TPMCalculator quantifies mRNA abundance directly from the alignments by parsing BAM files. The input parameters are the same GTF files used to generate the alignments, and one or multiple input BAM file(s) containing either single-end or paired-end sequencing reads. The TPMCalculator output is comprised of four files per sample reporting the TPM values and raw read counts for genes, transcripts, exons and introns respectively.

References:

• Roberto Vera Alvarez, Lorinc Sandor Pongor, Leonardo Mariño-Ramírez, David Landsman; TPMCalculator: one-step software to quantify mRNA abundance of genomic features, Bioinformatics, , bty896, https://doi.org/10.1093/bioinformatics/bty896

• TPMCalculator on github
• TPMCalculator Wiki

• Module Name: TPMCalculator (see the modules page for more information)
• Singlethreaded.
• Example files in /usr/local/apps/TPMCalculator/TEST_DATA
• Reference data in fdb/igenomes/

Allocate an interactive session and run the program.
Sample session (user input in bold):

[user@biowulf]$ sinteractive
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 TPMCalculator

[user@cn3144 ~]$ cp -r /usr/local/apps/TPMCalculator/TEST_DATA  /data/$USER

[user@cn3144 ~]$ cd /data/$USER/TEST_DATA

[user@cn3144 ~]$ TPMCalculator -v -g /fdb/igenomes/Homo_sapiens/UCSC/hg38/Annotation/Genes/genes.gtf -b SRR2126790_sorted.bam -q 255
Reading GTF file ...
Done in 57.6513 seconds
Parsing sample: SRR2126790_sorted
 3087949 reads processed in 80.0002 seconds
Printing results
Total time: 139.753 seconds

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

Create a batch input file (e.g. TPMCalculator.sh). For example, the following script will copy the sample data, run the program, and compare the output to the validated results.

#!/bin/bash
set -e
module load TPMCalculator

cp -r /usr/local/apps/TPMCalculator/TEST_DATA  /data/$USER

cd /data/$USER/TEST_DATA

./run.sh

Submit this job using the Slurm sbatch command.

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

Create a swarmfile (e.g. TPMCalculator.swarm). For example:

TPMCalculator -v -g /fdb/igenomes/Homo_sapiens/UCSC/hg38/Annotation/Genes/genes.gtf   -b file1.bam   -q 255
TPMCalculator -v -g /fdb/igenomes/Homo_sapiens/UCSC/hg38/Annotation/Genes/genes.gtf   -b file2.bam   -q 255
TPMCalculator -v -g /fdb/igenomes/Homo_sapiens/UCSC/hg38/Annotation/Genes/genes.gtf   -b file3.bam   -q 255
[...]

Submit this job using the swarm command.

swarm -f TPMCalculator.swarm [-g #] --module TPMCalculator