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1000 Titel
  • An efficient and robust laboratory workflow and tetrapod database for larger scale environmental DNA studies
1000 Autor/in
  1. Axtner, Jan |
  2. Crampton-Platt, Alex |
  3. Hörig, Lisa A. |
  4. Mohamed, Azlan |
  5. Xu, Charles |
  6. Yu, Douglas W. |
  7. Wilting, Andreas |
1000 Erscheinungsjahr 2019
1000 LeibnizOpen
1000 Publikationstyp
  1. Artikel |
1000 Online veröffentlicht
  • 2019-04-13
1000 Erschienen in
1000 Quellenangabe
  • 8(4):giz029
1000 FRL-Sammlung
1000 Copyrightjahr
  • 2019
1000 Lizenz
1000 Verlagsversion
  • https://doi.org/10.1093/gigascience/giz029 |
  • https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6461710/ |
1000 Ergänzendes Material
  • https://academic.oup.com/gigascience/article/8/4/giz029/5450733#supplementary-data |
1000 Publikationsstatus
1000 Begutachtungsstatus
1000 Sprache der Publikation
1000 Abstract/Summary
  • BACKGROUND: The use of environmental DNA for species detection via metabarcoding is growing rapidly. We present a co-designed lab workflow and bioinformatic pipeline to mitigate the 2 most important risks of environmental DNA use: sample contamination and taxonomic misassignment. These risks arise from the need for polymerase chain reaction (PCR) amplification to detect the trace amounts of DNA combined with the necessity of using short target regions due to DNA degradation. FINDINGS: Our high-throughput workflow minimizes these risks via a 4-step strategy: (i) technical replication with 2 PCR replicates and 2 extraction replicates; (ii) using multi-markers (12S,16S,CytB); (iii) a “twin-tagging,” 2-step PCR protocol; and (iv) use of the probabilistic taxonomic assignment method PROTAX, which can account for incomplete reference databases. Because annotation errors in the reference sequences can result in taxonomic misassignment, we supply a protocol for curating sequence datasets. For some taxonomic groups and some markers, curation resulted in >50% of sequences being deleted from public reference databases, owing to (i) limited overlap between our target amplicon and reference sequences, (ii) mislabelling of reference sequences, and (iii) redundancy. Finally, we provide a bioinformatic pipeline to process amplicons and conduct PROTAX assignment and tested it on an invertebrate-derived DNA dataset from 1,532 leeches from Sabah, Malaysia. Twin-tagging allowed us to detect and exclude sequences with non-matching tags. The smallest DNA fragment (16S) amplified most frequently for all samples but was less powerful for discriminating at species rank. Using a stringent and lax acceptance criterion we found 162 (stringent) and 190 (lax) vertebrate detections of 95 (stringent) and 109 (lax) leech samples. CONCLUSIONS: Our metabarcoding workflow should help research groups increase the robustness of their results and therefore facilitate wider use of environmental and invertebrate-derived DNA, which is turning into a valuable source of ecological and conservation information on tetrapods.
1000 Sacherschließung
lokal leeches
lokal environmental DNA
lokal invertebrate-derived -DNA
lokal metabarcoding
1000 Fächerklassifikation (DDC)
1000 Liste der Beteiligten
  1. https://orcid.org/0000-0003-1269-5586|https://frl.publisso.de/adhoc/uri/Q3JhbXB0b24tUGxhdHQsIEFsZXg=|https://frl.publisso.de/adhoc/uri/SMO2cmlnLCBMaXNhIEEu|https://orcid.org/0000-0003-3788-4383|https://orcid.org/0000-0001-6779-8879|https://frl.publisso.de/adhoc/uri/WXUsIERvdWdsYXMgVy4=|https://orcid.org/0000-0001-5073-9186
1000 Label
1000 Förderer
  1. Bundesministerium für Bildung und Forschung |
  2. Leibniz-Institut für Zoo- und Wildtierforschung |
  3. MEME Erasmus Mundus Programme in Evolutionary Biology |
  4. University of Groningen |
  5. National Natural Science Foundation of China |
  6. University of Science and Technology of China |
  7. Chinese Academy of Sciences |
  8. Ministry of Science and Technology of the People's Republic of China |
1000 Fördernummer
  1. 01LN1301A
  2. -
  3. -
  4. -
  5. 41661144002; 31670536; 31400470; 31500305
  6. QYZDY-SSW-SMC024; GJHZ1754; XDA20050202; XDB31000000
  7. 2012FY110800
  8. -
1000 Förderprogramm
  1. -
  2. -
  3. -
  4. Groningen University Fund; Marco Polo Fund
  5. -
  6. Key Research Program of Frontier Sciences
  7. -
  8. -
1000 Dateien
1000 Förderung
  1. 1000 joinedFunding-child
    1000 Förderer Bundesministerium für Bildung und Forschung |
    1000 Förderprogramm -
    1000 Fördernummer 01LN1301A
  2. 1000 joinedFunding-child
    1000 Förderer Leibniz-Institut für Zoo- und Wildtierforschung |
    1000 Förderprogramm -
    1000 Fördernummer -
  3. 1000 joinedFunding-child
    1000 Förderer MEME Erasmus Mundus Programme in Evolutionary Biology |
    1000 Förderprogramm -
    1000 Fördernummer -
  4. 1000 joinedFunding-child
    1000 Förderer University of Groningen |
    1000 Förderprogramm Groningen University Fund; Marco Polo Fund
    1000 Fördernummer -
  5. 1000 joinedFunding-child
    1000 Förderer National Natural Science Foundation of China |
    1000 Förderprogramm -
    1000 Fördernummer 41661144002; 31670536; 31400470; 31500305
  6. 1000 joinedFunding-child
    1000 Förderer University of Science and Technology of China |
    1000 Förderprogramm Key Research Program of Frontier Sciences
    1000 Fördernummer QYZDY-SSW-SMC024; GJHZ1754; XDA20050202; XDB31000000
  7. 1000 joinedFunding-child
    1000 Förderer Chinese Academy of Sciences |
    1000 Förderprogramm -
    1000 Fördernummer 2012FY110800
  8. 1000 joinedFunding-child
    1000 Förderer Ministry of Science and Technology of the People's Republic of China |
    1000 Förderprogramm -
    1000 Fördernummer -
1000 Objektart article
1000 Beschrieben durch
1000 @id frl:6419442.rdf
1000 Erstellt am 2020-03-26T13:40:33.074+0100
1000 Erstellt von 288
1000 beschreibt frl:6419442
1000 Bearbeitet von 25
1000 Zuletzt bearbeitet 2020-03-31T07:59:42.258+0200
1000 Objekt bearb. Tue Mar 31 07:59:41 CEST 2020
1000 Vgl. frl:6419442
1000 Oai Id
  1. oai:frl.publisso.de:frl:6419442 |
1000 Sichtbarkeit Metadaten public
1000 Sichtbarkeit Daten public
1000 Gegenstand von

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