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This page provides a basic introduction to Unix commands to HPC users with no previous knowledge.

Log into the HPC

ssh userID@lyra.qut.edu.au

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To go to a shared directory for your project named “kenna_team” type the following command and hit enter:

cd /work/kennachlamydia_teamcarey/

Display list of files in a directory

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Make a backup copy of the file

cp myfile.txt > myfile_copy.txt 

Move a copy of a file to a newly created folder - note it is recommended to make a copy of important files prior to modifying or executing commands on them.

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#hash tags are used to add comments on what a command line does
#several commands can be used including cat, less, more, head and tail
cat myfile_copy.txt

#example: less -S allows to visualise very large (wide) files
less -S myfile_copy.txt

#stop viewing a file using the above command
--> Type “Control” and “c” at the same time.
      Or “Control” and “d” at the same time.

#print the first 505 lines of a file
head -505 myfile_copy.txt

#print the last 205 lines of a file
tail -205 myfile_coy.txt

Go back to my personal space. Type 'cd' and hit enter. This will move you to /home/mystudentID/

cd

Interactive session:

qsub -I -S /bin/bash -l walltime=10:00:00 -l select=1:ncpus=2:mem=4gb

Running the nextflow nf-core/rnaseq pipeline

Requirements:

• index.csv → a file that provides a list of sample IDs and their associated FASTQ files (read 1 and read 2)

• launch.pbs → a script to submit the job to the HPC cluster

Example index.csv file for nf-core/rnaseq version 3.3:

group,fastq_1,fastq_2,strandedness
control_r1,/work/kenna_team/data/raw_data/SRR1039508_1.fastq.gz,/work/kenna_team/data/raw_data/SRR1039508_2.fastq.gz,unstranded
dex_r1,/work/kenna_team/data/raw_data/SRR1039509_1.fastq.gz,/work/kenna_team/data/raw_data/SRR1039509_2.fastq.gz,unstranded
control_r2,/work/kenna_team/data/raw_data/SRR1039512_1.fastq.gz,/work/kenna_team/data/raw_data/SRR1039512_2.fastq.gz,unstranded
dex_r2,/work/kenna_team/data/raw_data/SRR1039513_1.fastq.gz,/work/kenna_team/data/raw_data/SRR1039513_2.fastq.gz,unstranded
control_r3,/work/kenna_team/data/raw_data/SRR1039516_1.fastq.gz,/work/kenna_team/data/raw_data/SRR1039516_2.fastq.gz,unstranded
dex_r3,/work/kenna_team/data/raw_data/SRR1039517_1.fastq.gz,/work/kenna_team/data/raw_data/SRR1039517_2.fastq.gz,unstranded
control_r4,/work/kenna_team/data/raw_data/SRR1039520_1.fastq.gz,/work/kenna_team/data/raw_data/SRR1039520_2.fastq.gz,unstranded
dex_r4,/work/kenna_team/data/raw_data/SRR1039521_1.fastq.gz,/work/kenna_team/data/raw_data/SRR1039521_2.fastq.gz,unstranded

Example launch.pbs script:

#!/bin/bash -l
#PBS -N nfrnaseq
#PBS -l select=1:ncpus=2:mem=4gb
#PBS -l walltime=24:00:00

#Use the current directory to run the workflow
cd $PBS_O_WORKDIR

module load java
NXF_OPTS='-Xms1g -Xmx4g'

#run the nextflow RNA-seq pipeline:
nextflow run nf-core/rnaseq -profile singularity -r 3.3 --aligner star_salmon --input index.csv --genome GRCh38 -resume

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#!/bin/bash -l
#PBS -N nfrnaseq
#PBS -l select=1:ncpus=2:mem=4gb
#PBS -l walltime=24:00:00

#Use the current directory to run the workflow
cd $PBS_O_WORKDIR

module load java
NXF_OPTS='-Xms1g -Xmx4g'

#run the nextflow RNA-seq pipeline:
nextflow run nf-core/rnaseq -profile singularity -r 3.3 --aligner star_salmon --input index.csv --genome GRCh38 --clip_r1 10 --clip_r2 10 --three_prime_clip_r1 2 --three_prime_clip_r2 2 --save_trimmed

#allow access to others in the group
chmod -R g+rwX results
chmod -R g+rwX work

Session 2 exercises:

1. Run the nf-core/rnaseq pipeline using the Airway smooth muscle public data (PMID: 24926665. GEO: GSE52778) - aligner option set to ‘star_salmon’

2. Same as above but aligner option set to ‘star_rsem’

Create a new working folder:

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Access the HPC files from your laptop

Mac laptop (note: need to be connected via VPN)

• Open the ‘Finder' window

• Click on the search file tab and hit the “Command + K” keys simultaneously

• This will open a new window:

  

• Type the above to connect to the shared ‘work' space. To access your personal space replace ‘work’ with 'home’.

Next

Differential expression analysis using https://maayanlab.cloud/biojupies/analyze