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{"title"=>"Thar she blows! A novel method for DNA collection from cetacean blow", "type"=>"journal", "authors"=>[{"first_name"=>"Céline H.", "last_name"=>"Frère", "scopus_author_id"=>"24174363400"}, {"first_name"=>"Ewa", "last_name"=>"Krzyszczyk", "scopus_author_id"=>"36551158700"}, {"first_name"=>"Eric M.", "last_name"=>"Patterson", "scopus_author_id"=>"35991682300"}, {"first_name"=>"Sue", "last_name"=>"Hunter", "scopus_author_id"=>"55106214600"}, {"first_name"=>"Alison", "last_name"=>"Ginsburg", "scopus_author_id"=>"36550932200"}, {"first_name"=>"Janet", "last_name"=>"Mann", "scopus_author_id"=>"7402735648"}], "year"=>2010, "source"=>"PLoS ONE", "identifiers"=>{"scopus"=>"2-s2.0-77957872227", "doi"=>"10.1371/journal.pone.0012299", "issn"=>"19326203", "pui"=>"359754328", "sgr"=>"77957872227", "isbn"=>"1932-6203", "pmid"=>"20811619"}, "id"=>"fdf70528-a92e-34ab-ad7c-751ab41840d6", "abstract"=>"BACKGROUND: Molecular tools are now widely used to address crucial management and conservation questions. To date, dart biopsying has been the most commonly used method for collecting genetic data from cetaceans; however, this method has some drawbacks. Dart biopsying is considered inappropriate for young animals and has recently come under scrutiny from ethical boards, conservationists, and the general public. Thus, identifying alternative genetic collection techniques for cetaceans remains a priority, especially for internationally protected species. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated whether blow-sampling, which involves collecting exhalations from the blowholes of cetaceans, could be developed as a new less invasive method for DNA collection. Our current methodology was developed using six bottlenose dolphins, Tursiops truncatus, housed at the National Aquarium, Baltimore (USA), from which we were able to collect both blow and blood samples. For all six individuals, we found that their mitochondrial and microsatellite DNA profile taken from blow, matched their corresponding mitochondrial and microsatellite DNA profile collected from blood. This indicates that blow-sampling is a viable alternative method for DNA collection. CONCLUSION/SIGNIFICANCE: In this study, we show that blow-sampling provides a viable and less invasive method for collection of genetic data, even for small cetaceans. In contrast to dart biopsying, the advantage of this method is that it capitalizes on the natural breathing behaviour of dolphins and can be applied to even very young dolphins. Both biopsy and blow-sampling require close proximity of the boat, but blow-sampling can be achieved when dolphins voluntarily bow-ride and involves no harmful contact.", "link"=>"http://www.mendeley.com/research/thar-blows-novel-method-dna-collection-cetacean-blow", "reader_count"=>133, "reader_count_by_academic_status"=>{"Unspecified"=>2, "Professor > Associate Professor"=>4, "Librarian"=>1, "Researcher"=>35, "Student > Doctoral Student"=>7, "Student > Ph. D. Student"=>33, "Student > Postgraduate"=>6, "Student > Master"=>21, "Other"=>10, "Student > Bachelor"=>6, "Lecturer"=>2, "Lecturer > Senior Lecturer"=>3, "Professor"=>3}, "reader_count_by_user_role"=>{"Unspecified"=>2, "Professor > Associate Professor"=>4, "Librarian"=>1, "Researcher"=>35, "Student > Doctoral Student"=>7, "Student > Ph. D. Student"=>33, "Student > Postgraduate"=>6, "Student > Master"=>21, "Other"=>10, "Student > Bachelor"=>6, "Lecturer"=>2, "Lecturer > Senior Lecturer"=>3, "Professor"=>3}, "reader_count_by_subject_area"=>{"Unspecified"=>2, "Environmental Science"=>22, "Biochemistry, Genetics and Molecular Biology"=>4, "Agricultural and Biological Sciences"=>96, "Medicine and Dentistry"=>1, "Veterinary Science and Veterinary Medicine"=>1, "Business, Management and Accounting"=>1, "Psychology"=>1, "Social Sciences"=>2, "Immunology and Microbiology"=>1, "Earth and Planetary Sciences"=>2}, "reader_count_by_subdiscipline"=>{"Medicine