@prefix dcterms: <http://purl.org/dc/terms/> .
@prefix np: <http://www.nanopub.org/nschema#> .
@prefix npx: <http://purl.org/nanopub/x/> .
@prefix ns1: <https://w3id.org/np/snakemake/> .
@prefix orcid: <https://orcid.org/> .
@prefix prov: <http://www.w3.org/ns/prov#> .
@prefix rdfs: <http://www.w3.org/2000/01/rdf-schema#> .
@prefix schema: <https://schema.org/> .
@prefix sub: <https://w3id.org/np/RAK9xz_ccnu0Xhs4vX2KtqCxX44mmSt6nq-ePLeewMrFE/> .
@prefix this: <https://w3id.org/np/RAK9xz_ccnu0Xhs4vX2KtqCxX44mmSt6nq-ePLeewMrFE> .
@prefix xsd: <http://www.w3.org/2001/XMLSchema#> .

sub:Head {
  this: a np:Nanopublication;
    np:hasAssertion sub:assertion;
    np:hasProvenance sub:provenance;
    np:hasPublicationInfo sub:pubinfo .
}

sub:assertion {
  sub:config-1 dcterms:identifier "config.yml";
    schema:text """samples: samples.csv

ref:
    species: \"Drosophila melanogaster\"
    genome: \"\"
    annotation: \"\"
    accession: \"GCF_000001215.4\"
    ensembl_species: \"\"  # e.g., \"homo_sapiens\"
    build: \"\"  # e.g., \"GRCh38\"
    release: \"\"  # e.g., \"105\"

read_filter:
    min_length: 200

minimap2:
    index_opts: \"\"
    opts: \"\"
    maximum_secondary: 100
    secondary_score_ratio: 1.0

samtools:
    samtobam_opts: \"-b\"
    bamsort_opts: \"\"
    bamindex_opts: \"\"
    bamstats_opts: \"\"

quant:
    salmon_libtype: \"U\"

deseq2:
    fit_type: \"\"
    design_factors:
        - \"condition\"
    lfc_null: 1.0
    alt_hypothesis: \"greaterAbs\"
    point_width: 20
    mincount: 10
    alpha: 0.05
    threshold_plot: 10
    colormap: \"Blues\"
    figtype: \"png\"
    batch_effect:
      - \"\"

isoform_analysis:
    FLAIR: true
    qscore: 1
    exp_thresh: 10
    col_opts: \"--annotation_reliant generate --generate_map --stringent\"

protein_annotation:
    lambda: false
    uniref: \"https://ftp.imp.fu-berlin.de/pub/lambda/index/lambda3/gen_0/uniref50_20230713.lba.gz\"
    num_matches: 3""" .
  
  sub:dataset a schema:Dataset;
    ns1:describesWorkflow "RAjHDlPDghZzc9ZvQ3uJQNJ9Jd_KAYzZt7dk5PXKgjRyE";
    ns1:description """This workflow performs differential expression analysis of RNA-seq data obtained from Oxford Nanopore long-read sequencing technology.
First a transcriptome FASTA is constructed using gffread [https://github.com/gpertea/gffread]. Reads are then mapped to the transcriptome with the long-read optimized alignment tool minimap2 [https://github.com/lh3/minimap2].
Next quantification is performed using salmon [https://github.com/COMBINE-lab/salmon] before normalization and differential expression analysis are conducted by PyDESeq2 [https://github.com/owkin/PyDESeq2].
The workflow can optionally analyze splice-isoforms through integrating the FLAIR [https://github.com/BrooksLabUCSC/flair] workflow.
Additionaly, NanoPlot [https://github.com/wdecoster/NanoPlot] is employed to analyze initial sequencing data and QualiMap [https://github.com/EagleGenomics-cookbooks/QualiMap] is used to evaluate mapping results.""";
    ns1:generatedAt "2026-04-17T15:19:46.851430+00:00"^^xsd:dateTime;
    ns1:hasConfigurationSection sub:workflow-configuration .
  
  sub:job-1-alignment_qa a ns1:WorkflowJob;
    ns1:hasOutput "QC/qualimap/barcode10";
    ns1:ruleName "alignment_qa" .
  
