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Pre-requisites

• Nextflow installed → If not see: 1. Getting started with Nextflow Ask for help if you need assistance.

Overview:

• Create working directory folders for hands-on training exercises

• Copy data and scripts for today’s session

• (Optional ) run a test to verify Nextflow is properly installed and to assess the execution of the nf-core/rnaseq pipeline with example data provided by developers.

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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=4gb

• 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

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mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/scripts
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/scripts/local
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/data
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/data/human_disease
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/data/human_drosha
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/data/mydata
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/runs
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/runs/run0_test
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/runs/run1_human_miRBase
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/runs/run2_human_MirGeneDB
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/runs/run3_drosha_miRBase
mkdir -p $HOME/workshop/2024-2/session6_smallRNAseq/runs/run4_drosha_MirGeneDB

The above code creates the following folder structure:

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cp /work/training/2024/smallRNAseq/scripts/* $HOME/workshop/2024-2/session6_smallRNAseq/scripts
ls -l $HOME/workshop/2024-2/session6_smallRNAseq/scripts
cp /work/training/2024/smallRNAseq/scripts/local/* $HOME/workshop/2024-2/session6_smallRNAseq/scripts/local
ls -l $HOME/workshop/2024-2/session6_smallRNAseq/scripts/local

• 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/

• Line 2: List the files copied to the ‘script’ folder

• Line 3: Copies all files from /work/datasets/workshop/scripts/local as noted by an asterisk to the newly created folder $HOME/workshop/2024-2/session4_RNAseq/scripts/local

• Line 4: List the files copied to the ‘script’ folder

Copy public data to working folder

cp /work/training/2024/smallRNAseq/data/human_disease/* $HOME/workshop/2024-2/session6_smallRNAseq/data/human_disease
ls -l $HOME/workshop/2024-2/session6_smallRNAseq/data/human_disease
cp /work/training/2024/rnaseqsmallRNAseq/data/human_drosha/* $HOME/workshop/2024-2/session6_smallRNAseq/data/human_drosha
ls -l $HOME/workshop/2024-2/session6_smallRNAseq/data/human_drosha

• Line 1: Copies mouse RNAseq folder to created ~/data/mouse folder

• Line 2: List the files copied to the ‘data’ folder

• Line 3: Copies human RNAseq folder to created ~/data/human folder

• Line 4: List the files copied to the ‘data’ folder

(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.

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mkdir -p $HOME/workshop/2024-2/session4session6_RNAseqsmallRNAseq/runs/run0_test
cd $HOME/workshop/2024-2/session4session6_RNAseqsmallRNAseq/runs/run0_test

• Line 1: Create folder using the make directory “mkdir” command. We use the flap -p (parental) to create parental folders if not yet created.

• Line 2: Move to the newly created folder.

Copy the launch_nf-core_RNAseq_test.pbs from the ‘scripts’ folder to the current folder

cp $HOME/workshop/2024-2/session4session6_RNAseqsmallRNAseq/scripts/launch_nf-core_RNAseqsmallRNAseq_test.pbs .

View the content of the script as follows:

cat launch_nf-core_RNAseqsmallRNAseq_test.pbs

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• nextflow command: nextflow run

• pipeline name: nf-core/rnaseqsmrnaseq

• pipeline version (revision): -r 2.3.14.01

• 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_RNAseqsmallRNAseq_test.pbs

Monitoring the Run

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