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Validation of COI metabarcoding primers for terrestrial arthropods

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12 days ago
RT @VascoElbrecht: Interested in #metabarcoding primer validations? You can suggest your self as a reviewer of our @PeerJPreprints https://…
18 days ago
RT @VascoElbrecht: Interested in #metabarcoding primer validations? You can suggest your self as a reviewer of our @PeerJPreprints https://…
19 days ago
RT @VascoElbrecht: Interested in #metabarcoding primer validations? You can suggest your self as a reviewer of our @PeerJPreprints https://…
RT @VascoElbrecht: Interested in #metabarcoding primer validations? You can suggest your self as a reviewer of our @PeerJPreprints https://…
20 days ago
RT @VascoElbrecht: Interested in #metabarcoding primer validations? You can suggest your self as a reviewer of our @PeerJPreprints https://…
RT @VascoElbrecht: Interested in #metabarcoding primer validations? You can suggest your self as a reviewer of our @PeerJPreprints https://…
RT @VascoElbrecht: Interested in #metabarcoding primer validations? You can suggest your self as a reviewer of our @PeerJPreprints https://…
Interested in #metabarcoding primer validations? You can suggest your self as a reviewer of our @PeerJPreprints https://t.co/BGXSm1C1w1
RT @VascoElbrecht: Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) i…
RT @VascoElbrecht: Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) i…
Validation of COI metabarcoding primers for terrestrial arthropods https://t.co/ESUQXMmoDY
RT @windowmoon: Validation of COI metabarcoding primers for terrestrial arthropods https://t.co/AH6VHfbiQ3 昆虫COIメタバーコーディング用プライマーのベンチマーク。
RT @windowmoon: Validation of COI metabarcoding primers for terrestrial arthropods https://t.co/AH6VHfbiQ3 昆虫COIメタバーコーディング用プライマーのベンチマーク。
RT @windowmoon: Validation of COI metabarcoding primers for terrestrial arthropods https://t.co/AH6VHfbiQ3 昆虫COIメタバーコーディング用プライマーのベンチマーク。
Validation of COI metabarcoding primers for terrestrial arthropods https://t.co/AH6VHfbiQ3 昆虫COIメタバーコーディング用プライマーのベンチマーク。
45 days ago
RT @VascoElbrecht: Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) i…
46 days ago
RT @VascoElbrecht: Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) i…
RT @VascoElbrecht: Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) i…
46 days ago
RT @VascoElbrecht: Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) i…
RT @VascoElbrecht: Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) i…
RT @VascoElbrecht: Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) i…
Working on an update to our #metabarcoding primer test @PeerJPreprints! Taxa recovery for well working primers (green) is very consistent, no Order bias. Thanks 2 @M_Hajibabaei for the suggestion. Want to give feedback as well? Read the preprint here: https://t.co/BGXSm1C1w1
RT @VascoElbrecht: Doing #metabarcoding on insects? @ento_ben’s cat chewbacca demands you use the new BF3+BR2! You can download it here: ht…
RT @VascoElbrecht: Doing #metabarcoding on insects? @ento_ben’s cat chewbacca demands you use the new BF3+BR2! You can download it here: ht…
RT @VascoElbrecht: Doing #metabarcoding on insects? @ento_ben’s cat chewbacca demands you use the new BF3+BR2! You can download it here: ht…
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Supplemental Information

Figure S1: Mock community composition

DOI: 10.7287/peerj.preprints.27801v2/supp-1

Figure S2: Sequence alignment for 29 insect orders, including primer binding annotations. The alignment was used for primer development

DOI: 10.7287/peerj.preprints.27801v2/supp-2

Figure S3: Evaluation of Levenshtein distances for fusion primers used to metabarcode the 21 primer sets

DOI: 10.7287/peerj.preprints.27801v2/supp-3

Figure S4: Fusion primers used to metabarcode the 21 primer sets

DOI: 10.7287/peerj.preprints.27801v2/supp-4

Figure S5: Fusion primers used for gradient metabarcoding

DOI: 10.7287/peerj.preprints.27801v2/supp-5

Figure S6: Evaluation of Levenshtein distances for fusion primers used in gradient PCR

