sc2-sequencing/ont

Oxford Nanopore Technologies

These scripts are for use with a GridION or its output file, to make the files and folder structures correspond more closely to those required by downstream bioinformatics programs. Note that at the time of writing, the GridION runs Ubuntu 16.04 LTS, and mounts USB drives in the /data folder, and these scripts contain some "faults" to work around these idiosyncrasies (i.e. they will not pass in ShellCheck).

Usage

ont-transfer

ont-transfer [-s 1|6|12] FAP_FOLDER

Transfers all of the passing FAST5 and FASTQ files to the first available USB drive, skipping empty barcodes based on the number of the first skip passed through the -s parameter, consolidating all files of the same type into one folder for easier sorting and uploading, and concatenating and compressing FASTQ files into one per barcode.

ont-rename.jl

To be run with Julia.

julia --project=PATH_TO_ONT_FOLDER PATH_TO_ONT-RENAME.jl

In practice, this looks like

julia --project=C:\Users\MillironX\ont C:\Users\MillironX\ont\ont-rename.jl

ont-demux (deprecated)

ont-demux FAP_FOLDER

This functionality is now wrapped into ont-Transfer, which can produce the same results using far fewer resources and in far less time.

Consolidates all FAST5s into a single gzipped FASTQ using ONT's guppy_barcoder. Note that this is for MinKNOW's default settings of 4000 reads/file. It is highly recommended to set the "Number of Reads per File" entry in MinKNOW to 0, and checking "Compress FASTQs" to produce nearly the same results (this functionality has been taken out of recent updates to MinKNOW).

Installation

ont-transfer and ont-demux

Requirements

• git
• guppy (Get from ONT Community)
• GNU Parallel

See ONT for guppy installation instructions. Others may be installed by opening a terminal and running

sudo apt-get install git parallel -y

Instructions

Open a terminal (Ctrl+Alt+T), and use the following commands

git clone https://github.com/MillironX/sc2-sequencing
mkdir ~/bin
echo "export PATH=$PATH:$HOME/bin" >> ~/.bashrc
cp sc2-sequending/ont/* ~/bin
chmod +x ~/bin/*
source ~/.bashrc

ont-rename

These scripts require Julia.

Windows install instructions (I will assume Linux users can adapt):

1. Download and install Julia
   1. Select "Add Julia to PATH" as an option
   2. All other default options will work
2. Download the Repo's ZIP file (click green button on upper-right)
3. Press Win+R, type %USERPROFILE% and press Enter
4. Copy the ont folder from the ZIP file into the newly opened folder
5. Press Win+X, then I to open a new PowerShell window
6. Type julia --project=$env:USERPROFILE\ont and press Enter
7. Press the ] key, the prompt should change to say (ONTRename) pkg>
8. Run instantiate
9. Press the backspace key to return to the julia> prompt
10. Run exit()
11. Close PowerShell
12. Right-click the Windows desktop, and click "New | Shortcut..."
13. Type julia --project=%USERPROFILE%\ont %USERPROFILE%\ont\ont-rename.jl and click Next
14. Type Rename ONT FASTQs or something else memorable and click "Finish"