SLAMseq is a novel sequencing protocol that directly uncovers 4-thiouridine incorporation events in RNA by high-throughput sequencing. When combined with metabolic labeling protocols, SLAM-seq allows to study the intracellular RNA dynamics, from transcription, RNA processing to RNA stability. SlamDunk is a novel, fully automated software tool for automated, robust, scalable and reproducible SLAMseq data analysis.

References:

• Thiol-linked alkylation of RNA to assess expression dynamics, Herzog VA et al., Nature Methods, 2017

• slamdunk documentation

Slamdunk is available as a Docker container. While Docker is not available on Biowulf, the Singularity container system can run Docker images. You can therefore load the singularity module, pull the docker container, and run Slamdunk directly. It is important to set the SINGULARITY_BINDPATH environment variable to include any directories that you need mapped within the container.

Allocate an interactive session and run the program.
Sample session (user input in bold). In this example, the input files are in the user's /data area, and the reference genome file is in /fdb, so SINGULARITY_BINDPATH is set to "/data/$USER,/fdb"

[user@biowulf]$ sinteractive --cpus-per-task=8 --mem=20g
salloc.exe: Pending job allocation 46116226
salloc.exe: job 46116226 queued and waiting for resources
salloc.exe: job 46116226 has been allocated resources
salloc.exe: Granted job allocation 46116226
salloc.exe: Waiting for resource configuration
salloc.exe: Nodes cn3144 are ready for job

[user@cn3144 ~]$ module load singularity

[user@cn3144 ~]$ export SINGULARITY_BINDPATH="/data/$USER,/fdb"

[user@cn3144 ~]$ singularity run docker://tobneu/slamdunk

[user@cn3144 ~]$ slamdunk all -r /fdb/genome/hg19/chr_all.fa -b ./exampleBAM.bed \
      -o out -t $SLURM_CPUS_PER_TASK ./exampleBAM.bam
slamdunk all
Running slamDunk map for 1 files (8 threads)
.
Running slamDunk sam2bam for 1 files (8 threads)
.
Creating output directory: out/filter
Running slamDunk filter for 1 files (8 threads)
.
[...]

[user@cn3144 ~]$ exit
salloc.exe: Relinquishing job allocation 46116226
[user@biowulf ~]$

Create a batch input file (e.g. slamdunk.sh). For example:

#!/bin/bash
set -e
module load singularity
export SINGULARITY_BINDPATH="/data/$USER,/fdb,/lscratch"
singularity exec docker://tobneu/slamdunk \
      slamdunk all -r /fdb/genome/hg19/chr_all.fa -b ./exampleBAM.bed \
      -o out -t $SLURM_CPUS_PER_TASK ./exampleBAM.bam

Submit this job using the Slurm sbatch command.

sbatch [--cpus-per-task=#] [--mem=#] slamdunk.sh