding needs to be used alongside traditional [[taxonomy|taxonomic]] tools and alternative forms of molecular systematics so that problem cases can be identified and errors detected. Non-cryptic species can generally be resolved by either traditional or molecular taxonomy without ambiguity. However, more difficult cases will only yield to a combination of approaches. And finally, as most of the global [[biodiversity]] remains unknown, molecular barcoding can only hint at the existence of new taxa, but not delimit or describe them (DeSalle, 2006<ref>DeSalle, R. (2006): Species discovery versus species identification in DNA barcoding efforts: response to Rubinoff. ''Conservation Biology'' '''20'''(5): 1545-1547. {{DOI|10.1111/j.1523-1739.2006.00543.x}}</ref>; Rubinoff, 2006<ref>Rubinoff, D. (2006a): Utility of mitochondrial DNA barcodes in species conservation. ''Conservation Biology'' '''20'''(4): 1026–1033. {{DOI|10.1111/j.1523-1739.2006.00372.x}}</ref><ref>Rubinoff, D. (2006b): DNA Barcoding Evolves into the Familiar. ''Conservation Biology'' '''20'''(5): 1548–1549. {{DOI|10.1111/j.1523-1739.2006.00542.x}}</ref>). ==References==<!-- Cladistics20:47. Zootaxa1586:67. --> {{reflist|2}} ==See also== *[[DNA taxonomy]] *[[Consortium for the Barcode of Life]] ==External links== * [http://www.barcodinglife.org Barcode of Life Database] * [http://www.dnabarcoding.org/ International Barcode of Life] * [http://barcoding.si.edu/index_detail.htm Consortium for the Barcode of Life] * [http://www.fishbol.org Fish Barcode of Life Initiative (FISH-BOL)] * [http://barcoding.si.edu/AllBirds.htm All Birds Barcoding Initiative (ABBI)] * [http://www.polarbarcoding.org Polar Flora and Fauna Barcoding website] (Latest outpost in the Canadian Arctic in the field) * [http://phe.rockefeller.edu/barcode/blog/ The Barcode of Life Blog] *[http://www.barcoding.si.edu/PDF/Guidelines%20for%20non-CO1%20selection%20-%204%20June.pdf Guidelines for non COI gene selection] {{phylo}} [[Category:Taxonomy]] [[Category:Molecular genetics]] [[Category:Bioinformatics]] [[Category:Authentication methods]] [[Category:Biometrics]] [[de:DNA barcoding]] [[id:DNA barcode]] [[it:DNA barcoding]] [[nl:DNA-barcoding]]