and Dentistry"=>{"Medicine and Dentistry"=>1}, "Social Sciences"=>{"Social Sciences"=>2}, "Psychology"=>{"Psychology"=>1}, "Immunology and Microbiology"=>{"Immunology and Microbiology"=>1}, "Earth and Planetary Sciences"=>{"Earth and Planetary Sciences"=>2}, "Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>96}, "Business, Management and Accounting"=>{"Business, Management and Accounting"=>1}, "Biochemistry, Genetics and Molecular Biology"=>{"Biochemistry, Genetics and Molecular Biology"=>4}, "Unspecified"=>{"Unspecified"=>2}, "Environmental Science"=>{"Environmental Science"=>22}, "Veterinary Science and Veterinary Medicine"=>{"Veterinary Science and Veterinary Medicine"=>1}}, "reader_count_by_country"=>{"Colombia"=>1, "Romania"=>1, "United States"=>5, "United Kingdom"=>3, "United Arab Emirates"=>1, "Portugal"=>2, "Spain"=>1, "New Zealand"=>1, "Netherlands"=>1, "Brazil"=>3, "Denmark"=>1, "Italy"=>1, "Chile"=>1, "France"=>1}, "group_count"=>3}

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• http://doi.org/10.1021/ac5024217
• http://doi.org/10.1111/mec.13948
• http://doi.org/10.3354/esr00524
• http://doi.org/10.1371/journal.pone.0101427
• http://doi.org/10.1093/jmammal/gyx036
• http://doi.org/10.3390/metabo4030790
• http://doi.org/10.1002/ece3.2346
• http://doi.org/10.1038/s41598-018-28200-8
• http://doi.org/10.1071/MA15004
• http://doi.org/10.1371/journal.pone.0114062
• http://doi.org/10.1002/ecs2.2901
• http://doi.org/10.3389/fmars.2017.00148
• http://doi.org/10.1007/s10531-019-01709-8
• http://doi.org/10.1111/mms.12482
• http://doi.org/10.3390/genes10030192
• http://doi.org/10.1111/j.1758-2229.2011.00306.x
• http://doi.org/10.1292/jvms.18-0749
• http://doi.org/10.1093/conphys/cot006
• http://doi.org/10.1371/journal.pone.0215020
• http://doi.org/10.3354/meps13055
• http://doi.org/10.1371/journal.pone.0235537

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• {"type"=>"COMMENT", "annotationUri"=>"info:doi/10.1371/annotation/e1109d10-4563-4ef1-9881-544247b41b87", "title"=>"Other applications", "body"=>"I found this protocol very interesting regarding animal welfare in experimental sciences. I would like to know if there is posibble to obtain RNA from these method too. It will be useful for further analysis in gene expression modulation from differente conservative and veterinarian medicine approaches, and other research subjects. Thanks in advance for your comments.", "isRemoved"=>false, "created"=>"2010-09-05T19:32:59Z", "lastModified"=>"2010-09-05T19:32:59Z", "creator"=>{"userId"=>"136743"}, "highlightedText"=>"", "competingInterestStatement"=>{"creatorWasPrompted"=>true, "hasCompetingInterests"=>false}, "parentArticle"=>{"doi"=>"info:doi/10.1371/journal.pone.0012299", "state"=>"published", "journals"=>{"PLoSONE"=>{"journalKey"=>"PLoSONE", "eIssn"=>"1932-6203", "title"=>"PLOS ONE"}}}, "replyTreeSize"=>0, "mostRecentActivity"=>"2010-09-05T19:32:59Z", "replies"=>[]}

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Figshare

• {"files"=>["https://ndownloader.figshare.com/files/833452"], "description"=>"<p>To estimate whether the microsatellite profile collected from an individual's blow matched the microsatellite profile collected from its blood, we amplified 3 polymorphic dinucleotide microsatellite loci: Lobs_Di9 (black), Lobs-Di19 (red), and Lobs_Di21 (green) <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012299#pone.0012299-Cassens1\" target=\"_blank\">[24]</a>. The three loci were amplified using a Qiagen Multiplex KitTM (Qiagen). The annealing temperature was set at 53°C for 60 seconds for a total of 30 cycles. PCR products were then run on an ABI 3730 DNA Sequencer (Applied Biosystems). Microsatellite profiles were visualised using the software GeneMapper (version 4.0). For ease of presentation, we only include here a figure showing the microsatellite DNA profile of blood and blow for one individual. The remaining 5 microsatellite DNA profiles are included in supporting online information <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012299#pone.0012299.s001\" target=\"_blank\">Figure S1</a>. The top 3 panels represent the three microsatellite loci amplified from DNA extracted from blow.