Bologna University/Fluorescence Test

From 2007.igem.org

1. Growth O/N for 15h in LB (5ml) of:
-transformed E. coli with appropriate antibiotic;
-E. coli.
2. All measures must be done in M9 with OD=1.2.
3. The day after, in the morning, measure OD.
4. To obtain OD=1.2:
-centrifuge 4ml of bacterial culture at 4400rpm for 3min at 25°C;
-discard the supernatant;
-resuspend the cell pellet in 6ml of M9 medium with glucose and appropriate antibiotic (1000x [ ]).
5. Measure OD: adjust OD to 1.2 through further dilution or cell growth.
6. Adjust PMT offset with untransformed E. Coli.
7. Test culture fluorescence before IPTG induction.
8. IPTG induction:
-centrifugate bacteria culture at 4400rpm for 3min at 25°C;
-discard the supernatant;
-resuspend cell pellet in M9 medium and 2mM IPTG with appropriate antibiotic (1000x [ ]);
9. Incubate at 37°C.
10. Test fluorescence after 10min, 20min, 30min, 1h, 2h.

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