CK2
- Andrew replated transformation
- Nothing at all
- Will have to retransform
- Mystery: DNA stock was a miniprep from colony grown from transformation made when I needed to sequence
- The original transformation plate worked, hence the plate growth and colony selection
- Secondary transformation did not work
- Will do 2 transformations: one from the "stock" and one from the miniprep that was used to make the "stock"
Sub 4 mini-inductions
- Both colonies yielded liquid cultures
- Inoculated 1.5ml of A into 150ml LB + 150ul Kan in 500ml flask
- Grew to OD600 = 0.6 (~1.5hrs)
- Aliquoted 25ml into 6 125ml flasks
| Volume in ul (for 125ml) |
IPTG (mM) |
Temp |
Time (hr) |
IPTG (mM) |
Temp |
Time (hr)
|
| 10 |
0.4 |
37 |
2 |
0.4 |
25 |
6
|
| 20 |
0.8 |
37 |
2 |
0.8 |
25 |
6
|
| 25 |
1.0 |
37 |
2 |
1.0 |
25 |
6
|
- Save 1ml UI sample, spin, wash, resuspend, store -80C
- Cool flasks labeled 25C for 10mins before adding respective concentrations of IPTG
- At the end of all inductions:
- Save 1ml total I sample from each, spin, wash, resuspend, store -80C
- Rest: spin, wash, resuspend, add protease inhibitor, save 5ml I sample for lysing and observing protein solubility, store all -80C
|
Project name
