Difference between revisions of "Part:BBa K1896001"
| Line 3: | Line 3: | ||
<partinfo>BBa_K1896001 short</partinfo> | <partinfo>BBa_K1896001 short</partinfo> | ||
| − | + | ||
Variant of Green Fluorescent Protein (GFP) that contains the following mutations: | Variant of Green Fluorescent Protein (GFP) that contains the following mutations: | ||
* '''F99S/M153T/V163A''': GFPuv or ''cycle 3'' GFP was optimised for excitation by UV light (360-400nm). [1] | * '''F99S/M153T/V163A''': GFPuv or ''cycle 3'' GFP was optimised for excitation by UV light (360-400nm). [1] | ||
| Line 12: | Line 12: | ||
This combination of GFP mutations has not been reported in the literature. The UGent Belgium 2016 iGEM team used this part as a [[Part:BBa_K1896010|single protein]] and as a fusion partner with [[Part:BBa_K1896015|mSA2]] and [[Part:BBa_K1896016|INP<sub>RC</sub>]] | This combination of GFP mutations has not been reported in the literature. The UGent Belgium 2016 iGEM team used this part as a [[Part:BBa_K1896010|single protein]] and as a fusion partner with [[Part:BBa_K1896015|mSA2]] and [[Part:BBa_K1896016|INP<sub>RC</sub>]] | ||
| − | + | <gallery mode="packed" heights=300px> | |
| + | File:MGFPuv2_colonies.jpeg|Colonies of <i>E. coli</i> containing BBa_K1896010 imaged under long-wave UV light. | ||
| + | File:MGFPuv2_spectrum.png|Fluorescence spectrum recorded from culture expressing mGFPuv2. | ||
| + | </gallery> | ||
| + | |||
| + | <gallery mode="packed" heights=300px> | ||
| + | File:BBa K1896001 nativePage.jpg|Native PAGE analysis confirms that mGFPuv2 is indeed a monomeric protein. Left three bands: GFPuv, rightmost three bands: mGFPuv2. | ||
| + | File:BBa K1896001 fusionProteins.jpg|MGFPuv2 can be used in difficult fusion proteins, from left to right: non-fluorescent control, INP<sub>RC</sub>-mGFPuv2 (weak promoter), mGFPuv2-streptavidin, mGFPuv2-mSA2, mGFPuv2 | ||
| + | </gallery> | ||
| + | |||
| + | |||
<span class='h3bb'>Sequence and Features</span> | <span class='h3bb'>Sequence and Features</span> | ||
Revision as of 22:00, 21 October 2016
mGFPuv2
Variant of Green Fluorescent Protein (GFP) that contains the following mutations:
- F99S/M153T/V163A: GFPuv or cycle 3 GFP was optimised for excitation by UV light (360-400nm). [1]
- S208L: Increases the fluorescence intensity of GFPuv, resulting in GFPuv2. [2]
- A206K: Monomerizes most GFP variants by disrupting a hydrophobic patch. [3]
Usage and Biology
This combination of GFP mutations has not been reported in the literature. The UGent Belgium 2016 iGEM team used this part as a single protein and as a fusion partner with mSA2 and INPRC
Colonies of E. coli containing BBa_K1896010 imaged under long-wave UV light.
Fluorescence spectrum recorded from culture expressing mGFPuv2.
Native PAGE analysis confirms that mGFPuv2 is indeed a monomeric protein. Left three bands: GFPuv, rightmost three bands: mGFPuv2.
MGFPuv2 can be used in difficult fusion proteins, from left to right: non-fluorescent control, INPRC-mGFPuv2 (weak promoter), mGFPuv2-streptavidin, mGFPuv2-mSA2, mGFPuv2
Sequence and Features
Assembly Compatibility:
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
References
- von Stetten, D., Noirclerc-Savoye, M., Goedhart, J., Gadella, T. W., & Royant, A. (2012). Structure of a fluorescent protein from Aequorea victoria bearing the obligate-monomer mutation A206K. Acta Crystallographica Section F: Structural Biology and Crystallization Communications, 68(8), 878-882.
- Ito, Y., Suzuki, M., & Husimi, Y. (1999). A novel mutant of green fluorescent protein with enhanced sensitivity for microanalysis at 488 nm excitation. Biochemical and biophysical research communications, 264(2), 556-560.
- Crameri, A., Whitehorn, E. A., Tate, E., Stemmer, W. P., Crameri, A., Kitts, P. A., & Kitts, P. A. (1996). Improved green fluorescent protein by molecular evolution using. Nat. Biotechnol, 14(3), 315-319.