featurecounts-rna-counting

npx skills add BioTender-max/awesome-bio-agent-skills --skill featurecounts-rna-counting --agent claude-code

Same command for any agent — swap --agent for codex, cursor, copilot.

# featureCounts — RNA-seq Read Counting ## Overview featureCounts (part of the Subread package) assigns sequencing reads in BAM files to genomic features defined in a GTF/GFF annotation. It counts how many reads overlap each gene (or exon, intron, or custom feature), producing a gene × sample count matrix suitable for differential expression analysis with DESeq2 or edgeR. featureCounts processes multiple BAM files in a single command, reporting read assignment statistics (assigned, unassigned by category) alongside the count matrix. It is the standard counting step after STAR alignment in RNA-seq pipelines. ## When to Use - Generating gene-level count matrices from STAR-aligned BAM files for DESeq2 or edgeR - Counting reads from multiple samples simultaneously in a single featureCounts command - Handling stranded RNA-seq libraries where sense/antisense assignment matters - Producing exon-level or custom-feature counts (e.g., for splicing analysis with DEXSeq) - Verifying strandedness of an RNA-seq library when protocol documentation is unavailable - Use **Salmon** instead when no BAM file exists and fast pseudoalignment is preferred - Use **HTSeq-count** as an alternative with slow

Install with conda (recommended)
conda install -c bioconda subread
Verify
featureCounts -v
featureCounts v2.0.6
sudo apt-get install subread
Count reads for multiple samples (unstranded paired-end RNA-seq)
featureCounts \
results/sample1/Aligned.sortedByCoord.out.bam \
results/sample2/Aligned.sortedByCoord.out.bam