Merge pull request #582 from pavanvidem/rnaseq-de

first release of RNAseq DE analysis, filtering and plotting workflow

workflows/transcriptomics/rnaseq-de/.dockstore.yml

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+version: 1.2
+workflows:
+- name: main
+  subclass: Galaxy
+  publish: true
+  primaryDescriptorPath: /rnaseq-de-filtering-plotting.ga
+  testParameterFiles:
+  - /rnaseq-de-filtering-plotting-tests.yml
+  authors:
+  - name: Pavankumar Videm
+    orcid: 0000-0002-5192-126X

workflows/transcriptomics/rnaseq-de/CHANGELOG.md

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+## [0.1] 2024-10-25
+
+First release.

workflows/transcriptomics/rnaseq-de/README.md

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+# RNA-seq Differential expression and filtering workflow
+
+This workflow works only with an experimental setup containing exactly 2 conditions with at least 2 replicates per condition.
+
+## Inputs dataset
+
+- Counts from changed condition: Counts from experimental condition or changed condition. For eg. counts from treatment or knockdown samples.
+- Counts from reference condition: Counts from reference condition or base condition. For eg. counts from untreated or wildtype samples.
+- Gene Annotaton: The same GTF file used for mapping and quantification. It is used to annotate the DESeq2 results table. Ideally, the GTF file should contain `gene_id`, `gene_biotype` and `gene_name` attributes.
+
+## Inputs values
+
+- Count files have header: Indicate whether your input count files have a header line. Usually, count files generated from featureCounts tool have a header line whereas count files from RNA-STAR do not have.
+- Adjusted p-value threshold: All the genes with an adjusted p-value less than this value are considered as differentially expressed. With a value of 0.05, expect 5% of false positives in the differentially expressed genes list. If empty, a default value of 0.05 is used.
+- log2 fold change threshold: All the genes with an absolute fold change (regarless of up or down regulation) more than this value are selected. A log2 FC of 3 equals to an absolute fold change of 8 (2³). If empty, a default value of 1.0 is used.
+
+## Processing
+
+- The workflow uses DESeq2 for performing differential expression analysis. In addition to the results table, it also produces normalized counts table.
+- The results table is annotated with gene positions, biotypes, gene symbols.
+- The annotated results table is further filtered with the input adjusted p-value and log2 fold change thresholds.
+- A valcano plot is generated with top 10 significantly differentially expressed genes.
+- A heatmap of log trasformed normalized counts and another heatmap of Z-scores is generated.

workflows/transcriptomics/rnaseq-de/rnaseq-de-filtering-plotting-tests.yml

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+- doc: Test outline for RNAseq_DE_filtering_plotting
+  job:
+    Gene Annotaton:
+      class: File
+      location: https://zenodo.org/records/14056162/files/Saccharomyces_cerevisiae.R64-1-1.113.gtf
+      filetype: gtf
+    Counts from changed condition:
+      class: Collection
+      collection_type: list
+      elements:
+      - class: File
+        identifier: SRR5085169 Counts Table
+        location: https://zenodo.org/records/14056162/files/SRR5085169.tabular
+      - class: File
+        identifier: SRR5085170 Counts Table
+        location: https://zenodo.org/records/14056162/files/SRR5085170.tabular
+    Counts from reference condition:
+      class: Collection
+      collection_type: list
+      elements:
+      - class: File
+        identifier: SRR5085167 Counts Table
+        location: https://zenodo.org/records/14056162/files/SRR5085167.tabular
+      - class: File
+        identifier: SRR5085168 Counts Table
+        location: https://zenodo.org/records/14056162/files/SRR5085168.tabular
+    Count files have header: true
+    Adjusted p-value threshold: 0.1
+    log2 fold change threshold: 0.5
+  outputs:
+    Annotated DESeq2 results table:
+        has_text_matching:
+            expression: "YML123C\t122.984408142053\t-1.67[0-9]*\t0.21[0-9]*\t-7.66[0-9]*\t1.81[0-9]*e-14\t5.04[0-9]*e-[0-9]*\tchrXIII\t24036\t25800\t-\tprotein_coding\tPHO84"
+            expression: "YKL081W\t264.71[0-9]*\t-0.54[0-9]*\t0.15[0-9]*\t-3.46[0-9]*\t0.00[0-9]*\t0.09[0-9]*\tchrXI\t282890\t284455\t+\tprotein_coding\tTEF4"
+    Heatmap of Z-scores:
+      has_size:
+        value: 19510
+        delta: 1000
+    DESeq2 Normalized Counts:
+        has_text_matching:
+            expression: "YML123C\t210.50[0-9]*\t180.36[0-9]*\t48.64[0-9]*\t52.43[0-9]*"
+            expression: "YKL081W\t313.76[0-9]*\t322.37[0-9]*\t223.48[0-9]*\t199.24[0-9]*"
+    DESeq2 Plots:
+      has_size:
+        value: 1193021
+        delta: 60000
+    Volcano Plot of DE genes:
+      has_size:
+        value: 301346
+        delta: 15000
+    Heatmap of log transformed normalized counts:
+      has_size:
+        value: 19501
+        delta: 1000