03/02/15 - Alexander Ellingson
PCR-verify activation domain primers
Try different cDNA libraries for the CARM1, P300, and MYB fragment to account for any mutations or deletions
- U2OS C002, 1:1000 cDNA dilution; CARM1 = 1827 bp: CARM1 F1/ CARM1 R1; P300 = 1848 bp: P300 F1/ P300 R1; MYB = 159 bp: MYB F1/ MYB R1.
- SKNSH C001, 1:1000 cDNA dilution; same.
- K562 C001, 1:1000 cDNA dilution; same.
| Reagent
|
Rxn1
|
Rxn2
|
Rxn3
|
Expected:
1. U2OS CARM1 = 1827 bp
1. U2OS P300 = 1848 bp
1. U2OS MYB = 159 bp
2. SKNSH CARM1 = 1827 bp
2. SKNSH P300 = 1848 bp
2. SKNSH MYB = 159 bp
3. K562 CARM1 = 1827 bp
3. K562 P300 = 1848 bp
3. K562 MYB = 159 bp
|
Hover name
10 μL/lane, 1% agarose; Ladder
|
| Template |
1.0 |
1.0 |
1.0
|
| 10 uM fwd primer |
1.0 |
1.0 |
1.0
|
| 10 uM rev primer |
1.0 |
1.0 |
1.0
|
| 2x GoTaq green |
12.5 |
12.5 |
12.5
|
| dH2O |
9.5 |
9.5 |
9.5
|
| |
25.0 |
25.0 |
25.0
|
Program: GOTAQ35cyc
- 95°C, 3 min
- 35x[95°C, 1 min; 57°C, 1 min; 72°C, 3 min]
- 72°C, 3 min
- 4°C ∞
|
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