</p>", "links"=>[], "tags"=>["dna"], "article_id"=>503824, "categories"=>["Genetics", "Ecology"], "users"=>["Céline H. Frère", "Ewa Krzyszczyk", "Eric M. Patterson", "Sue Hunter", "Alison Ginsburg", "Janet Mann"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0012299.g002", "stats"=>{"downloads"=>1, "page_views"=>29, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Microsatellite_DNA_profile_of_blow_and_blood_from_individual_1_/503824", "title"=>"Microsatellite DNA profile of blow and blood from individual 1.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-08-25 01:03:44"}
• {"files"=>["https://ndownloader.figshare.com/files/415287", "https://ndownloader.figshare.com/files/415297"], "description"=>"<div><h3>Background</h3><p>Molecular tools are now widely used to address crucial management and conservation questions. To date, dart biopsying has been the most commonly used method for collecting genetic data from cetaceans; however, this method has some drawbacks. Dart biopsying is considered inappropriate for young animals and has recently come under scrutiny from ethical boards, conservationists, and the general public. Thus, identifying alternative genetic collection techniques for cetaceans remains a priority, especially for internationally protected species.</p><h3>Methodology/Principal Findings</h3><p>In this study, we investigated whether blow-sampling, which involves collecting exhalations from the blowholes of cetaceans, could be developed as a new less invasive method for DNA collection. Our current methodology was developed using six bottlenose dolphins, <em>Tursiops truncatus</em>, housed at the National Aquarium, Baltimore (USA), from which we were able to collect both blow and blood samples. For all six individuals, we found that their mitochondrial and microsatellite DNA profile taken from blow, matched their corresponding mitochondrial and microsatellite DNA profile collected from blood. This indicates that blow-sampling is a viable alternative method for DNA collection.</p><h3>Conclusion/Significance</h3><p>In this study, we show that blow-sampling provides a viable and less invasive method for collection of genetic data, even for small cetaceans. In contrast to dart biopsying, the advantage of this method is that it capitalizes on the natural breathing behaviour of dolphins and can be applied to even very young dolphins. Both biopsy and blow-sampling require close proximity of the boat, but blow-sampling can be achieved when dolphins voluntarily bow-ride and involves no harmful contact.</p></div>", "links"=>[], "tags"=>["thar", "dna", "cetacean"], "article_id"=>141997, "categories"=>["Genetics", "Ecology"], "users"=>["Céline H. Frère", "Ewa Krzyszczyk", "Eric M. Patterson", "Sue Hunter", "Alison Ginsburg", "Janet Mann"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0012299.s001", "https://dx.doi.org/10.1371/journal.pone.0012299.s002"], "stats"=>{"downloads"=>0, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/Thar_She_Blows_A_Novel_Method_for_DNA_Collection_from_Cetacean_Blow/141997", "title"=>"Thar She Blows! A Novel Method for DNA Collection from Cetacean Blow", "pos_in_sequence"=>0, "defined_type"=>4, "published_date"=>"2010-08-25 00:33:17"}
