Hi all, I'm trying to run this pipeline for the first time (1.04), and I'm hitting a wall when it gets to

STAR_SALMON:DRIMSEQ_FILTER

:

ERROR ~ Error executing process > 'NFCORE_RNASPLICE:RNASPLICE:DRIMSEQ_DEXSEQ_DTU_STAR_SALMON:DRIMSEQ_FILTER (1)'

Caused by:
  Process `NFCORE_RNASPLICE:RNASPLICE:DRIMSEQ_DEXSEQ_DTU_STAR_SALMON:DRIMSEQ_FILTER (1)` terminated with an error exit status (1)


Command executed:

  run_drimseq_filter.R salmon.merged.txi.dtu.rds tximport.tx2gene.tsv samplesheet.csv \
      4 \
      2 \
      2 \
      10 \
      0.1 \
      10
  
  cat <<-END_VERSIONS > versions.yml
  "NFCORE_RNASPLICE:RNASPLICE:DRIMSEQ_DEXSEQ_DTU_STAR_SALMON:DRIMSEQ_FILTER":
      r-base: $(echo $(R --version 2>&1) | sed 's/^.*R version //; s/ .*$//')
      bioconductor-drimseq: $(Rscript -e "library(DRIMSeq); cat(as.character(packageVersion('DRIMSeq')))")
  END_VERSIONS

Command exit status:
  1

Command output:
  (empty)

Command error:
  INFO:    Environment variable SINGULARITYENV_TMPDIR is set, but APPTAINERENV_TMPDIR is preferred
  INFO:    Environment variable SINGULARITYENV_NXF_TASK_WORKDIR is set, but APPTAINERENV_NXF_TASK_WORKDIR is preferred
  INFO:    Environment variable SINGULARITYENV_NXF_DEBUG is set, but APPTAINERENV_NXF_DEBUG is preferred
  INFO:    Environment variable SINGULARITYENV_SNIC_TMP is set, but APPTAINERENV_SNIC_TMP is preferred
  WARNING: Skipping mount /var/apptainer/mnt/session/etc/resolv.conf [files]: /etc/resolv.conf doesn't exist in container
  
  Attaching package: 'DRIMSeq'
  
  The following object is masked from 'package:base':
  
      proportions
  
  Error in .local(x, ...) : 
    min_samps_gene_expr >= 0 && min_samps_gene_expr <= ncol(x@counts) is not TRUE
  Calls: <Anonymous> -> <Anonymous> -> .local -> stopifnot
  Execution halted

I have only one sample per condition, so if this an issue with a lack of replicates? My samplesheet:

sample,fastq_1,fastq_2,strandedness,condition
ox10,ox10_1.fq.gz,ox10_2.fq.gz,unstranded,control
ox20,ox20_1.fq.gz,ox20_2.fq.gz,unstranded,control
ox40,ox40_1.fq.gz,ox40_2.fq.gz,unstranded,treatment

My contrasts:

contrast,treatment,control
ox40_ox10,ox40,ox10
ox40_ox20,ox40,ox20
ox20_ox10,ox20,ox10

Any help appreciated!

You need replicates for any kind of statistics. Don't know the details of the failing process, but that may well be the reason. If you really want to know, you can run the same sample under different names. (Obviously don't use those results.) That might give other errors, though, since the intra-group variance would be zero.

This was my hunch too, but the odd thing is that it tries to kick off

DRIMseq

process before the STAR alignment has been done, therefore there are no counts to work on. I had to resume the pipeline many times before most steps were done. It still crashes at this point, but now at least I have something to work with in the output which is totally fine.