Paris/August 1
From 2007.igem.org
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(→DGAT Assembly PCR: Megaprimer PCR) |
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* After purification: <br> | * After purification: <br> | ||
2µL of purification product (2µL of ladder): <br> | 2µL of purification product (2µL of ladder): <br> | ||
| - | - well 0: ladder | + | - well 0: ladder<br> |
| - | - well 1: negative control of purification | + | - well 1: negative control of purification<br> |
| - | - well 2: purification product | + | - well 2: purification product<br> |
[[Image: DGATDPst108012007purification.jpg|right|100px]] | [[Image: DGATDPst108012007purification.jpg|right|100px]] | ||
Revision as of 14:37, 2 August 2007
DGAT Assembly PCR: Megaprimer PCR
- Material:
- DNA plasmid (pKS::DGAT): 1.5µL (200ng)
- Tampon: 10µL
- dNTP: 1µL (final 200µmol each)
- Taq: 0.5µL
- Water: 29µL
- Primer: ForDGAT1: 4µL (final 0.8µM)
- Megaprimer: DGAT2: 4µL (50ng)
- PCR Program:
5 cycles only with DGAT2 as primer:
- 98°C: 5min
- 98°C: 1min (X5)
- 60°C: 30s (X5)
- 72°C: 2min (X5)
- 72°C: 10 min
- 4°C: inf
Add primer ForDGAT1:
30 cycles:
- 98°C: 5min
- 98°C: 1min (X5)
- 60°C: 30s (X5)
- 72°C: 2min (X5)
- 72°C: 10 min
- 4°C: inf
- Quantity: 5µg/µL
- Expected size: 1443 bp
- After purification:
2µL of purification product (2µL of ladder):
- well 0: ladder
- well 1: negative control of purification
- well 2: purification product
