Quantify Viral Gene Expression in RNA-seq: STAR & featureCounts

Описание: Master viral gene expression quantification in host RNA-seq data! Learn to create dual-genome references (viral + human), align reads with STAR, quantify viral transcripts with featureCounts, and perform differential expression analysis of viral genes.

📚 FULL TUTORIAL WITH CODE:
https://ngs101.com/how-to-analyze-rna...

⏱️ TIMESTAMPS:
0:00 - Introduction: Two Strategies for Viral Quantification
1:46 - Strategy 1: Build Viral-Only Reference Index
4:38 - Strategy 1: Align Reads to Viral Genome with STAR
6:09 - Strategy 2: Build Integrated Viral + Host Reference
7:11 - Quantify Viral Gene Expression with featureCounts

🔬 WHAT YOU'LL LEARN:
✅ Build combined viral + host genome references
✅ Prepare viral GTF annotation files
✅ Align RNA-seq reads to dual genomes with STAR
✅ Quantify viral gene expression with featureCounts
✅ Separate host vs viral read counts
✅ Perform differential viral gene expression
✅ Analyze viral gene regulation patterns
✅ Interpret viral transcript dynamics
✅ Compare viral expression across samples

🧬 COMPLETE WORKFLOW:

*Step 1: Reference Preparation*
→ Download viral genome sequences (NCBI/ViralZone)
→ Download viral annotation (GTF format)
→ Combine with human genome reference
→ Build STAR index for dual genome

*Step 2: Alignment to Dual Genome*
→ Align reads with STAR to combined reference
→ Generate sorted BAM files
→ Separate host-mapped vs virus-mapped reads
→ Quality control on alignment rates

*Step 3: Viral Gene Quantification*
→ Run featureCounts on viral genes
→ Extract viral gene count matrix
→ Calculate viral gene TPM/FPKM
→ Quantify viral vs host read ratios

*Step 4: Differential Expression*
→ Apply limma/DESeq2/edgeR to viral counts
→ Identify differentially expressed viral genes
→ Analyze viral gene co-expression patterns
→ Correlate with host gene expression

🔧 SOFTWARE & TOOLS:
STAR aligner - dual genome alignment
featureCounts - viral gene quantification
samtools - BAM file processing
limma/DESeq2/edgeR - differential expression
R/Bioconductor - statistical analysis
IGV - visualization of viral alignments