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Team:British Columbia/Protocols/ConsortiaFluor
From 2012.igem.org
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6. Measure the respective O.D 600 with a spectrometer. | 6. Measure the respective O.D 600 with a spectrometer. | ||
| - | 7. Inoculate appropriate co-culture of auxotrophs (each with | + | 7. Inoculate appropriate co-culture of auxotrophs (each with a starting O.D.600 of 0.05) in a 96 well plate containing 200uL M9 with ampicillin (100mg/mL). |
8. Incubate in Tecan Plate Scanner, taking measurements of O.D.600 and fluorescence outputs for time intervals of every 15 minutes for 24 hours. | 8. Incubate in Tecan Plate Scanner, taking measurements of O.D.600 and fluorescence outputs for time intervals of every 15 minutes for 24 hours. | ||
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EYFP - Excitation: 514nm Emission: 527nm | EYFP - Excitation: 514nm Emission: 527nm | ||
| + | |||
ECFP - Excitation: 439nm Emission: 476nm | ECFP - Excitation: 439nm Emission: 476nm | ||
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ERFP - Excitation: 584nm Emission: 607nm | ERFP - Excitation: 584nm Emission: 607nm | ||
Revision as of 23:46, 3 October 2012
Consortia Fluorescence Monitoring
1. Grow overnight 5mL cultures of auxotrophs(MetA-, TrpA-, and TyrA-)with their respective fluorescent proteins in LB media and ampicillin (100mg/mL) .
2. Spin down cells using a centrifuge (1600g, 10min). Pour out LB supernatant.
3. Wash and resuspend in 5mL M9 media. Spin down cells using a centrifuge (1600g, 10min)
4. Perform step 3 three times.
5. Resuspend in 5mL M9 Media.
6. Measure the respective O.D 600 with a spectrometer.
7. Inoculate appropriate co-culture of auxotrophs (each with a starting O.D.600 of 0.05) in a 96 well plate containing 200uL M9 with ampicillin (100mg/mL).
8. Incubate in Tecan Plate Scanner, taking measurements of O.D.600 and fluorescence outputs for time intervals of every 15 minutes for 24 hours.
Note:
EYFP - Excitation: 514nm Emission: 527nm
ECFP - Excitation: 439nm Emission: 476nm
ERFP - Excitation: 584nm Emission: 607nm
