bulkRNAseq_Oct2025_workshop

Author: Qiaoshan Lin, Ph.D.

These materials are provided as part of the workshop Practical Workshop on Bulk RNA-seq Bioinformatics Pipelines held on October 10th, 2025, at the University of Chicago. The workshop covers the basic usage of the nextflow RNAseq pipeline and the in-house application for downstream DE analysis.

Section 1: Run Nextflow RNAseq Pipeline on Randi

Step 1: Log into Randi

ssh <replace_with_your_account_name>@randi.cri.uchicago.edu

Step 2: Change to workshop directory

cd /gpfs/data/biocore-workshop/bulkRNAseq_Oct2025_workshop

List contents:

ls -l

Step 3: Build your working directory

cd testRun

Check current working directory (where you are at):

pwd

Check your user name on server:

echo $USER

Make your own working directory

mkdir -p $USER

Go into your working directory

cd $USER

Check current working directory again (where you are at):

pwd

Step 4: Prepare scripts and configurations

Copy all necessary files from template to your own working directory:

cp ../template/* .

Check the contents:

ls -l

Check the file text:

cat nextflow.config

cat nextflow.slurm

Step 5: Submit the job

Submit a batch job

sbatch nextflow.slurm

Check the job status:

squeue -u $USER

or

squeue -j <JOB_ID>

Check the error and out logs:

cat nextflow_*.err
cat nextflow_*.out

Section 2: In-House Downstream Analysis

Step 1: Setup experimental groups

Check the contents in the metadata table

cat metadata.txt

Step 2: Run the slurm script

Check the slurm script for running the app:

cat app.slurm

Check out log to confirm the pipeline is completed successfully:

cat nextflow_*.out

Remove intermediate files once nextflow completion is confirmed:

rm -r -f work/ tmp/ singularity/ .nextflow*

Submit the batch job for running shiny app:

sbatch app.slurm

Check the job status:

squeue -u $USER

or

squeue -j <JOB_ID>

Step 3: Fill out the form and submit

Open a new terminal on your local computer.

Run the following from your local terminal:

ssh -N -f -L 3838:<replace_with_your_shinyApp_running_node>:3838 <replace_with_username>@randi.cri.uchicago.edu

or

ssh -N -J <replace_with_username>@randi.cri.uchicago.edu -L 3838:0.0.0.0:3838 <replace_with_username>@shinyApp_running_node

Open http://localhost:3838/ in your local browser (e.g. Chrome)

Note that: There is a chance that when many users are running on the same node, the port 3838 gets congested. When a port conflict happens, change the port number and re-submit app.slurm.

Note: If your nextflow run is not completed yet, please use the result folder at /gpfs/data/biocore-workshop/bulkRNAseq_Oct2025_workshop/testRun/template/results as the Nextflow Output Directory in the form.

Step 4: Cancel your jobs

Kill all your jobs:

scancel -u $USER

Kill one job:

scancel <replace_with_your_job_ID>