Functional Genomics, Screening, and Causal Gene Discovery
From Hypothesis to Genome-Wide Perturbation
Scope
Functional genomics links large-scale perturbation methods with sequencing-based readouts to infer which genes, elements, or interactions control a phenotype.
Screen types
Loss-of-function screen
- Identifies genes required for a phenotype
- Often uses CRISPR knockout or interference
- Can reveal essential genes
Gain-of-function screen
- Tests sufficiency or activation effects
- Often uses CRISPR activation or cDNA libraries
- Can uncover dosage-sensitive drivers
Which CRISPR modality is commonly used to repress transcription without cutting DNA?
CRISPR interference uses catalytically dead Cas proteins fused to repressors to reduce transcription.
Correct answer: CRISPRi
Why are pooled screens powerful?
This enables scalable discovery across the genome.
Correct answer: They allow many perturbations to be tested in parallel with sequencing-based readout
Readout logic
After perturbation, the phenotype may be inferred from cell abundance, reporter intensity, transcriptomic state, survival, or lineage barcodes.
Common screening readouts
| Readout | What it measures |
|---|---|
| Cell survival | Fitness under selection |
| Fluorescence | Reporter or pathway activity |
| Sequencing abundance | Representation of guide or barcode |
| Single-cell RNA-seq | Transcriptomic response per cell |
What is a major advantage of single-cell readouts in perturbation screens?
Single-cell methods reveal heterogeneous responses hidden in bulk measurements.
Correct answer: They preserve cellular heterogeneity information
What does CRISPRa do?
CRISPR activation recruits transcriptional activators to the target locus.
Correct answer: It activates gene expression using a targeted DNA-binding platform