06/02/13
- Transfection: KAH126 for flow cytometry on Monday
Lipo transfection
> Cells were 100% confluent
> Re-plate 1:2 dilution prior to transfection in p/s-free medium
| Wells |
Plasmid |
DNA |
Volume |
Lipo |
Opti-MEM (total)
|
| 1 |
KAH126 |
2 μg |
9.0 μL |
4 μL |
500 μL
|
| 2 |
KAH126 |
1 μg |
4.5 μL |
" |
"
|
| 3 |
KAH126 |
0.5 μg |
2.3 μL |
" |
"
|
| 4 |
KAH126 |
0.2 μg |
0.9 μL |
" |
"
|
| 5 |
KAH126 |
0.1 μg |
0.5 μL |
" |
"
|
| 6 |
none |
none |
--- |
none |
none
|
> Add Lipo to 250 μL Opti-MEM --> R.T/ 5 min.
> Add DNA to 250 μL Opti-MEM
> Add DNA mix to Lipo mix --> R.T./ 20 min.
> Remove 500 μL medium from each well; Add 500 μL complexes to each well (4 ml med. each)
> Add 1 μg/μL dox
> Grow cells at 37°C overnight
Next day
> When harvesting cells for flow cytometry, use 1/4 of cells to seed new plate (for outgrowth)
> Might run cells again to see when RFP reaches steady state
| 
Project name
