Portable sequencing as a teaching tool in conservation and biodiversity research

Watsa, Mrinalini; Erkenswick, Gideon; Pomerantz, Aaron; Prost, Stefan

doi:10.1371/journal.pbio.3000667

Cited by 38 publications

(36 citation statements)

References 20 publications

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“…Marine ecosystems, which are dynamic and subject to fluctuations in tides, reproductive cycles, seasonal disasters and stochastic events, require the generation of actionable data in a timelier manner. For such applications, we developed the fieldable eDNA sequencing (FeDS) kit which utilizes the field-deployable MinION sequencing platform Oxford Nanopore Technologies (ONT) [reviewed in Jain et al (2016); Pomerantz et al (2018), Krehenwinkel et al (2019); Watsa et al (2020)].…”

Section: Introductionmentioning

confidence: 99%

Fieldable Environmental DNA Sequencing to Assess Jellyfish Biodiversity in Nearshore Waters of the Florida Keys, United States

et al. 2021

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Recent advances in molecular sequencing technology and the increased availability of fieldable laboratory equipment have provided researchers with the opportunity to conduct real-time or near real-time gene-based biodiversity assessments of aquatic ecosystems. In this study, we developed a workflow and portable kit for fieldable environmental DNA sequencing (FeDS) and tested its efficacy by characterizing the breadth of jellyfish (Medusozoa) taxa in the coastal waters of the Upper and Lower Florida Keys. Environmental DNA was isolated from seawater collection events at eight sites and samples were subjected to medusozoan 16S rRNA gene and metazoan mitochondrial cytochrome oxidase 1 gene profiling via metabarcoding onsite. In total, FeDS yielded 175,326 processed sequence reads providing evidence for 53 medusozoan taxa. Our most salient findings revealed eDNA from: (1) two venomous box jellyfish (Cubozoa) species, including taxa whose stings cause the notorious Irukandji envenomation syndrome; (2) two species of potentially introduced stalked jellyfish (Staurozoa); and (3) a likely cryptic species of upside-down jellyfish (Scyphozoa). Taken together, the results of this study highlight the merits of FeDS in conducting biodiversity surveys of endemic and introduced species, and as a potential tool for assessing envenomation and/or conservation-related threats.

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Section: Introductionmentioning

confidence: 99%

Fieldable Environmental DNA Sequencing to Assess Jellyfish Biodiversity in Nearshore Waters of the Florida Keys, United States

et al. 2021

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“…The MinION’s potential as an effective teaching tool was recognized early on and it has been used in classroom settings ( Salazar et al , 2020 ; Zaaijer and Erlich 2016 ; Zeng and Martin 2017 ) as well as in the field ( Watsa et al , 2020 ). The presented study illustrates that inexpensive nanopore-based sequencing along with published short-read data, as well as memory and run-time efficient genome assembly tools offer great potential to generate high quality chromosome-level assemblies, even of more complex vertebrate genomes, as part of university courses.…”

Section: Resultsmentioning

confidence: 99%

Improving the Chromosome-Level Genome Assembly of the Siamese Fighting Fish (Betta splendens) in a University Master’s Course

Prost

Petersen

Grethlein

et al. 2020

G3 Genes|Genomes|Genetics

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Ever decreasing costs along with advances in sequencing and library preparation technologies enable even small research groups to generate chromosome-level assemblies today. Here we report the generation of an improved chromosome-level assembly for the Siamese fighting fish (Betta splendens) that was carried out during a practical university master’s course. The Siamese fighting fish is a popular aquarium fish and an emerging model species for research on aggressive behavior. We updated the current genome assembly by generating a new long-read nanopore-based assembly with subsequent scaffolding to chromosome-level using previously published Hi-C data. The use of ∼35x nanopore-based long-read data sequenced on a MinION platform (Oxford Nanopore Technologies) allowed us to generate a baseline assembly of only 1,276 contigs with a contig N50 of 2.1 Mbp, and a total length of 441 Mbp. Scaffolding using the Hi-C data resulted in 109 scaffolds with a scaffold N50 of 20.7 Mbp. More than 99% of the assembly is comprised in 21 scaffolds. The assembly showed the presence of 96.1% complete BUSCO genes from the Actinopterygii dataset indicating a high quality of the assembly. We present an improved full chromosome-level assembly of the Siamese fighting fish generated during a university master’s course. The use of ∼35× long-read nanopore data drastically improved the baseline assembly in terms of continuity. We show that relatively in-expensive high-throughput sequencing technologies such as the long-read MinION sequencing platform can be used in educational settings allowing the students to gain practical skills in modern genomics and generate high quality results that benefit downstream research projects.

