Team:Hong Kong HKUST/InterLab
INTERLAB
OUR INTERLAB OBJECTIVES
The 2018 year iGEM interlab aimed at reducing Lab-to-lab variability in fluorescence measurement. Since in previous years interlab studies show that there are variation in measuring GFP expression level per cell. This large variability is the use of optical density (OD) as approximation of cell number. So this year the interlab will try in two ways: absolute cell count and colony forming units, to replace the use of OD.
Method:
All procedures are followed to the protocols iGEM given. Except the OD measurement was changed from OD600 to OD595, due to limited options of plate reader in HKUST. After contact with iGEM headquarter, we retain the data to be OD595.
Machines, materials and parts:
- Envision Multilabel Reader
Machines:
*To know more about the setting of EnVision multilabel reader, please click
- LUDOX CL-X,/b>: 45% colloidal silica suspension, use as single reference point for converting absorbance (Abs600) to OD600.
- Silica beads: Microsphere suspension that mimic the shape and size of typical E.coli cell. With known concentration, it is used as conversion of absorbance measurement to universal standard concentration of bead measurement.
- Fluorescein: fluorescent molecules for create standard fluorescence curve.
- E.coli strain DH5αCompetent cell: use for transformation, the protocol used for making it can view in here
parts:
| Parts | Parts location on the kits plate | Parts used as the promoter(strength) | Parts used as the RBS(Efficiency) | Reporter Gene | Parts used as the Terminator |
|---|---|---|---|---|---|
| Positive Control(BBa_I20270) | Plate 7 Well 2B | BBa_J23151 (nil) | BBa_B0032 (0.3) | GFP | BBa_B0010, BBa_B0012 |
| Negative Control (BBa_R0040) | Plate 7 Well 2D | BBa_R0040 (nil) | nil | ||
| Test Device 1 (BBa_J364000) | Plate 7 Well 2F | BBa_J23101 (1791au) | BBa_B0034 (1.0) | ||
| Test Device 2 (BBa_J364001) | Plate 7 Well 2H | BBa_J23106 (1185au) | |||
| Test Device 3 (BBa_J364002) | Plate 7 Well 2J | BBa_J23117 (162au) | |||
| Test Device 4 (BBa_J364007) | Plate 7 Well 2L | BBa_J23100(2547au) | BBa_B0034* (nil) | ||
| Test Device 4 (BBa_J364007) | Plate 7 Well 2L | BBa_J23100(2547au) | BBa_B0034* (nil) |
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