  sub:job-10-alignment_qa_report a ns1:WorkflowJob;
    ns1:hasOutput "qualimap/barcode13/qualimapReport.html";
    ns1:ruleName "alignment_qa_report" .
  
  sub:job-11-alignment_qa_report a ns1:WorkflowJob;
    ns1:hasOutput "qualimap/barcode15/qualimapReport.html";
    ns1:ruleName "alignment_qa_report" .
  
  sub:job-12-alignment_qa_report a ns1:WorkflowJob;
    ns1:hasOutput "qualimap/barcode16/qualimapReport.html";
    ns1:ruleName "alignment_qa_report" .
  
  sub:job-13-bam_sort a ns1:WorkflowJob;
    ns1:hasOutput "sorted_alignments/barcode10_sorted.bam";
    ns1:ruleName "bam_sort" .
  
  sub:job-14-bam_sort a ns1:WorkflowJob;
    ns1:hasOutput "sorted_alignments/barcode11_sorted.bam";
    ns1:ruleName "bam_sort" .
  
  sub:job-15-bam_sort a ns1:WorkflowJob;
    ns1:hasOutput "sorted_alignments/barcode12_sorted.bam";
    ns1:ruleName "bam_sort" .
  
  sub:job-16-bam_sort a ns1:WorkflowJob;
    ns1:hasOutput "sorted_alignments/barcode13_sorted.bam";
    ns1:ruleName "bam_sort" .
  
  sub:job-17-bam_sort a ns1:WorkflowJob;
    ns1:hasOutput "sorted_alignments/barcode15_sorted.bam";
    ns1:ruleName "bam_sort" .
  
  sub:job-18-bam_sort a ns1:WorkflowJob;
    ns1:hasOutput "sorted_alignments/barcode16_sorted.bam";
    ns1:ruleName "bam_sort" .
  
  sub:job-19-bam_stats a ns1:WorkflowJob;
    ns1:hasOutput "QC/bamstats/barcode10.txt";
    ns1:ruleName "bam_stats" .
  
  sub:job-2-alignment_qa a ns1:WorkflowJob;
    ns1:hasOutput "QC/qualimap/barcode11";
    ns1:ruleName "alignment_qa" .
  
  sub:job-20-bam_stats a ns1:WorkflowJob;
    ns1:hasOutput "QC/bamstats/barcode11.txt";
    ns1:ruleName "bam_stats" .
  
  sub:job-21-bam_stats a ns1:WorkflowJob;
    ns1:hasOutput "QC/bamstats/barcode12.txt";
    ns1:ruleName "bam_stats" .
  
  sub:job-22-bam_stats a ns1:WorkflowJob;
    ns1:hasOutput "QC/bamstats/barcode13.txt";
    ns1:ruleName "bam_stats" .
  
  sub:job-23-bam_stats a ns1:WorkflowJob;
    ns1:hasOutput "QC/bamstats/barcode15.txt";
    ns1:ruleName "bam_stats" .
  
  sub:job-24-bam_stats a ns1:WorkflowJob;
    ns1:hasOutput "QC/bamstats/barcode16.txt";
    ns1:ruleName "bam_stats" .
  
  sub:job-25-build_flair_genome_index a ns1:WorkflowJob;
    ns1:hasOutput "index/flair_genome_index.mmi";
    ns1:ruleName "build_flair_genome_index" .
  
  sub:job-26-build_minimap_index a ns1:WorkflowJob;
    ns1:hasOutput "index/transcriptome_index.mmi";
    ns1:ruleName "build_minimap_index" .
  
  sub:job-27-correct_transcriptome a ns1:WorkflowJob;
    ns1:hasOutput "transcriptome/corrected_transcriptome.fa";
    ns1:ruleName "correct_transcriptome" .
  
  sub:job-28-count_reads a ns1:WorkflowJob;
    ns1:hasOutput "counts/barcode10_salmon/quant.sf";
    ns1:ruleName "count_reads" .
  
