IGEM:IMPERIAL/2007/Projects/Reporters: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
Ben Yi Tew (talk | contribs) |
Ben Yi Tew (talk | contribs) mNo edit summary |
||
| Line 7: | Line 7: | ||
===Firefly luciferase=== | ===Firefly luciferase=== | ||
*Gene name: ''lucff'' | *Gene name: ''lucff'' | ||
*'''Description''': | *'''Description''': Oxidizes luciferin with the help of oxygen, releases light as a product of oxidation. | ||
*Enzyme: [http://www.expasy.org/enzyme/1.13.12.7 EC 1.13.12.7] | *Enzyme: [http://www.expasy.org/enzyme/1.13.12.7 EC 1.13.12.7] | ||
*Reaction: Photinus luciferin + O<sub>2</sub> + ATP <=> oxidized Photinus luciferin + CO<sub>2</sub> + AMP + diphosphate + light | *Reaction: Photinus luciferin + O<sub>2</sub> + ATP <=> oxidized Photinus luciferin + CO<sub>2</sub> + AMP + diphosphate + light | ||
*Response Time: ~30 minutes | *Response Time: ~30 minutes | ||
*Steady State Time: | *Steady State Time: 4 hours | ||
*Concentration sensitivity: Moderate | *Concentration sensitivity: Moderate | ||
*Advantages: | *Advantages: | ||
| Line 18: | Line 17: | ||
**Can be initiated by addition of luciferin | **Can be initiated by addition of luciferin | ||
*Disadvantages: | *Disadvantages: | ||
**Metabolic | **Metabolic stress, ATP is consumed | ||
**Substrate limiting | **Substrate limiting | ||
| Line 31: | Line 30: | ||
*Advantages: | *Advantages: | ||
**Fast response | **Fast response | ||
**Can be initiated by addition of substrate | |||
*Disadvantages: | *Disadvantages: | ||
**Five subunits required | **Five subunits required | ||
Revision as of 06:56, 27 July 2007
Useful Reporters
GUS reporter system
- Description: Analyze the activity of a promoter (in terms of expression of a gene under that promoter) either in a quantitative way or through visualization of its activity in different tissues. The technique is based on beta-glucuronidase, an enzyme from the bacterium Escherichia coli. This enzyme when incubated with some specific colorless or not fluorescent substrates, can transform them into coloured or fluorescent products.
- Potential use: Fluorescent output
- Link: http://en.wikipedia.org/wiki/GUS_reporter_system
Firefly luciferase
- Gene name: lucff
- Description: Oxidizes luciferin with the help of oxygen, releases light as a product of oxidation.
- Enzyme: EC 1.13.12.7
- Reaction: Photinus luciferin + O2 + ATP <=> oxidized Photinus luciferin + CO2 + AMP + diphosphate + light
- Response Time: ~30 minutes
- Steady State Time: 4 hours
- Concentration sensitivity: Moderate
- Advantages:
- Fast response
- Can be initiated by addition of luciferin
- Disadvantages:
- Metabolic stress, ATP is consumed
- Substrate limiting
Bacterial luciferase
- Gene name: luxCDABE
- Description: 5 subunit protein which oxidizes FMN with the help of oxygen to generate FMN and H2 complex. This complex breaks down to form light.
- Enzyme: EC 1.14.14.3
- Reaction: RCHO + reduced FMN + O2 <=> RCOOH + FMN + H2O + light
- Response Time: ~30 minutes
- Steady State Time: 8 hours
- Concentration sensitivity: Moderate
- Advantages:
- Fast response
- Can be initiated by addition of substrate
- Disadvantages:
- Five subunits required
- Metabolic stress, FMN is consumed
- Substrate limiting
Questions: What substrate does this need?
