Team:Paris Bettencourt/Notebook/Interlab Study
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Notebook
Interlab Study
July
July 4th
We will participate in the iGEM interlab study: https://2014.igem.org/Tracks/Measurement/Interlab_study
The three specific devices teams are required to measure fluorescence data for are given below.
Existing device: BBa_I20260 (J23101-B0032-E0040-B0015) in the pSB3K3 vector.
- Kit location
- Plate 4, Well 18A
E.Coli NEBTurbo competent cells
To use the DNA in the Distribution Kit you may follow these instructions:
- With a pipette tip, punch a hole through the foil cover into the corresponding well of the Biobrick™-standard part that you want.
- Pipette 10uL of dH2O (distilled water) into the well. Pipette up and down a few times and let sit for 5 minutes to make sure the dried DNA is fully resuspended.
- Transform 1 or 2uL of the resuspended DNA into your desired competent cells,
Heat Shock Transformation of E. coli
This protocol can be used to transform chemically competent (i.e. from CaCl2) with a miniprepped plasmid or a ligation product.
Note: Never vortex competent cells. Mix cells by gentle shaking.
- Thaw competent cells on ice. These can be prepared using the CaCl2 protocol.
- Place 20 ul of cells in a pre-chilled Eppendorf tube.
- For an Intact Vector: Add 0.5 ul or less to the chilled cells
- Mix gently by flicking the tube.
- Chill on ice for 10 minutes. This step is optional, but can improve yields when transforming a ligation product.
- Heat shock at 42 °C for 30 seconds.
- Return to ice for 2 minutes.
- Add 200 ul LB medium and recover the cells by shaking at 37 °C.Kanamycin: 120 minutes
- Plate out the cells on selective LB.Note: 200 ul is the maximum volume of liquid that an LB plate can absorb.
- Incubate at 37 °C. Transformants should appear within 12 hrs.
Result: Bacteria grow and made big ans small colonies
4.Pick a single colony and inoculate broth (again, with the correct antibiotic) and grow for 16 hours.
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