• {"files"=>["https://ndownloader.figshare.com/files/833332"], "description"=>"<p>A test tube was held inverted over the dolphin's blowhole and the dolphins were trained to exhale on cue (Figure 1). Four to six exhalations were collected per tube. We collected two sets of blow samples from each individual using sterile 50 ml polypropylene tubes (Fisherbrand) to test two different storage and transportation methods (detailed below). Both sets of blow samples were stored on dry ice for transportation to Georgetown University (Washington DC, USA). Once in the laboratory, the first set of blow samples were resuspended in 1 ml of 100% ethanol. The sides were scraped down using sterilized policemen, vortexed, and finally centrifuged at 3000 rpm for 3 minutes. Samples were then transferred to 1.5 ml microcentrifuge tubes to dry off the ethanol in a speed vacuum. The second set of blow samples were resuspended in 500 µl of TE buffer and centrifuged at 3000 rpm for 3 minutes. Excess TE buffer was carefully removed from all samples leaving a small amount at the bottom of the tube. DNA extraction of both sets of blow samples and blood samples were completed using a Qiagen DNeasy Blood and Tissue kit, using the animal tissues spin-column protocol. After DNA purification, blow samples were stored at −80°C until transportation to the University of Queensland (Brisbane, Australia) for DNA fingerprinting. A control sample of seawater was taken along with each blow sample set to assure that any DNA results were from blow samples and not seawater contamination.</p>", "links"=>[], "tags"=>["sterile", "50", "ml", "polypropylene"], "article_id"=>503692, "categories"=>["Genetics", "Ecology"], "users"=>["Céline H. Frère", "Ewa Krzyszczyk", "Eric M. Patterson", "Sue Hunter", "Alison Ginsburg", "Janet Mann"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0012299.g001", "stats"=>{"downloads"=>1, "page_views"=>17, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Blow_sample_collection_using_sterile_50_ml_polypropylene_tubes_/503692", "title"=>"Blow sample collection using sterile 50 ml polypropylene tubes.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-08-25 01:01:32"}
• {"files"=>["https://ndownloader.figshare.com/files/833612"], "description"=>"<p>To estimate whether the mitochondrial DNA profile collected from an individual's blow matched the mitochondrial DNA profile collected from its blood, we amplified a 426 base-pair fragment of the maternally inherited mitochondrial DNA control region using the primers dlp1.5 <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012299#pone.0012299-Baker2\" target=\"_blank\">[25]</a> and dlp5 <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012299#pone.0012299-Krtzen1\" target=\"_blank\">[26]</a>. The PCR products were cleaned using ExoSAP-IT from Affymetrix. Cycle sequencing was conducted with the BigDyeTM Terminator Cycle Sequencing Ready Reaction kit (Applied Biosystems). After a magnesium sulphate (MgSO4) clean-up, the sequencing fragments were run on an ABI 3730 DNA Sequencer (Applied Biosystems). The sequences were visualised and edited using Geneious Pro version 5.0. Out of the 434 base-pair fragments amplified across the seven individuals, we identified only two polymorphic sites. As a result and for ease of presentation, we only include here a figure showing an 80 base-pair fragment including the two polymorphic sites for three of the six individuals (178 bp-255 bp). The remaining three individuals' mtDNA profile is available in the supporting online information <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012299#pone.0012299.s002\" target=\"_blank\">Figure S2</a>. The full sequences for all six individuals have been made available on GenBank (HM581690-HM58701).</p>", "links"=>[], "tags"=>["dna", "individuals"], "article_id"=>503981, "categories"=>["Genetics", "Ecology"], "users"=>["Céline H. Frère", "Ewa Krzyszczyk", "Eric M. Patterson", "Sue Hunter", "Alison Ginsburg", "Janet Mann"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0012299.g003", "stats"=>{"downloads"=>0, "page_views"=>4, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Mitochondrial_DNA_profile_of_blow_and_blood_from_individuals_1_to_3_/503981", "title"=>"Mitochondrial DNA profile of blow and blood from individuals 1 to 3.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2010-08-25 01:06:21"}

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F1000Prime