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nf-core / differentialabundance

Differential abundance analysis for feature/ observation matrices from platforms such as RNA-seq
https://nf-co.re/differentialabundance
MIT License
63 stars 37 forks source link

Normalized/Raw data mishap on the report #271

Closed rekren closed 5 months ago

rekren commented 5 months ago

Description of the bug

Hi there, I am using nf-core/differentialabundance v.1.5.0.

It seems that in the "Exploratory analysis" section of the "Results" part, Raw values and Normalized values needs to replace each other. It looks like they are switched by mistake of the generated Rmarkdown code.

here are the representative plots of the generated -suspiciously- looking plots. image image

Command used and terminal output

nextflow run nf-core/differentialabundance -r 1.5.0 --input dif_ab_samplesheet.csv --matrix outs/merged_gene_counts_table.tsv --transcript_length_matrix outs/merged_gene_lengths_table.tsv --contrasts dif_ab_contrast.csv --gtf /home/rekren/save/refs/Homo_sapiens.GRCh38.111.gtf.gz -profile genotoul --outdir DiffAbundance --report_author 'Rüçhan Ekren' --logo_file inf_logo_smaller.png --report_title 'Report' --report_description ' Explanation of the report' --shinyngs_build_app --report_scree

Relevant files

{ "input": "dif_ab_samplesheet.csv", "study_name": "study", "study_type": "rnaseq", "study_abundance_type": "counts", "contrasts": "dif_ab_contrast.csv", "querygse": null, "matrix": "outs/merged_gene_counts_table.tsv", "transcript_length_matrix": "outs/merged_gene_lengths_table.tsv", "control_features": null, "sizefactors_from_controls": false, "logo_file": "inf_logo_smaller.png", "css_file": "/home/rekren/.nextflow/assets/nf-core/differentialabundance/assets/nf-core_style.css", "citations_file": "/home/rekren/.nextflow/assets/nf-core/differentialabundance/CITATIONS.md", "report_file": "/home/rekren/.nextflow/assets/nf-core/differentialabundance/assets/differentialabundance_report.Rmd", "report_title": Title", "report_author": "R\u00fc\u00e7han Ekren", "report_contributors": null, "report_description": "This is description.", "report_scree": true, "report_round_digits": 4, "observations_type": "sample", "observations_id_col": "sample", "observations_name_col": null, "features": null, "features_type": "gene", "features_id_col": "gene_id", "features_name_col": "gene_name", "features_metadata_cols": "gene_id,gene_name,gene_biotype", "features_gtf_feature_type": "transcript", "features_gtf_table_first_field": "gene_id", "affy_cel_files_archive": null, "affy_file_name_col": "file", "affy_background": true, "affy_bgversion": 2, "affy_destructive": false, "affy_cdfname": null, "affy_rm_mask": false, "affy_rm_outliers": false, "affy_rm_extra": false, "affy_build_annotation": true, "proteus_measurecol_prefix": "LFQ intensity", "proteus_norm_function": "normalizeMedian", "proteus_plotsd_method": "violin", "proteus_plotmv_loess": true, "proteus_palette_name": "Set1", "filtering_min_samples": 1.0, "filtering_min_abundance": 1.0, "filtering_min_proportion": null, "filtering_grouping_var": null, "filtering_min_proportion_not_na": 0.5, "filtering_min_samples_not_na": null, "exploratory_main_variable": "auto_pca", "exploratory_clustering_method": "ward.D2", "exploratory_cor_method": "spearman", "exploratory_n_features": 500, "exploratory_whisker_distance": 1.5, "exploratory_mad_threshold": -5, "exploratory_assay_names": "raw,normalised,variance_stabilised", "exploratory_final_assay": "variance_stabilised", "exploratory_log2_assays": "raw,normalised", "exploratory_palette_name": "Set1", "differential_file_suffix": null, "differential_feature_id_column": "gene_id", "differential_feature_name_column": "gene_name", "differential_fc_column": "log2FoldChange", "differential_pval_column": "pvalue", "differential_qval_column": "padj", "differential_min_fold_change": 2.0, "differential_max_pval": 1.0, "differential_max_qval": 0.05, "differential_foldchanges_logged": true, "differential_palette_name": "Set1", "differential_subset_to_contrast_samples": false, "deseq2_test": "Wald", "deseq2_fit_type": "parametric", "deseq2_sf_type": "ratio", "deseq2_min_replicates_for_replace": 7, "deseq2_use_t": false, "deseq2_lfc_threshold": 0, "deseq2_alt_hypothesis": "greaterAbs", "deseq2_independent_filtering": true, "deseq2_p_adjust_method": "BH", "deseq2_alpha": 0.1, "deseq2_minmu": 0.5, "deseq2_vs_method": "vst", "deseq2_shrink_lfc": true, "deseq2_cores": 1, "deseq2_vs_blind": true, "deseq2_vst_nsub": 1000, "limma_ndups": null, "limma_spacing": null, "limma_block": null, "limma_correlation": null, "limma_method": "ls", "limma_proportion": 0.01, "limma_stdev_coef_lim": "0.1,4", "limma_trend": false, "limma_robust": false, "limma_winsor_tail_p": "0.05,0.1", "limma_adjust_method": "BH", "limma_p_value": 1.0, "limma_lfc": 0, "limma_confint": false, "gsea_run": false, "gsea_nperm": 1000, "gsea_permute": "phenotype", "gsea_scoring_scheme": "weighted", "gsea_metric": "Signal2Noise", "gsea_sort": "real", "gsea_order": "descending", "gsea_set_max": 500, "gsea_set_min": 15, "gsea_norm": "meandiv", "gsea_rnd_type": "no_balance", "gsea_make_sets": true, "gsea_median": false, "gsea_num": 100, "gsea_plot_top_x": 20, "gsea_rnd_seed": "timestamp", "gsea_save_rnd_lists": false, "gsea_zip_report": false, "gprofiler2_run": false, "gprofiler2_organism": null, "gprofiler2_significant": true, "gprofiler2_measure_underrepresentation": false, "gprofiler2_correction_method": "gSCS", "gprofiler2_sources": null, "gprofiler2_evcodes": false, "gprofiler2_max_qval": 0.05, "gprofiler2_token": null, "gprofiler2_background_file": "auto", "gprofiler2_background_column": null, "gprofiler2_domain_scope": "annotated", "gprofiler2_min_diff": 1, "gprofiler2_palette_name": "Blues", "shinyngs_build_app": true, "shinyngs_guess_unlog_matrices": true, "shinyngs_deploy_to_shinyapps_io": false, "shinyngs_shinyapps_account": null, "shinyngs_shinyapps_app_name": null, "gene_sets_files": null, "genome": null, "igenomes_base": "s3://ngi-igenomes/igenomes/", "igenomes_ignore": true, "outdir": "DiffAbundance", "publish_dir_mode": "copy", "email": null, "email_on_fail": null, "plaintext_email": false, "monochrome_logs": false, "hook_url": null, "help": false, "version": false, "config_profile_name": null, "config_profile_description": "The Genotoul cluster profile", "custom_config_version": "master", "gtf": "/home/rekren/save/refs/Homo_sapiens.GRCh38.111.gtf.gz", "custom_config_base": "https://raw.githubusercontent.com/nf-core/configs/master", "config_profile_contact": "support.bioinfo.genotoul@inra.fr", "config_profile_url": "http://bioinfo.genotoul.fr/", "max_memory": { "mega": 122880, "kilo": 125829120, "giga": 120, "bytes": 128849018880 }, .nextflow.log

