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Area of Science:

  • Genomics and Bioinformatics
  • Environmental DNA (eDNA) Metabarcoding
  • Portable Sequencing Technologies

Background:

  • The MinION nanopore sequencer offers portable DNA sequencing for field applications.
  • Current MinION DNA barcoding workflows have limited sample capacity (n ≤ 10).
  • In-situ species identification is crucial for biodiversity monitoring in remote environments.

Purpose of the Study:

  • To enhance the sample processing capability of portable MinION DNA barcoding.
  • To develop and validate a workflow for simultaneous processing of 32 samples.
  • To assess the performance of the R10.3 nanopore flow cell for DNA barcoding.

Main Methods:

  • Assembled a portable sequencing setup using BentoLab and MinION.
  • Developed a workflow for processing up to 32 DNA samples concurrently.
  • Conducted onboard-vessel DNA barcoding of marine samples collected at Sisters' Islands Marine Park, Singapore.

Main Results:

  • Generated 105 MinION barcodes in under 9 hours, including 19 from fresh metazoans.
  • Demonstrated a viable setup for enhanced portable DNA barcoding capabilities.
  • Achieved ~99.9% accuracy for R10.3 nanopore barcodes compared to Illumina references, with 80% showing zero base ambiguities.

Conclusions:

  • The developed portable workflow significantly increases MinION DNA barcoding capacity for field applications.
  • The R10.3 nanopore flow cell offers improved accuracy and homopolymer resolution for DNA barcoding.
  • This enhanced setup is highly recommended for fortifying portable DNA barcoding efforts in marine and other remote environments.