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Team:British Columbia/Protocols/ConsortiaFluor
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| + | <h1>Consortia Fluorescence Monitoring</h1> | ||
| + | |||
| + | #Grow overnight 5mL cultures of auxotrophs(MetA<sup>-</sup>, TrpA<sup>-</sup>, and TyrA<sup>-</sup>)with their respective fluorescent proteins in LB media and ampicillin (100 mg/mL) . | ||
| + | #Spin down cells using a centrifuge (1600 g, 10 min). Pour out LB supernatant. | ||
| + | #Wash and resuspend in 5mL M9 media. Spin down cells using a centrifuge (1600 g, 10 min) | ||
| + | #Perform step 3 three times. | ||
| + | #Resuspend in 5 mL M9 Media. | ||
| + | #Measure the respective O.D 600 with a spectrometer. | ||
| + | #Inoculate appropriate co-culture of auxotrophs (each with a starting O.D.600 of 0.05) in a 96 well plate containing 200 μL M9 with ampicillin (100 mg/mL). | ||
| + | #Incubate in Tecan plate scanner, taking measurements of O.D.600 and fluorescence outputs for time intervals of every 15 minutes for 24 hours. | ||
| + | |||
| + | <h1>Excitation and Emission Wavelengths of Fluorescent Proteins</h1> | ||
| + | |||
| + | *EYFP - Excitation: 514nm Emission: 527nm | ||
| + | |||
| + | *ECFP - Excitation: 439nm Emission: 476nm | ||
| + | |||
| + | *ERFP - Excitation: 584nm Emission: 607nm | ||
Latest revision as of 03:42, 4 October 2012
Consortia Fluorescence Monitoring
- Grow overnight 5mL cultures of auxotrophs(MetA-, TrpA-, and TyrA-)with their respective fluorescent proteins in LB media and ampicillin (100 mg/mL) .
- Spin down cells using a centrifuge (1600 g, 10 min). Pour out LB supernatant.
- Wash and resuspend in 5mL M9 media. Spin down cells using a centrifuge (1600 g, 10 min)
- Perform step 3 three times.
- Resuspend in 5 mL M9 Media.
- Measure the respective O.D 600 with a spectrometer.
- Inoculate appropriate co-culture of auxotrophs (each with a starting O.D.600 of 0.05) in a 96 well plate containing 200 μL M9 with ampicillin (100 mg/mL).
- Incubate in Tecan plate scanner, taking measurements of O.D.600 and fluorescence outputs for time intervals of every 15 minutes for 24 hours.
Excitation and Emission Wavelengths of Fluorescent Proteins
- EYFP - Excitation: 514nm Emission: 527nm
- ECFP - Excitation: 439nm Emission: 476nm
- ERFP - Excitation: 584nm Emission: 607nm