"max_cpus": 48,
"max_time": {
    "days": 4,
    "millis": 345600000,
    "hours": 96,
    "minutes": 5760,
    "seconds": 345600,
    "durationInMillis": 345600000
},
"validationFailUnrecognisedParams": false,
"validation-fail-unrecognised-params": false,
"validationLenientMode": false,
"validation-lenient-mode": false,
"validationSchemaIgnoreParams": "genomes,igenomes_base",
"validation-schema-ignore-params": "genomes,igenomes_base",
"validationShowHiddenParams": false,
"validation-show-hidden-params": false,
"validate_params": true,
"save_reference": true,
"igenomesIgnore": true,
"igenomes-ignore": true,
"genomes": {

}

}

System information

Nextflow version : version 23.10.0, build 5889 Hardware: HPC (Genotoul) Executor: Slurm Container: Singularity OS: Linux Version of nf-core/differentialabundance : 1.5

rekren commented 5 months ago

@pinin4fjords and Oskar Wacker checked the label assignments with the test data on test profile, everything seemed good.

I have tested with another dataset and also tested with nf-core/differential_abundance v1.4. They were seeming appropriately assigning the assay names. I think everything is indeed in order and this was only visual "mishap". For the above figures when I check the actual png figures they were looking a bit better than the rendered plots of the report. image image

For this specific dataset, I believe data plots shrunken a little bit more than usual and made us question the correctness of the labels on the report rendering part .

Closing the issue with this comments.