DOI: 10.7287/peerj.preprints.27801v2/supp-6

Figure S7: Gradient PCR gels for the initial 36 primer combinations

DOI: 10.7287/peerj.preprints.27801v2/supp-7

Figure S8: Amplicon concentration of the 36 primer sets after the first gradient PCR test

DOI: 10.7287/peerj.preprints.27801v2/supp-8

Figure S9: Sequencing depth for the mock community metabarcoding run

DOI: 10.7287/peerj.preprints.27801v2/supp-9

Figure S10: Distribution of read lengths after paired end merging for the mock community metabarcoding run

DOI: 10.7287/peerj.preprints.27801v2/supp-10

Figure S11: Rarefaction curves showing taxon recovery for the mock sample with different primer sets

DOI: 10.7287/peerj.preprints.27801v2/supp-11

Figure S12: Heat map showing taxon recovery for the mock sample with different primer sets

DOI: 10.7287/peerj.preprints.27801v2/supp-12

Figure S13: Principal component analysis of the metabarcoding OTU table for the mock community metabarcoding run

DOI: 10.7287/peerj.preprints.27801v2/supp-13

Figure S14: Jaccard similarity and Bray-Curtis distance based on taxa recovered from the mock community metabarcoding run

DOI: 10.7287/peerj.preprints.27801v2/supp-14

Figure S15: Plot showing the similarity between taxon recovery at 46 °C with primers of both the mock community metabarcoding and the final gradient metabarcoding run

DOI: 10.7287/peerj.preprints.27801v2/supp-15

Figure S16: Heat map showing taxon recovery with four primer sets at different annealing temperatures (40 - 56 °C)

DOI: 10.7287/peerj.preprints.27801v2/supp-16

Figure S17: Distribution of read length after paired end merging for the final gradient run

DOI: 10.7287/peerj.preprints.27801v2/supp-17

Figure S18: Heat map showing taxon recovery for the Malaise trap metabarcoding run with 21 primer sets

DOI: 10.7287/peerj.preprints.27801v2/supp-18

Figure S19: Rarefaction curves showing taxon recovery for the Malaise trap metabarcoding run with different primer sets

DOI: 10.7287/peerj.preprints.27801v2/supp-19

Scripts S1: R scripts used for bioinformatics processing, figure generation and statistical analysis

DOI: 10.7287/peerj.preprints.27801v2/supp-20

Table S1: Raw OTU table for both the 21 primer and the gradient metabarcoding run, as well as details on mock sample composition

DOI: 10.7287/peerj.preprints.27801v2/supp-21

Table S2: Primer sequences and primer combinations evaluated in this study

DOI: 10.7287/peerj.preprints.27801v2/supp-22

Table S3: NCBI SRA accession numbers for demultiplexed samples and raw MiSeq sequencing files

DOI: 10.7287/peerj.preprints.27801v2/supp-23

Manuscript file, for you to download and provide feedback using track changes!

DOI: 10.7287/peerj.preprints.27801v2/supp-24

Additional Information

Competing Interests

The authors declare that they have no competing interests.

Author Contributions

Vasco Elbrecht conceived and designed the experiments, performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, prepared figures and/or tables, authored or reviewed drafts of the paper.

Thomas WA Braukmann performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, authored or reviewed drafts of the paper.

Natalia V Ivanova contributed reagents/materials/analysis tools.

Sean WJ Prosser contributed reagents/materials/analysis tools.

Mehrdad Hajibabaei contributed reagents/materials/analysis tools.

Michael Wright contributed reagents/materials/analysis tools.

Evgeny V Zakharov contributed reagents/materials/analysis tools.

Paul DN Hebert authored or reviewed drafts of the paper.

Dirk Steinke conceived and designed the experiments, authored or reviewed drafts of the paper.

DNA Deposition

The following information was supplied regarding the deposition of DNA sequences:

Raw sequence data is available on the NCBI SRA archive; accession SRX4908948 provides data for the mock community, accession SRX5175597 for the Malaise sample, and accession SRX4908947 for the gradient PCR experiment.

Data Deposition

The following information was supplied regarding data availability:

Scripts are available as supporting informations, the metabarcoding pipeline is available on GitHub https://github.com/VascoElbrecht/JAMP

Funding

This work was funded by Food from Thought. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.


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