Hi all, I had a successful run with the test_full profile, but when I tried the pipeline with a minimal dataset, these 7 tasks failed. Despite multiple attempts and resuming, these tasks still fail. I've attached my dummy sample sheet and contrast sheet. Could you please advise on the root cause of the failure for these tasks? Compute environment Platform: azure-batch Nextflow version: 23.10.1 build 5891 Pipeline parameters:

{
  "input": "#####/RNASplice-Samplesheet.csv",
  "contrasts": "https://#####/RNASplice-ContrastSamplesheet-try1.csv",
  "source": "fastq",
  "outdir": "<az://myoutdir>",
  "save_merged_fastq": false,
  "genome": "GRCh38",
  "gencode": false,
  "gtf_extra_attributes": "gene_name",
  "gtf_group_features": "gene_id",
  "save_reference": false,
  "igenomes_base": "<s3://ngi-igenomes/igenomes/>",
  "igenomes_ignore": false,
  "custom_config_version": "master",
  "custom_config_base": "<https://raw.githubusercontent.com/nf-core/configs/master>",
  "max_cpus": 16,
  "max_memory": "128.GB",
  "max_time": "240.h",
  "help": false,
  "version": false,
  "publish_dir_mode": "copy",
  "plaintext_email": false,
  "max_multiqc_email_size": "25.MB",
  "monochrome_logs": false,
  "validate_params": true,
  "validationShowHiddenParams": false,
  "validationFailUnrecognisedParams": false,
  "validationLenientMode": false,
  "save_trimmed": false,
  "skip_trimming": false,
  "skip_trimgalore_fastqc": false,
  "min_trimmed_reads": 10000,
  "skip_fastqc": false,
  "skip_bigwig": true,
  "aligner": "star_salmon",
  "pseudo_aligner": "salmon",
  "bam_csi_index": false,
  "star_ignore_sjdbgtf": false,
  "skip_alignment": false,
  "save_unaligned": false,
  "save_align_intermeds": false,
  "rmats": true,
  "rmats_splice_diff_cutoff": 0.0001,
  "rmats_paired_stats": true,
  "rmats_read_len": 40,
  "rmats_novel_splice_site": false,
  "rmats_min_intron_len": 50,
  "rmats_max_exon_len": 500,
  "dexseq_exon": true,
  "save_dexseq_annotation": false,
  "alignment_quality": 10,
  "aggregation": true,
  "save_dexseq_plot": true,
  "n_dexseq_plot": 10,
  "edger_exon": true,
  "save_edger_plot": true,
  "n_edger_plot": 10,
  "dexseq_dtu": true,
  "dtu_txi": "dtuScaledTPM",
  "min_samps_gene_expr": 4,
  "min_samps_feature_expr": 2,
  "min_samps_feature_prop": 2,
  "min_gene_expr": 10,
  "min_feature_expr": 10,
  "min_feature_prop": 0.1,
  "sashimi_plot": true,
  "miso_genes": "ENSG00000004961, ENSG00000005302, ENSG00000147403",
  "miso_genes_file": "",
  "miso_read_len": 75,
  "fig_height": 7,
  "fig_width": 7,
  "suppa": true,
  "suppa_per_local_event": true,
  "suppa_per_isoform": true,
  "generateevents_pool_genes": true,
  "generateevents_event_type": "SE SS MX RI FL",
  "generateevents_boundary": "S",
  "generateevents_threshold": 10,
  "generateevents_exon_length": 100,
  "psiperevent_total_filter": 0,
  "diffsplice_local_event": true,
  "diffsplice_isoform": true,
  "diffsplice_method": "empirical",
  "diffsplice_area": 1000,
  "diffsplice_lower_bound": 0,
  "diffsplice_gene_correction": true,
  "diffsplice_paired": true,
  "diffsplice_alpha": 0.05,
  "diffsplice_median": false,
  "diffsplice_tpm_threshold": 0,
  "diffsplice_nan_threshold": 0,
  "clusterevents_local_event": true,
  "clusterevents_isoform": true,
  "clusterevents_dpsithreshold": 0.05,
  "clusterevents_eps": 0.05,
  "clusterevents_metric": "euclidean",
  "clusterevents_min_pts": 20,
  "clusterevents_method": "DBSCAN"
}

moreinfo

To help we'd love a few more details please: 1. The Nextflow command you used to launch the pipeline 2. Any custom Nextflow config files 3. The full error message 4. The corresponding

.nextflow.log

file, where you are running the analysis. Remember to use triple backslashes on their own lines for code blocks. Also take a look at https://nf-co.re/usage/troubleshooting for more information on debugging failed pipelines. Thanks!

More info for attempt no.1: • Command line:

nextflow run '<https://github.com/nf-core/rnasplice>' \
		 -name RNASplice-Samplesheet-S2-tno1-Run01_11 \
		 -params-file '<https://api.tower.nf/ephemeral/34_-9wThVGzGcw3Az_B2_w.json>' \
		 -with-tower \
		 -r 2b7adfa697f8c2f34bd39f3dfb8ddfdb83fc390e \
		 -resume d5411dc7-2ed3-4c81-96a6-590a88a00751

• Attached: Console Output, Nextflow Log File and Execution Time Line • The full error message: Attached

How are you supplying the tools? Docker, Singularity/Apptainer, a Conda environment?

I'm using the Seqera Platform https://cloud.seqera.io/. @Thomas Danhorn

Sorry, I have never used that, but my understanding is that Seqera Platform is a "dashboard" that controls pipelines running e.g. in some kind of cloud environment or a cluster, which in turn would have some container system or conda. I'm wondering if your errors could be due to an incomplete installation of the software.

Thank you @Thomas Danhorn!. I believe that I may have missed something on the sample sheet and contrast sheet.

No, sorry, I some experience with splicing analysis, but just getting started running this pipeline. Your error is about some

GBR

object not found in R. If you sample sheet is invalid somehow, that should have ideally been caught at the validation step. It could be a bug in the code, possibly triggered by an unusual sample or contrast sheet, or something missing in the software, but I'm just guessing ...

Thank you everyone. it fixed. It was a fault in matching the contrast and condition tables between the sample sheet and the contrast sheet.