| Activation: |
|
|
|
|
|
| Test |
pMW112 |
pBait |
(White/light blue) |
(White/light blue) |
(No) |
| Negative control |
pMW112 |
pEG202-Ras |
White |
White |
No |
| Weak positive control |
pMW112 |
pEG202-hsRPB7 |
Light blue |
Bluish |
No |
| Strong positive control |
pMW112 |
pSH17-4 |
Dark blue |
Blue |
Yes |
|
| Repression: |
|
|
|
|
|
| Test |
pJK101 |
pBait |
(White) |
(Lighter blue) |
|
| Negative control |
pJK101 |
pGKS3a |
White |
Blue |
No |
| Positive control |
pJK101 |
pEG202-Ras |
White |
Lighter blue |
|
|
|
Test by Western blot |
|
| Expression: |
|
|
|
|
| Test |
Use clones from activation assay |
pBait |
(Single band) |
|
| Positive control 1 |
pEG202-Ras |
Single band |
~45 kD |
| Positive control 2 |
pSH17-4 |
Single band |
~35 kD |
|
|
|
|
|
| apGKS3 can be substituted for any other yeast HIS3 plasmid not expressing LexA.
|
| (Adapted from Table 18-7, Sambrook and Russell 2001.) |