  sub:job-29-count_reads a ns1:WorkflowJob;
    ns1:hasOutput "counts/barcode11_salmon/quant.sf";
    ns1:ruleName "count_reads" .
  
  sub:job-3-alignment_qa a ns1:WorkflowJob;
    ns1:hasOutput "QC/qualimap/barcode12";
    ns1:ruleName "alignment_qa" .
  
  sub:job-30-count_reads a ns1:WorkflowJob;
    ns1:hasOutput "counts/barcode12_salmon/quant.sf";
    ns1:ruleName "count_reads" .
  
  sub:job-31-count_reads a ns1:WorkflowJob;
    ns1:hasOutput "counts/barcode13_salmon/quant.sf";
    ns1:ruleName "count_reads" .
  
  sub:job-32-count_reads a ns1:WorkflowJob;
    ns1:hasOutput "counts/barcode15_salmon/quant.sf";
    ns1:ruleName "count_reads" .
  
  sub:job-33-count_reads a ns1:WorkflowJob;
    ns1:hasOutput "counts/barcode16_salmon/quant.sf";
    ns1:ruleName "count_reads" .
  
  sub:job-34-deseq2 a ns1:WorkflowJob;
    ns1:hasOutput "de_analysis/condition_wt_vs_mt_MA_plot.svg", "de_analysis/condition_wt_vs_mt_count_heatmap.svg",
      "de_analysis/condition_wt_vs_mt_dispersion_plot.svg", "de_analysis/condition_wt_vs_mt_l2fc.tsv",
      "de_analysis/condition_wt_vs_mt_sample_heatmap.svg", "de_analysis/condition_wt_vs_mt_top_count_heatmap.svg";
    ns1:ruleName "deseq2" .
  
  sub:job-35-deseq2_init a ns1:WorkflowJob;
    ns1:hasOutput "de_analysis/all.rds", "de_analysis/normcounts.tsv";
    ns1:ruleName "deseq2_init" .
  
  sub:job-36-download_ncbi_annotation a ns1:WorkflowJob;
    ns1:hasOutput "references/ncbi_dataset_annotation.zip";
    ns1:ruleName "download_ncbi_annotation" .
  
  sub:job-37-download_ncbi_genome a ns1:WorkflowJob;
    ns1:hasOutput "references/ncbi_dataset_genome.zip";
    ns1:ruleName "download_ncbi_genome" .
  
  sub:job-38-filter_reads a ns1:WorkflowJob;
    ns1:hasOutput "filter/barcode10_filtered.fq";
    ns1:ruleName "filter_reads" .
  
  sub:job-39-filter_reads a ns1:WorkflowJob;
    ns1:hasOutput "filter/barcode11_filtered.fq";
    ns1:ruleName "filter_reads" .
  
  sub:job-4-alignment_qa a ns1:WorkflowJob;
    ns1:hasOutput "QC/qualimap/barcode13";
    ns1:ruleName "alignment_qa" .
  
  sub:job-40-filter_reads a ns1:WorkflowJob;
    ns1:hasOutput "filter/barcode12_filtered.fq";
    ns1:ruleName "filter_reads" .
  
  sub:job-41-filter_reads a ns1:WorkflowJob;
    ns1:hasOutput "filter/barcode13_filtered.fq";
    ns1:ruleName "filter_reads" .
  
  sub:job-42-filter_reads a ns1:WorkflowJob;
    ns1:hasOutput "filter/barcode15_filtered.fq";
    ns1:ruleName "filter_reads" .
  
  sub:job-43-filter_reads a ns1:WorkflowJob;
    ns1:hasOutput "filter/barcode16_filtered.fq";
    ns1:ruleName "filter_reads" .
  
  sub:job-44-flair_align a ns1:WorkflowJob;
    ns1:hasOutput "iso_analysis/align/flair.bam", "iso_analysis/align/flair.bam.bai",
      "iso_analysis/align/flair.bed";
    ns1:ruleName "flair_align" .
  
  sub:job-45-flair_collapse a ns1:WorkflowJob;
    ns1:hasOutput "iso_analysis/collapse/flair.isoforms.bed", "iso_analysis/collapse/flair.isoforms.fa";
    ns1:ruleName "flair_collapse" .
  
