nf-core/scrnaseq: A single-cell RNAseq pipeline for 10X genomics data

Prepared by the eResearch Office, QUT.

This page provides a guide to QUT users on how to install and run the NextFlow https://nf-co.re/scrnaseq/1.1.0workflow on the HPC.

Further details on setting up, running and output of the nf-core/scrnaseq workflow can be found here:

https://nf-co.re/scrnaseq/1.1.0

https://nf-co.re/scrnaseq/1.1.0/usage

https://nf-co.re/scrnaseq/1.1.0/parameters

https://nf-co.re/scrnaseq/1.1.0/output

Table of contents

Analysis environment

Nextflow is designed to be run on Linux command-line. This guide assumes the QUT HPC (high performance computing cluster) will be used for this. When running a complex analysis (e.g. a Nextflow workflow) on the HPC, commands are submitted into a job queue using a system called PBS.

If you don’t have a HPC account or haven’t used the HPC/PBS before, click on the below link for details on how to create a HP account and how to use the PBS job submission system:

Start using the HPC

This nf-core/scrnaseq guide is designed so that people with even minimal Linux command-line experience can use it. However, some familiarity with using Linux at a command line is recommended. If you have no experience at all using Linux commands, we advise you watch an introductory video or video series on the topic, such as:

Linux Command Line Tutorial For Beginners 1 - Introduction

Install Nextflow

The nf-core/scrnaseq workflow requires Nextflow to be installed in your account on the HPC. Find details on how to install and test Nextflow here, including preparing a nextflow.config file and running a PBS pro submission script for Nextflow pipelines.

Additional information available here: https://nf-co.re/usage/installation

Test the nf-core/scrnaseq workflow

The nf-core/scrnaseq workflow contains a built-in dataset to test. The below code chunk is the basic command to run the nf-core/scrnaseq test.

However, as mentioned, commands should be submitted to the queue, rather than copy and pasted directly. See the below section for how to submit the above as a PBS job.

Submitting as a PBS job

To create a PBS job script to run the above nf-core/scrnaseq test, follow these steps:

1. Load a text editor, such as nano (copy and paste the following code into the command-line)

2. Create an empty file called ‘launch.pbs’ using nano

3. Copy and paste the following into nano

4. Exit and save the file. Control-x exits nano which then asks you if you want to save.

5. The job can then be submitted by:

Viewing the workflow progress

When submitted as a PBS job, Nextflow splits the analysis up into multiple sub-processes that are run and queued according to available HPC memory and CPUs. Nextflow Tower has been set up on the HPC so you can observe sub-processes as they are submitted, run and completed, as well as a multitude of other details.

To access Nextflow Tower, click on the below link.

https://nftower.qut.edu.au/

This will ask you for an email address. Submit your QUT email address and you will be sent a link to access Tower.

You can now watch the progress of your nf-core/scrnaseq test workflow (or any other Nextflow workflows you are running). When all the sub-processes are complete and the nf-core/scrnaseq turns green, the test is successful.

Full workflow

nextflow run nf-core/scrnaseq -profile test,singularity --aligner kallisto -resume --fasta https://github.com/nf-core/test-datasets/raw/scrnaseq/reference/GRCm38.p6.genome.chr19.fa --gtf https://github.com/nf-core/test-datasets/raw/scrnaseq/reference/gencode.vM19.annotation.chr19.gtf

Mouse genome

http://ftp.ensembl.org/pub/release-104/fasta/mus_musculus/dna/Mus_musculus.GRCm39.dna.primary_assembly.fa.gz

Mouse GTF

http://ftp.ensembl.org/pub/release-104/gtf/mus_musculus/Mus_musculus.GRCm39.104.chr.gtf.gz