A package for including transposable elements in differential enrichment analysis of sequencing datasets.

References:

• Ying Jin, O. H. Tam, E. Paniagua and M. Hammell. TEtranscripts: a package for including transposable elements in differential expression analysis of RNA-seq datasets. Bioinformatics 2015, 23:btv422. Pubmed |  PMC |  Journal

• GitHub
• Home page

• Module Name: tetoolkit (see the modules page for more information)
• Example files in $TETOOLKIT_TEST_DATA
• Reference data in /fdb/tetoolkit/

Allocate an interactive session and run the program. Sample session:

[user@biowulf]$ sinteractive --mem=10g --cpus-per-task=12 --gres=lscratch:50
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 tetoolkit
[user@cn3144]$ cp -rL $TETOOLKIT_TEST_DATA/01fastq .
[user@cn3144]$ cp $TETOOLKIT_TEST_DATA/GTF/dm6_rmsk_TE.gtf.gz .
[user@cn3144]$ gunzip dm6_rmsk_TE.gtf.gz

This test data contains 4 untreated samples (GSM461176, GSM461177, GSM461178, GSM461182) and 3 samples with RNAi depleted Pasilla gene expression (GSM461179, GSM461180, GSM461181). First, let's align it in a way that is compatible with TEtranscripts. This will take about 1h.

[user@cn3144]$ module load STAR/2.7.3a
[user@cn3144]$ genome=/fdb/STAR_indices/2.7.3a/UCSC/dm6/genes-75
[user@cn3144]$ mkdir -p 02aln
[user@cn3144]$ for fq in 01fastq/*.fastq.gz ; do
     STAR --runThreadN $SLURM_CPUS_PER_TASK \
          --genomeDir $genome \
          --readFilesCommand zcat \
          --outSAMtype BAM Unsorted \
          --winAnchorMultimapNmax 200 \
          --outFilterMultimapNmax 100 \
          --readFilesIn "$fq" \
          --outFileNamePrefix "./02aln/$(basename $fq .fastq.gz)"
 done

Now we can run TEtoolkit on the alignments.

[user@cn3144]$ untreated=(
     02aln/GSM461176Aligned.out.bam
     02aln/GSM461177Aligned.out.bam
     02aln/GSM461178Aligned.out.bam
     02aln/GSM461182Aligned.out.bam )
[user@cn3144]$ treated=(
     02aln/GSM461179Aligned.out.bam
     02aln/GSM461180Aligned.out.bam
     02aln/GSM461181Aligned.out.bam )
[user@cn3144]$ TEtranscripts --mode multi \
    --TE dm6_rmsk_TE.gtf \
    --GTF /fdb/STAR_indices/2.7.3a/UCSC/dm6/genes.gtf \
    --minread 100 \
    --project test \
    -c "${untreated[@]}" \
    -t "${treated[@]}"
...
[user@cn3144]$ ls -1
drwxr-xr-x 2 user group 4.0K Apr 10 08:50 01fastq
drwxr-xr-x 2 user group 4.0K Apr 10 09:38 02aln
-rw-r--r-- 1 user group 5.3M Apr 10 08:51 dm6_rmsk_TE.gtf
-rw-r--r-- 1 user group 521K Apr 10 14:13 test.cntTable
-rw-r--r-- 1 user group  735 Apr 10 14:13 test_DESeq2.R
-rw-r--r-- 1 user group 878K Apr 10 14:13 test_gene_TE_analysis.txt
-rw-r--r-- 1 user group  94K Apr 10 14:13 test_sigdiff_gene_TE.txt

[user@cn3144]$ head -5 test_sigdiff_gene_TE.txt
baseMean        log2FoldChange  lfcSE   stat    pvalue  padj
AGBE    1668.50772310032        0.489686064951153       0.162938655205519       3.00534004244418        0.00265284088823233   0.0288848429674012
AGO2    8993.79284144723        -0.265578080875274      0.0892885493032728      -2.97438006270239       0.0029358119956592    0.0313958714926737
ATP7    1750.30129130752        0.526853871760357       0.161346748429612       3.26535165343106        0.00109328255464026   0.0139041285415587
Aats-cys        1256.3667600502 -0.670235513690024      0.105205551162389       -6.37072384759898       1.8813812974601e-10   1.13699601511619e-08

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

Create a batch input file (e.g. tetoolkit.sh), which uses the input file 'tetoolkit.in'. For example:

#! /bin/bash
# this is tetranscripts.sh

module load tetoolkit/2.1.4 || exit 1
untreated=(
     02aln/GSM461176Aligned.out.bam
     02aln/GSM461177Aligned.out.bam
     02aln/GSM461178Aligned.out.bam
     02aln/GSM461182Aligned.out.bam )
treated=(
     02aln/GSM461179Aligned.out.bam
     02aln/GSM461180Aligned.out.bam
     02aln/GSM461181Aligned.out.bam )
TEtranscripts --mode multi \
    --TE dm6_rmsk_TE.gtf \
    --GTF /fdb/STAR_indices/2.7.3a/UCSC/dm6/genes.gtf \
    --minread 100 \
    --project test \
    -c "${untreated[@]}" \
    -t "${treated[@]}"

Submit this job using the Slurm sbatch command.

sbatch --cpus-per-task=2 --mem=10g tetoolkit.sh