PCR/Agarose Gel Electrophoresis
- PCR: Candidate cell lines (3 primers + 1 error = 4)
| Reagent
|
Vol
|
Mix* (x #)
|
Expected:
1. 4-x, P1 = 1145
2. 4-x, P3 = 1046
3. 4-x, P6 = 1139
4. 6-x, P1 = 1146
5. 6-x, P3 = 1046
6. 6-x, P6 = 1146
|
5 μL/lane; 1% agarose; Ladder
|
| genomic OR plasmid DNA |
1.0 / 0.5 |
4.0 / 2.0
|
| 10 μM forward primer |
1.0 |
---
|
| 10 μM reverse primer |
1.0 |
---
|
| 2x GoTaq green |
10.0 |
40.0
|
| dH2O |
7.0 / 7.5 |
28.0 / 30
|
| |
20.0 μL |
72.0 μL
|
Note: *Prep one Mix per template
- For each template, aliquot 18.0 master mix into 3 tubes
Thermal cycler: Labnet - GOTAQ
- 95°C 3 min.
- 30x[95°C, 30 sec; 57°C 30 sec; 72°C 30 sec]
- 72°C 3 min.
- 4°C, ∞
- Conclusions
- Top image, top row, 6-1, 6-2, 6-3, P1, 3, and 6 for each.
- Top image, bottom row, 6-4, 6-5, 6-6, P1, 3, and 6 for each.
- Bottom image, top row, 4-1, 4-2, 4-3, P1, 3, and 6 for each.
- Bottom image, bottom row, 4-4, 4-7, 4-9, P1, 3, and 6 for each.
|
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