Protein domains/Reporters
Introduction to Reporters
What are reporters? Why call them reporters and not fluorescent proteins, beta-galactosidase/Xgal assays or pigments? The answer is that in synthetic biology, they are often used as a mechanism to display information. If you design a biosensor system that is sensitive to substrate A, you ideally want a way of displaying these results without needing an additional assay. Using a reporter protein is a great way of achieving this property.
Reporter protein coding sequences encode proteins whose presence in the cell or organism is readily observed. For example, fluorescent proteins cause a cell to fluoresce when excited with light of a particular wavelength, luciferases cause a cell to catalyze a reaction that produces light, and enzymes such as beta-galactosidase convert a substrate to a colored product. Reporters are frequently used to quantify the strength or activity of upstream gene expression parts such as promoters and ribosome binding sites. Reporters, when fused in frame to other protein coding sequences, can also be used to identify where a protein is located in a cell or organism.
There are several different ways to measure or quantify a reporter depending on the particular reporter and what kind of characterization data is desired. Generally speaking, microscopy is useful for obtaining both spatial and temporal information on reporter activity, particularly at the single cell level. Flow cytometers are best suited for measuring the distribution in reporter activity across a large population of cells. Plate readers are generally best for taking population average measurements of many different samples over time.
Some iGEM teams have excellent projects that create new reporters, or make great use of existing ones. Please see the tables below for fluorescent proteins, chromoproteins, luciferase proteins and enzymes that produce colored substrates in the Registry.
| Name | Description | AA sequence | Length |
|---|---|---|---|
| BBa_K165014 | CFP x2 | 1494 | |
| BBa_K157004 | Fluoresceine-A-binding | 522 | |
| BBa_K3484002 | Intein-mediated T3(THRB) → eGFP | 2274 | |
| BBa_K3484000 | Intein-mediated T3(THRB) → sfGFP | 2268 | |
| BBa_K103009 | linker fused to alpha fragment of TEM beta-lactamase | 578 | |
| BBa_K103013 | linker fused to omega fragment of TEM beta-lactamase | 335 | |
| BBa_J176005 | mCh | 705 | |
| BBa_K165004 | mCherry, yeast optimized for fusion | 735 | |
| BBa_K103014 | OmpA_linker_alpha_linker | 1049 | |
| BBa_K103016 | OmpA_linker_omega_linker | 809 | |
| BBa_J70595 | RFC12 Green Fluorescent Protein | 711 | |
| BBa_K157024 | SP-AntiNIP-GGGSlinker-Tm-betaLactamase(Bla1) | 1488 | |
| BBa_K157025 | SP-AntiNIP-GGGSlinker-Tm-betaLactamase(Bla2) | 1233 | |
| BBa_K157028 | SP-AntiNIP-GGGSlinker-Tm-Fluolinker-cCFP | 1263 | |
| BBa_K157026 | SP-AntiNIP-GGGSlinker-Tm-Fluolinker-cYFP | 1263 | |
| BBa_K157029 | SP-AntiNIP-GGGSlinker-Tm-nCFP | 1428 | |
| BBa_K157027 | SP-AntiNIP-GGGSlinker-Tm-nYFP | 1431 | |
| BBa_K157031 | SP-AntiNIP-GGGSlinker-Tm-Splitluciferase1/2 | 1239 | |
| BBa_K157030 | SP-AntiNIP-GGGSlinker-Tm-Splitluciferase2/2 | 1239 | |
| BBa_K157032 | SP-Lipocalin-GGGSlinker-Tm-betaLactamase(Bla1) | 1275 | |
| BBa_K157033 | SP-Lipocalin-GGGSlinker-Tm-betaLactamase(Bla2) | 1020 | |
| BBa_K157036 | SP-Lipocalin-GGGSlinker-Tm-Fluolinker-cCFP | 1050 | |
| BBa_K157034 | SP-Lipocalin-GGGSlinker-Tm-Fluolinker-cYFP | 1050 | |
| BBa_K157037 | SP-Lipocalin-GGGSlinker-Tm-nCFP | 1215 | |
| BBa_K157035 | SP-Lipocalin-GGGSlinker-Tm-nYFP | 1218 | |
| BBa_K157039 | SP-Lipocalin-GGGSlinker-Tm-Splitluciferase1/2 | 978 | |
| BBa_K157038 | SP-Lipocalin-GGGSlinker-Tm-Splitluciferase2/2 | 1026 | |
| BBa_K5466030 | Split mCerulean C-terminal. Saccharomyces cerevisiae codon optimized | 273 | |
| BBa_K5466029 | Split mCerulean N-terminal. Saccharomyces cerevisiae codon optimized | 540 | |
| BBa_K157005 | Split-Cerulean-cCFP | 243 | |
| BBa_K157006 | Split-Cerulean-nCFP | 465 | |
| BBa_K157007 | Split-Venus-cYFP | 243 | |
| BBa_K157008 | Split-Venus-nYFP | 468 | |
| BBa_K157019 | Tm-betaLactamase(Bla2) | 363 | |
| BBa_K157018 | Tm-betaLactamaseBla1 | 618 | |
| BBa_K157014 | Tm-Fluo-linker-cYFP | 393 | |
| BBa_K157016 | Tm-Fluolinker-cCFP | 393 | |
| BBa_K157017 | Tm-nCFP | 558 | |
| BBa_K157015 | Tm-nYFP | 561 | |
| BBa_K157020 | Tm-Split-Luciferase1/2 | 321 | |
| BBa_K157021 | Tm-Splitluciferase2/2 | 369 |
References
<biblio>
- Pardy pmid=7866866
</biblio>
Fluorescent proteins
<biblio>
- Chalfie pmid=8303295
- Prasher pmid=1347277
- Cormack pmid=8707053
- Tsien pmid=9759496
- Andersen pmid=9603842
- Leveau pmid=11698362
- Iafolla pmid=18350571
- Dong pmid=18352052
- Bagh pmid=18352063
- Baird pmid=11050229
- Patterson pmid=15583657
</biblio>
Luciferase
<biblio>
- Bronstein pmid=8080073
</biblio>
β-galactosidase
<biblio>
- Griffith pmid=11779182
- Villarejo pmid=4552692
- Zamenhof pmid=4552986
- Ullman pmid=1345751
</biblio> [http://openwetware.org/index.php?title=Beta-Galactosidase_Assay_%28A_better_Miller%29 Miller Assay on OpenWetWare] by Sean Moore