  sub:job-46-flair_correct a ns1:WorkflowJob;
    ns1:hasOutput "iso_analysis/align/flair_all_corrected.bed";
    ns1:ruleName "flair_correct" .
  
  sub:job-47-flair_diffexp a ns1:WorkflowJob;
    ns1:hasOutput "iso_analysis/diffexp/genes_deseq2_QCplots_wt_v_mt.pdf", "iso_analysis/diffexp/genes_deseq2_wt_v_mt.tsv",
      "iso_analysis/diffexp/isoforms_deseq2_QCplots_wt_v_mt.pdf", "iso_analysis/diffexp/isoforms_deseq2_wt_v_mt.tsv",
      "iso_analysis/diffexp/isoforms_drimseq_wt_v_mt.tsv";
    ns1:ruleName "flair_diffexp" .
  
  sub:job-48-flair_plot_isoforms a ns1:WorkflowJob;
    ns1:hasOutput "iso_analysis/plots";
    ns1:ruleName "flair_plot_isoforms" .
  
  sub:job-49-flair_quantify a ns1:WorkflowJob;
    ns1:hasOutput "iso_analysis/quantify/flair.counts.tsv";
    ns1:ruleName "flair_quantify" .
  
  sub:job-5-alignment_qa a ns1:WorkflowJob;
    ns1:hasOutput "QC/qualimap/barcode15";
    ns1:ruleName "alignment_qa" .
  
  sub:job-50-genome_to_transcriptome a ns1:WorkflowJob;
    ns1:hasOutput "references/genomic.fa.fai", "transcriptome/transcriptome.fa";
    ns1:ruleName "genome_to_transcriptome" .
  
  sub:job-51-get_annotation a ns1:WorkflowJob;
    ns1:hasOutput "references/genomic.gff";
    ns1:ruleName "get_annotation" .
  
  sub:job-52-get_genome a ns1:WorkflowJob;
    ns1:hasOutput "references/genomic.fa";
    ns1:ruleName "get_genome" .
  
  sub:job-53-gff_to_gtf a ns1:WorkflowJob;
    ns1:hasOutput "references/standardized_genomic.gtf";
    ns1:ruleName "gff_to_gtf" .
  
  sub:job-54-iso_analysis_report a ns1:WorkflowJob;
    ns1:hasOutput "iso_analysis/report/isoforms", "iso_analysis/report/usage";
    ns1:ruleName "iso_analysis_report" .
  
  sub:job-55-map_reads a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode10.sam";
    ns1:ruleName "map_reads" .
  
  sub:job-56-map_reads a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode11.sam";
    ns1:ruleName "map_reads" .
  
  sub:job-57-map_reads a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode12.sam";
    ns1:ruleName "map_reads" .
  
  sub:job-58-map_reads a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode13.sam";
    ns1:ruleName "map_reads" .
  
  sub:job-59-map_reads a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode15.sam";
    ns1:ruleName "map_reads" .
  
  sub:job-6-alignment_qa a ns1:WorkflowJob;
    ns1:hasOutput "QC/qualimap/barcode16";
    ns1:ruleName "alignment_qa" .
  
  sub:job-60-map_reads a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode16.sam";
    ns1:ruleName "map_reads" .
  
  sub:job-61-merge_read_counts a ns1:WorkflowJob;
    ns1:hasOutput "merged/all_counts.tsv";
    ns1:ruleName "merge_read_counts" .
  
  sub:job-62-reads_manifest a ns1:WorkflowJob;
    ns1:hasOutput "iso_analysis/reads_manifest.tsv";
    ns1:ruleName "reads_manifest" .
  
  sub:job-63-sam_to_bam a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode10.bam";
    ns1:ruleName "sam_to_bam" .
  
  sub:job-64-sam_to_bam a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode11.bam";
    ns1:ruleName "sam_to_bam" .
  
  sub:job-65-sam_to_bam a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode12.bam";
    ns1:ruleName "sam_to_bam" .
  
