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Pre-requisites

Overview:

  • Create working directory folders for hands-on training exercises

  • Copy data and scripts for today’s session

  • (Optional - those that did not attend Session 3) run a test to verify Nextflow is properly installed and to test the execution of the nf-core/rnaseq pipeline with example data provided by developers.

Prepare Working Directory Space for Session 4

Before we start using the HPC, let’s start an interactive session:

qsub -I -S /bin/bash -l walltime=10:00:00 -l select=1:ncpus=2:mem=8gb

  • where:

    • ‘walltime’ is amount of time requested to run the interactive session

    • ‘cpus’ number of CPUs to be used by the interactive session

    • ‘mem’ amount of memory assigned to the interactive session

Create working directories

Let’s create the working directories for today’s session:

mkdir $HOME/workshop/2024-2/session4_RNAseq
mkdir $HOME/workshop/session4_RNAseq/scripts
mkdir $HOME/workshop/session4_RNAseq/data
mkdir $HOME/workshop/session4_RNAseq/runs
mkdir $HOME/workshop/session4_RNAseq/runs/run1_QC
mkdir $HOME/workshop/session4_RNAseq/runs/run2_RNAseq
mkdir $HOME/workshop/session4_RNAseq/runs/run3_RNAseq_T2T

The folder structure will look like this: 

session4_RNAseq/
├── data
├── runs
│   ├── run1_QC
│   ├── run2_RNAseq
│   └── run3_RNAseq_T2T
└── scripts

Get a copy of the scripts

Now let’s copy the scripts necessary for today’s session:

cp /work/training/2024/rnaseq/scripts/* $HOME/workshop/2024-2/session4_RNAseq/scripts

  • Line 1: Copies all files from /work/datasets/workshop/scripts/ as noted by an asterisk to the newly created folder $HOME/workshop/2024-2/session4_RNAseq/scripts/

Copy public data to your $HOME

cp /work/training/2024/rnaseq/data/* $HOME/workshop/2024-2/session4_RNAseq/data/

  • Line 1: Copies all files from /work/datasets/workshop/scripts/ folder as noted by an asterisk to newly created folder $HOME/workshop/2024-2/session4_RNAseq/data/

Create a folder for running the nf-RNA-seq pipeline

Let’s create an “runs” folder in the ~/workshop/2024/rnaseq folder to run the nf-core/rnaseq pipeline. For example:

mkdir -p $HOME/workshop/2024/rnaseq/runs
mkdir $HOME/workshop/2024/rnaseq/runs/run1_test
mkdir $HOME/workshop/2024/rnaseq/runs/run2_QC
mkdir $HOME/workshop/2024/rnaseq/runs/run3_RNAseq
cd $HOME/workshop/2024/rnaseq/runs

  • Lines 1-4: create sub-folders for each exercise

  • Line 5: change the directory to the folder “run1_test”

  • Line 6: print the current working directory

(Optional ): Running a test with nf-core sample data

First, let’s assess the execution of the nf-core/rnaseq pipeline by running a test using sample data.

Copy the launch_nf-core_RNAseq_test.pbs to the working directory

cd $HOME/workshop/2024-2/session4_RNAseq/runs/run1_test
cp $HOME/workshop/2024-2/session4_RNAseq/scripts/launch_nf-core_RNAseq_test.pbs .

View the content of the script as follows:

cat launch_nf-core_RNAseq_test.pbs

#!/bin/bash -l

#PBS -N nfrnaseq_test

#PBS -l select=1:ncpus=2:mem=4gb

#PBS -l walltime=24:00:00

 

#work on current directory

cd $PBS_O_WORKDIR

 

#load java and set up memory settings to run nextflow

module load java

export NXF_OPTS='-Xms1g -Xmx4g'

 

nextflow run nf-core/rnaseq -r 3.14.0 -profile test,singularity --outdir results

  • nextflow command: nextflow run

  • pipeline name: nf-core/rnaseq

  • pipeline version: -r 3.14.0

  • container type and sample data: -profile test,singularity

  • output directory: --outdir results

Submitting the job

Submit the test job to the HPC cluster as follows:

qsub launch_nf-core_RNAseq_test.pbs

Monitoring the Run

qjobs

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