Error: gff_merge with wf-transcriptomics workflow.

[Aravind-mss]

Operating System

Other Linux (please specify below)

Other Linux

Rocky Linux

Workflow Version

v1.5.0

Workflow Execution

Command line (Cluster)

Other workflow execution

No response

EPI2ME Version

No response

CLI command run

nextflow run epi2me-labs/wf-transcriptomes \ --fastq $input \ --de_analysis \ --ref_annotation "$genomedir/GRCh38.p14_genomic.gtf" \ --ref_genome "$genomedir/GRCh38.p14_genomic.fa" \ --sample_sheet "$mydir/samplesheet.csv" \ --out_dir "$mydir/transcriptomics_fastq" -profile singularity

Workflow Execution - CLI Execution Profile

None

What happened?

I have tried with both fastq , uBMA and BAM inputs but unable make any progress beyond this step.
I have used the below way of running the pipeline on my last try. But no luck. The same step failed like before. Please help.

Relevant log output

executor > pbspro (110)
[46/625c71] validate_sample_sheet | 1 of 1 ✔
[67/de8deb] fastcat (1) | 6 of 6 ✔
[bc/af7649] pip…:preprocess_ref_annotation | 1 of 1 ✔
[e7/1a7ccf] pip…stqIngressResultsInDir (6) | 6 of 6 ✔
[c8/6eda3f] pipeline:getVersions | 1 of 1 ✔
[c0/8add90] pipeline:getParams | 1 of 1 ✔
[eb/ffbc6a] pipeline:preprocess_reads (5) | 6 of 6 ✔
[b9/260bbb] pip…ne:build_minimap_index (1) | 1 of 1 ✔
[2c/ebb78b] pip…nce_assembly:map_reads (6) | 6 of 6 ✔
[15/bf5486] pipeline:split_bam (6) | 6 of 6 ✔
[52/f6a5c4] pip…:assemble_transcripts (58) | 67 of 67 ✔
[cb/b8006b] pipeline:merge_gff_bundles (3) | 1 of 6, failed: 1
[- ] pipeline:run_gffcompare -
[- ] pipeline:get_transcriptome -
[1a/56fee6] pip…ne:check_annotation_strand | 1 of 1 ✔
[- ] pipeline:merge_transcriptomes -
[12/4005f3] pip…ckSampleSheetCondition (1) | 1 of 1 ✔
[- ] pip…inimap_index_transcriptome -
[- ] pip…pression:map_transcriptome -
[- ] pip…pression:count_transcripts -
Plus 6 more processes waiting for tasks…
ERROR ~ Error executing process > 'pipeline:merge_gff_bundles (3)'

Caused by:
Process pipeline:merge_gff_bundles (3) terminated with an error exit status (1)

Command executed:

echo '##gff-version 2' >> BEL14169A3.gff;
echo '#pipeline-nanopore-isoforms: stringtie' >> BEL14169A3.gff;

for fn in BEL14169A3_batch_7.gff BEL14169A3_batch_8.gff BEL14169A3_batch_6.gff BEL14169A3_batch_3.gff BEL14169A3_batch_9.gff BEL14169A3_batch_4.gff BEL14169A3_batch_5.gff BEL14169A3_batch_2.gff BEL14169A3_batch_1.gff;
do
grep -v '#' $fn >> BEL14169A3.gff

done

workflow-glue summarise_gff BEL14169A3.gff BEL14169A3 transcriptome_summary.pickle

Command exit status:
1
Command output:
(empty)

Application activity log entry

No response

Were you able to successfully run the latest version of the workflow with the demo data?

yes

Other demo data information

No response

[nrhorner]

Hi @Aravind-mss

Could you share the whole .nextflow.log please? Also would you be able to post the output of -wc -l <work_dir>/*. The workdir path should be near the the bottom of the full error message.

[Aravind-mss]

Hi @nrhorner
Here is the output from the <work_dir>
wc -l /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/* 6514 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_1.gff 8153 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_2.gff 8202 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_3.gff 7143 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_4.gff 6742 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_5.gff 6699 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_6.gff 7490 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_7.gff 4199 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_8.gff 2 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3_batch_9.gff 26584 /share/bioinfo/wf_transcriptomics/BAMs/work/cb/b8006be2802d4204d78ace6acd56fc/BEL14169A3.gff 81728 total
Attached the nexflow log for review. Thanks.
nextflow.log

[nrhorner]

Thanks for that @Aravind-mss

The log you sent does not match the original error. It has this instead, which indicates a different issue.

Caused by:
  Process `pipeline:differential_expression:deAnalysis (1)` terminated with an error exit status (1)

Can you send the log associated with the original error please?

[Aravind-mss]

Apologies @nrhorner , it appears that the log was overwritten.

Here is the output requested and the log associated with the error after a rerun.

wc -l /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/* 6514 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_1.gff 8153 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_2.gff 8202 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_3.gff 7143 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_4.gff 6742 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_5.gff 6699 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_6.gff 7490 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_7.gff 4199 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_8.gff 2 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_9.gff 37494 /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3.gff 92638 total
transcriptomics.nextflow.log

[nrhorner]

Hi @Aravind-mss

The following gff file looks suspiciously small, could you post the contents of it please?
/share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3_batch_8.gff

[nrhorner]

Actually, this file would also be useful, if you're able to share: /share/bioinfo/z35/ONT_BEL14169/wf_transcriptomics/BAMs/work/e6/a6674b98090329e41bcbcc51503a3c/BEL14169A3.gff

[Aravind-mss]

BEL14169A3_batch_8.zip
Hi @nrhorner Both the files required are in this zip file.

[nrhorner]

Thanks for those @Aravind-mss

Those files seem ok to me. But they don't seem to be the files from the work directory that caused the issue.

I have the number of lines in each at

37494 BEL14169A3.gff
4199 BEL14169A3_batch_8.gff

while you report 92638, and 2

Could you rerun the workflow again, and if it fails, zip up the whole wokdir and post it?

[Aravind-mss]

Hi @nrhorner
I have run the pipeline again and it failed at the same step. I have made the work directory and the nextflow log via the link below.
https://mydata.ramaciotti.unsw.edu.au/s/ACP2j5gg5Ko9mjr/download/transcriptomics_work.tar

[nrhorner]

thanks @Aravind-mss I'll take a look ASAP

[nrhorner]

HI @Aravind-mss

I've recreated your issue. It's caused by an empty GFF file at this step. Sorry for the inconvenience. I will get this fixed ASAP.

[nrhorner]

Hi @Aravind-mss

v1.6.1 has just been released, which will hopefully fix your issue.