  sub:job-66-sam_to_bam a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode13.bam";
    ns1:ruleName "sam_to_bam" .
  
  sub:job-67-sam_to_bam a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode15.bam";
    ns1:ruleName "sam_to_bam" .
  
  sub:job-68-sam_to_bam a ns1:WorkflowJob;
    ns1:hasOutput "alignments/barcode16.bam";
    ns1:ruleName "sam_to_bam" .
  
  sub:job-69-sample_qa_plot a ns1:WorkflowJob;
    ns1:hasOutput "NanoPlot/barcode10/NanoPlot-report.html";
    ns1:ruleName "sample_qa_plot" .
  
  sub:job-7-alignment_qa_report a ns1:WorkflowJob;
    ns1:hasOutput "qualimap/barcode10/qualimapReport.html";
    ns1:ruleName "alignment_qa_report" .
  
  sub:job-70-sample_qa_plot a ns1:WorkflowJob;
    ns1:hasOutput "NanoPlot/barcode11/NanoPlot-report.html";
    ns1:ruleName "sample_qa_plot" .
  
  sub:job-71-sample_qa_plot a ns1:WorkflowJob;
    ns1:hasOutput "NanoPlot/barcode12/NanoPlot-report.html";
    ns1:ruleName "sample_qa_plot" .
  
  sub:job-72-sample_qa_plot a ns1:WorkflowJob;
    ns1:hasOutput "NanoPlot/barcode13/NanoPlot-report.html";
    ns1:ruleName "sample_qa_plot" .
  
  sub:job-73-sample_qa_plot a ns1:WorkflowJob;
    ns1:hasOutput "NanoPlot/barcode15/NanoPlot-report.html";
    ns1:ruleName "sample_qa_plot" .
  
  sub:job-74-sample_qa_plot a ns1:WorkflowJob;
    ns1:hasOutput "NanoPlot/barcode16/NanoPlot-report.html";
    ns1:ruleName "sample_qa_plot" .
  
  sub:job-75-standardize_gff a ns1:WorkflowJob;
    ns1:hasOutput "references/standardized_genomic.gff";
    ns1:ruleName "standardize_gff" .
  
  sub:job-76-total_sample_qa_plot a ns1:WorkflowJob;
    ns1:hasOutput "NanoPlot/NanoPlot-report.html";
    ns1:ruleName "total_sample_qa_plot" .
  
  sub:job-77-transcriptid_to_gene a ns1:WorkflowJob;
    ns1:hasOutput "merged/all_counts_gene.tsv", "merged/transcriptid_to_gene_plot.svg";
    ns1:ruleName "transcriptid_to_gene" .
  
  sub:job-8-alignment_qa_report a ns1:WorkflowJob;
    ns1:hasOutput "qualimap/barcode11/qualimapReport.html";
    ns1:ruleName "alignment_qa_report" .
  
  sub:job-9-alignment_qa_report a ns1:WorkflowJob;
    ns1:hasOutput "qualimap/barcode12/qualimapReport.html";
    ns1:ruleName "alignment_qa_report" .
  
  sub:workflow-configuration rdfs:label "from workflow configuration";
    ns1:hasConfigurationFile sub:config-1 .
}

sub:provenance {
  sub:assertion prov:generatedAtTime "2026-04-17T17:19:48.418353"^^xsd:dateTime;
    prov:wasAttributedTo orcid:0000-0003-2408-7588 .
}

sub:pubinfo {
  this: dcterms:created "2026-04-17T15:19:46.851430+00:00"^^xsd:dateTime;
    dcterms:creator orcid:0000-0003-2408-7588;
    npx:hasNanopubType schema:Dataset;
    npx:signedBy orcid:0000-0003-2408-7588;
    rdfs:label "Snakemake workflow metadata: RAjHDlPDghZzc9ZvQ3uJQNJ9Jd_KAYzZt7dk5PXKgjRyE";
    prov:generatedAtTime "2026-04-17T17:19:48.418353"^^xsd:dateTime;
    prov:wasAttributedTo orcid:0000-0003-2408-7588 .
  
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}