Reporter Types
LacZ (Beta Galactosidase)
LacZ encodes bacterial beta galactosidase, which converts X gal substrate to an insoluble blue precipitate:
LacZ remains the most widely used reporter for detailed expression pattern analysis in fixed tissues.
Fluorescent Proteins
Fluorescent protein reporters enable live imaging and flow cytometry:
Luciferase
Luciferase reporters produce light through enzymatic reaction with luciferin substrate:
Luciferase is ideal for tracking gene expression dynamics in living animals without sacrifice.
Fluorescent Protein Options
| Reporter | Color | Excitation/Emission | Applications |
|---|---|---|---|
| EGFP | Green | 488/507 nm | General purpose, live imaging |
| tdTomato | Red | 554/581 nm | Bright, photostable, lineage tracing |
| mCherry | Red | 587/610 nm | Live imaging, FRET |
| YFP | Yellow | 514/527 nm | Multicolor imaging |
| CFP | Cyan | 433/475 nm | FRET donor |
| mScarlet | Red | 569/594 nm | Bright, monomeric |
Reporter Comparison
| Factor | LacZ | Fluorescent Protein | Luciferase |
|---|---|---|---|
| Live imaging | No | Yes | Yes |
| Cellular resolution | Yes | Yes | Limited |
| Tissue sections | Yes | Yes | No |
| Flow cytometry | No | Yes | No |
| In vivo imaging | No | Limited depth | Yes |
| Quantification | Semi | Yes | Yes |
| Substrate required | Yes (X gal) | No | Yes (luciferin) |
Reporter Knockin Designs
Replacement (Knockout Reporter)
- Reporter replaces the target gene coding sequence
- Reports expression pattern of disrupted gene
- Creates null allele (gene is inactivated)
- Useful when knockout phenotype is not lethal
- Common in knockout-first allele design
Fusion Reporter
- Reporter fused to endogenous protein
- Protein function typically preserved
- Reports protein localization, not just expression
- Enables tracking of protein dynamics
- Requires careful design to avoid disrupting protein function
IRES Reporter
- Reporter expressed from bicistronic mRNA via internal ribosome entry site
- Endogenous gene expression preserved
- Reporter expressed from same transcript
- Lower reporter expression than fusion or replacement
- Does not report protein localization
2A Peptide Reporter
- Reporter separated from endogenous protein by self cleaving 2A peptide
- Near stoichiometric expression of reporter and target protein
- Both proteins released as separate polypeptides
- Higher reporter expression than IRES
- Small peptide tag remains on proteins
Applications
Gene Expression Analysis
Reporter knockins reveal endogenous gene expression patterns:
- Developmental expression timing
- Tissue and cell type distribution
- Expression level changes in disease or treatment
- Validation of transcriptomic data at cellular level
Lineage Tracing
Cre reporter systems enable permanent marking of cell lineages:
- Cross Cre knockin to Rosa26 reporter (Rosa26 tdTomato, Rosa26 Confetti)
- All descendants of Cre expressing cells permanently labeled
- Track cell fate during development or regeneration
- Identify progeny of stem or progenitor populations
Cell Sorting and Isolation
Fluorescent reporters enable isolation of specific cell populations:
- FACS isolation of reporter positive cells
- Single cell analysis of defined populations
- Primary cell culture from sorted populations
- Enrichment for transplantation studies
In Vivo Imaging
Luciferase and fluorescent reporters enable non invasive imaging:
- Track gene expression changes longitudinally
- Monitor tumor growth or metastasis
- Assess therapeutic responses in real time
- Reduce animal numbers through repeated measures
Technical Considerations
Reporter Selection Factors
Strain Background
Reporter visibility can vary with strain background:
Selected Publications
Reporter knockin models generated by ingenious targeting laboratory:
Tsvilovskyy V, Ottenheijm R, Kriebs U, Schütz A, et al. (2024).
OCaR1 endows exocytic vesicles with autoregulatory competence by preventing uncontrolled Ca2+ release, exocytosis, and pancreatic tissue damage. ↗J Clin Invest 134(7): e169428
Clausen BE et al. (1999).
Conditional gene targeting in macrophages and granulocytes using LysMcre mice. ↗Transgenic Research 8(4): 265-277
What Researchers Say
“We are very happy with this mouse line and we're not done yet. There's still plenty to do. The versatility of the F.A.S.T.™ approach is really unparalleled. There are so many possibilities to use the cassette that one paper does not showcase it all. Also, the addition of N-terminal GFP has helped quite a bit to understand gene expression patterns.”
— Frank Bosmans, PhD
Ghent University, Belgium
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