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Scaling up DNA metabarcoding for freshwater macrozoobenthos monitoring

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RT @VascoElbrecht: @eDNAScience @QIAGEN @dirch3 @BraukmannThomas @BiodiversityIn1 Thanks :D very happy with the 96 plate extractions and pc…
@eDNAScience @QIAGEN @dirch3 @BraukmannThomas @BiodiversityIn1 Thanks :D very happy with the 96 plate extractions and pcr. There is actually a sophisticated sample layout behind this, allowing to detect contamination and other things that might go wrong. See @PeerJPreprints: https://t.co/tIDCoCtDxO ☺️
@KristenFerns Maybe you find this video helpful concerning fusion primer design : ) https://t.co/k6q0EMdfW6 here is also the related @PeerJPreprints https://t.co/tIDCoCtDxO please let me know if i can help with anything #metabarcoding related : )
Getting things ready for 96 well plate extractions! =) Will use single tubes for digestions as the powder does too quickly disperse by static charging. But then switching to @Quiagen plate extractions after that. Scaling up #metabarcoding! @PeerJPreprints: https://t.co/tIDCoCc29e https://t.co/HhxiKqIBCV
@Gnarly_Larvae @dirch3 Yes, that’s probably the reason. The fusion primer system works brilliantly with 5% phiX, heterogeneity spacers (also used at in-line barcodes), and parallel sequencing. See also https://t.co/tIDCoCtDxO for recommendations : ) please let me know if you have any questions!
Hundreds of grinding chambers have arrived and @QIAGEN plate extraction kits for #metabarcoding at large scale! More information on scaling things up, here: https://t.co/tIDCoCc29e https://t.co/ZmCTAj0ZJh
A bit more shifting has to be done till for the 21 #metabarcoding primer sets to be tagged and tested = ) A Levenshtein distance of 1-2 bp means the primers are too similar -> Tagswitching can happen! scripts and more info in this related @PeerJPreprints: https://t.co/tIDCoCc29e https://t.co/KOm08QCFWL
RT @VascoElbrecht: Update of our scaling up #metabarcoding @PeerJPreprints is online! =) some minor improvements + new example sample layou…
98 days ago
RT @VascoElbrecht: Update of our scaling up #metabarcoding @PeerJPreprints is online! =) some minor improvements + new example sample layou…
Update of our scaling up #metabarcoding @PeerJPreprints is online! =) some minor improvements + new example sample layout that does use a unique primer combination for each replicate. Thanks to Arne Beermann from @leeselab for the suggestion! https://t.co/tIDCoCc29e https://t.co/9LUp8RyKyg
Scaling up DNA metabarcoding for freshwater macrozoobenthos monitoring https://t.co/we2ltjAECx
99 days ago
Scaling up DNA metabarcoding for freshwater macrozoobenthos monitoring https://t.co/R4YV4H5oPF
179 days ago
RT @VascoElbrecht: Updated @PeerJPreprints! Finally: better fusion #metabarcoding BF2+BR2 primer versions now available (the P7 we used so…
RT @VascoElbrecht: Updated @PeerJPreprints! Finally: better fusion #metabarcoding BF2+BR2 primer versions now available (the P7 we used so…
RT @metabar_papers: Scaling up DNA metabarcoding for freshwater macrozoobenthos monitoring https://t.co/h1E32gm4FB
180 days ago
RT @metabar_papers: Scaling up DNA metabarcoding for freshwater macrozoobenthos monitoring https://t.co/h1E32gm4FB
180 days ago
Scaling up DNA metabarcoding for freshwater macrozoobenthos monitoring https://t.co/fuVTWZNdUV
Scaling up DNA metabarcoding for freshwater macrozoobenthos monitoring https://t.co/h1E32gm4FB
Updated @PeerJPreprints! Finally: better fusion #metabarcoding BF2+BR2 primer versions now available (the P7 we used so far at @leeselab were not ideal, but did still work). I also did add a few hints on how to set things up with indexing for 1000+ samples https://t.co/tIDCoCc29e https://t.co/gFnTNxvO8g
@MatSeymour @BangorUniPress @alpineedna @NERCscience Hey mat, great. Write me a pm if you need details/ me to look over things. There is also a new (untested) BF3 primer i can send you under the counter. Also https://t.co/YIxDCZ9cG1 might be helpful. Minor update soon, with tagging 1000+ samples and updated P5 (cured one is false).
RT @VascoElbrecht: here is your weekend read: Scaling up #metabarcoding for monitoring macroinvertebrates! Practical guide to @illumina fus…
RT @VascoElbrecht: here is your weekend read: Scaling up #metabarcoding for monitoring macroinvertebrates! Practical guide to @illumina fus…
RT @VascoElbrecht: here is your weekend read: Scaling up #metabarcoding for monitoring macroinvertebrates! Practical guide to @illumina fus…
RT @VascoElbrecht: Updated version of our workflow manuscript now available @PeerJPreprints: Further improved tags and added analysis tools…
Updated version of our workflow manuscript now available @PeerJPreprints: Further improved tags and added analysis tools for Levenshtein distance to catch indels that might affect the tags! https://t.co/tIDCoCc29e
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Supplemental Information

Figure S1: Newly developed fusion primer sets (BF2+BR2), suitable for tagging 288 individual wells

DOI: 10.7287/peerj.preprints.3456v4/supp-1

Figure S2: Base composition of the inline tagging region

DOI: 10.7287/peerj.preprints.3456v4/supp-2

Figure S3: Hamming distance between tags for all fusion primers

DOI: 10.7287/peerj.preprints.3456v4/supp-3

Figure S4: Levenshtein distance between tags for all fusion primers

DOI: 10.7287/peerj.preprints.3456v4/supp-4

Script S1: R script used for randomly generating inline barcodes for the given primer sets (includes visualization, as shown in Figure S3 and Figure 3)

DOI: 10.7287/peerj.preprints.3456v4/supp-5

Table S1: Table providing an overview of proposed tagging combinations (as shown in Figure 3)

DOI: 10.7287/peerj.preprints.3456v4/supp-6

Manuscript file for providing feedback

(Pleas use track changes)

DOI: 10.7287/peerj.preprints.3456v4/supp-7

Additional Information

Competing Interests

The authors declare that they have no competing interests.

Author Contributions

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

Dirk Steinke authored or reviewed drafts of the paper.

Data Deposition

The following information was supplied regarding data availability:

primer sequences and scripts to generated them are provided as supporting information

Funding

V.E. and D.S. are supported by funding through the Canada First Research Excellence Fund. This work represents a contribution to the ‘Food From Thought’ research program and the EU Action ‘DNAqua-Net’ (CA15219). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.


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