ONTbarcoder (available from https://github.com/asrivathsan/ONTbarcoder) allows you to call DNA barcodes fom Nanopore sequence data. It allows the user to ONTbarcoder is designed to handle dual tagged amplicon pools. In the Conventional mode, it accepts a fastq file obtained after base-calling nanopore data and a demultiplexing file that specifies the tag combination for every sample. The original paper describes in detail how such tagged amplicons can be obtained and the pipeline implemented in ONTbarcoder. In the real-time barcoding mode, described here, it accepts either fastq or raw files to call barcodes real-time, given user-defined demultiplexing information.
ONTbarcoder has been optimized for protein coding gene like COI. While there are ways to obtain consensus for length variable non coding genes, this has not been extensively tested.
Please check the releases in this repository: https://github.com/asrivathsan/ONTbarcoder/releases. The ONTBarcoder_manual.pdf details more detailed guidelines for troubleshooting in case of issues. It is installed by unzipping the folder with the version of the program that supports the operating system on your computer. The folders are available from releases in Github. For MacOS, an .app bundle has been created. Kindly download the bundle relevant to the OS version. For further notes on MacOS compatibility and permissions: see the MacOS section. Currently the software has been tested in OSX 12.5 and 13.5.
Current version: Unzip ONTbarcoder2.3.0_win_exe.zip and double click on the executable. In default mode the output files are stored in the same directory as the executable. Windows has a character limit of path length as 256 characters. Therefore, please avoid placing the software deep inside the directory structure. Avoid spaces in folder names
Older version: Unzip ONTBarcoder_0.1.9_exe.win-amd64-2.7.zip and double click on the executable. In default mode the output files are stored in the same directory as the executable
Use ONTbarcoder2.3.0_OSX.zip. In some tests, it was noted that permissions can be an issue in Mac. Transferring the app to Applications folder and running worked smoothly. But you may have to right click and select open
For older version of ONTbarcoder: Use the DMGs for the version corresponding to your Mac. We recommend using the upgraded Mac (macOS 11) and corresponding dmg file ONTbarcoder_0.1.9_OS11.dmg. We have also compiled for older versions; ONTbarcoder_0.1.9_OSX10_13.dmg has been tested in macOS 10.13 and 10.12. We would suggesting using this in first pass for 10.14/15
In case DMG files give problems due to permission, you may use the standalone.tgz: macOS 11, macOS 10.13 files. These will contain numerous executables and the software should be run using ONTbarcoder_multiprocessing.
Current version: Uncompress the tgz archive ONTbarcoder2.3.0_linux.tgz Start the software from the terminal as by
cd directory_containing_ONTbarcoderfiles
./ONTbarcoder2Older version: Uncompress the tgz archive ONTbarcoder_multiprocessing_0.1.9_linux.tgz.
There are two types of analysis implemented in the software
Please use the Dataset_accessibility_and_descriptions file in the repository for access to test datasets. We recommend use of datasets A,B,C for initial testing before running the larger dataset.