Contributors: Esther, Chloe, Alexander
Selection plate conditions established
Four plate conditions for the cellobiose selection assay:
| Condition | Composition |
|---|---|
| (a) | LB + chloramphenicol (34 µg/mL) |
| (b) | M9 + 1% cellobiose + chloramphenicol (34 µg/mL) |
| (c) | M9 + 1% cellobiose + 0.4% glucose + chloramphenicol (34 µg/mL) |
| (d) | M9 + 0.4% glucose + chloramphenicol (34 µg/mL) |
Stock concentrations: Chloramphenicol 34 mg/mL stock in ethanol → 34 µg/mL working. Agar powder at 30 g/L. Cellobiose 5% w/v stock → 1% working. Glucose 20% stock → 0.4% working. M9 comes at 4×, diluted to 1×.
Media prepared
- M9 stock: 380 ml 4× stock + 1170 ml distilled water, inverted to mix.
- M9 for autoclaving (solid plates): 250 ml 1× M9 + 7.5 g agar powder, autoclaved.
- Cellobiose stock: 10 g cellobiose + 200 ml filtered water, mixed 2 min, re-filtered.
- LB + chloramphenicol plates (condition a): 10× plates poured (Addgene protocol).