Bulk FAST5 file for 16S Oxford Nanopore Barcoded Sequencing
Authors/Creators
-
1.
Indian Institute of Technology Delhi
Description
Real-time taxonomic classification of nanopore amplicon sequencing data enables rapid insights into microbial communities, with applications in clinical diagnostics, environmental monitoring, and outbreak surveillance. However, bridging the gap between long-read data and interpretable results often requires specialized bioinformatics expertise, highlighting the need for integrated, user-friendly software that combines live data acquisition with downstream microbiome analysis.
To address this need, we developed NANOTAXI, a fully automated Shiny-based graphical user interface (GUI) for classifying barcoded 16S rRNA gene sequences generated by Oxford Nanopore sequencing. NANOTAXI supports four taxonomic classifiers integrated with five reference databases, offering flexible classification strategies tailored to user requirements and computational resources.
The dataset accompanying this submission comprises a bulk FAST5 file from a Nanopore sequencing run of barcoded 16S rRNA genes. This bulk FAST5 file enables users to replay the sequencing run using Oxford Nanopore Technologies' MinKNOW software, allowing the testing of novel methods or analytical techniques without performing additional sequencing experiments, thereby saving both time and resources.
Methods
Reference strains from ATCC were subcultured on blood agar at 37°C. Single colonies were grown overnight in LB broth to an OD600 of 0.8–1.2. Genomic DNA (gDNA) was extracted using the QIAamp DNA Mini Kit (Qiagen) and analysed using a NanoDrop.
Four mock communities (A-D) were constructed to simulate mixed populations. gDNA from E. coli, P. aeruginosa, S. aureus, and the ZymoBIOMICS community was mixed equimolarly in biological triplicates (barcodes 01-03). In the remaining samples, one species dominated (80% DNA), with the other two at 10% each. Three biological replicates of the ZymoBIOMICS mock community were also included (barcodes 13-15).
Libraries were prepared using the 16S Barcoding Kit 24 V14 (ONT) according to the standard protocol. 16S rRNA gene amplification and barcoding, magnetic bead purification, equimolar pooling, and sequencing on a MinION Mk1D with an R10.4.1 flow cell.
Files
Files
(62.6 GB)
| Name | Size | Download all |
|---|---|---|
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md5:7cfd88c13b7a6f754abd6402c609fb71
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62.6 GB | Download |
Additional details
Software
- Repository URL
- https://github.com/Nirmal2310/NANOTAXI/tree/main
- Programming language
- R , Python , Shell
- Development Status
- Active