11/19/09
- ✓ Transfections: test Gal4/mCh/VP64 ATF's (trial 2)
Transfections
> HEK293 cells + Gal4/mCh/VP64 constructs
> Use Fugene, 24-well format
> Dilute each plasmid to 100 ng/ul in st dH2O
Wells |
Construct |
DNA |
Volumes |
Fugene |
Opti-MEM
|
1-6 |
KAH59/VNpc |
25-800 ng |
0.3, 0.5, 1.0, 2.0, 4.0, 8.0 ul |
1.8 ul |
18.2 ul
|
7-12 |
KAH60/VNpc |
" |
" |
" |
"
|
13-18 |
KAH54/VNpc |
" |
" |
" |
"
|
19-24 |
KAH66/MV5 |
" |
" |
" |
"
|
25-30 |
KAH65/MV5 |
" |
" |
" |
"
|
31-36 |
KAH59/MV5 |
" |
" |
" |
"
|
37-42 |
KAH36 |
" |
" |
" |
"
|
43-48 |
none |
" |
" |
" |
"
|
> Fugene + Opti-MEM --> R.T/ 5 min.
> Add DNA --> R.T./ 20 min.
> Add complexes to each well (1 ml med. each); 37°C
> Assay RFP/ Luc after 18-24 hours
|