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“…For example, the one described by (Sahlin, Lim et al 2021) involves minibar/qcat and nanofilt, while NGSpeciesID relies on isONclust SPOA, Parasail, and optionally, Medaka (Daily 2016, Krehenwinkel, Pomerantz et al 2019, Sahlin and Medvedev 2020). These dependencies and complexities meant that Watsa et al (2020) recommended bioinformatics training before MinION barcoding could be used in schools (e.g., training in UNIX command-line) and additionally required the installation of several software tools onto the teaching computers. Neither is needed for ONTbarcoder, which runs on a regular laptop and has been extensively tested (>4000 direct comparisons to Sanger and Illumina barcodes).…”

Section: Discussionmentioning

confidence: 99%

MinION barcodes: biodiversity discovery and identification by everyone, for everyone

Srivathsan

Lee

Katoh

et al. 2021

Preprint

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DNA barcodes are a useful tool for discovering, understanding, and monitoring biodiversity. This is critical at a time when biodiversity loss is a major problem for many countries. However, widespread adoption of barcoding programs requires the process to be cost-effective and simple to apply. We here present a workflow that satisfies these conditions. It was developed via 'innovation through subtraction' and thus requires minimal lab equipment, can be learned within days, reduces the barcode sequencing cost to <10 cents, and allows fast turnaround from specimen to sequence by using the real-time sequencer MinION. We first describe cost-effective and rapid procedures in a comprehensive workflow for obtaining tagged amplicons. We then demonstrate how a portable MinION device can be used for real-time sequencing of tagged amplicons in many settings (field stations, biodiversity labs, citizen science labs, schools). Small projects can use the flow cell dongle ('Flongle') while large projects can rely on MinION flow cells that can be stopped and re-used after collecting sufficient data for a given project. We also provide amplicon coverage recommendations that are based on several runs of MinION flow cells (R10.3) involving >24,000 specimen barcodes, which suggest that each run can generate >10,000 barcodes. Additionally, we present a novel software, ONTbarcoder, that overcomes the bioinformatics challenges posed by the sequencing errors of MinION reads. This software is compatible with Windows10, Macintosh, and Linux, has a graphical user interface (GUI), and can generate thousands of barcodes on a standard laptop within hours based on two input files (FASTQ, demultiplexing file). Next, we document that MinION barcodes are virtually identical to Sanger and Illumina barcodes for the same specimens (>99.99%). Lastly, we demonstrate how rapidly MinION data have improved by comparing the performance of sequential flow cell generations. We overall assert that barcoding with MinION is the way forward for government agencies, universities, museums, and schools because it combines low consumable and capital cost with scalability. Biodiversity loss is threatening the planet and the use of MinION barcodes will help with enabling an army of researchers and citizen scientists, which is necessary for effective biodiversity discovery and monitoring.

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Portable sequencing as a teaching tool in conservation and biodiversity research

Cited by 38 publications

References 20 publications

Fieldable Environmental DNA Sequencing to Assess Jellyfish Biodiversity in Nearshore Waters of the Florida Keys, United States

Fieldable Environmental DNA Sequencing to Assess Jellyfish Biodiversity in Nearshore Waters of the Florida Keys, United States

Improving the Chromosome-Level Genome Assembly of the Siamese Fighting Fish (Betta splendens) in a University Master’s Course

MinION barcodes: biodiversity discovery and identification by everyone, for everyone

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