Paris/August 1
From 2007.igem.org
< Paris(Difference between revisions)
(→DGAT Assembly PCR: Megaprimer PCR) |
Nicolas C. (Talk | contribs) |
||
| (15 intermediate revisions not shown) | |||
| Line 1: | Line 1: | ||
| + | [[Paris/July 31|yesterday]] -- [[Paris/August 2|tomorrow]] <br> | ||
== DGAT Assembly PCR: Megaprimer PCR == | == DGAT Assembly PCR: Megaprimer PCR == | ||
| Line 11: | Line 12: | ||
- Megaprimer: DGAT2: 4µL (50ng)<br> | - Megaprimer: DGAT2: 4µL (50ng)<br> | ||
| - | [[Image: DGATDPst108012007.jpg|right]] | + | [[Image: DGATDPst108012007.jpg|right|100px]] |
* PCR Program:<br> | * PCR Program:<br> | ||
| Line 34: | Line 35: | ||
- 4°C: inf | - 4°C: inf | ||
| - | Quantity: | + | * '''Quantity:''' 5ng/µL <br> |
| - | Expected size: 1443 bp | + | * '''Expected size:''' 1443 bp |
| + | |||
| + | * After purification: <br> | ||
| + | [[Image: DGATDPst108012007purification.jpg|right|100px]] | ||
| + | 2µL of purification product (2µL of ladder): <br> | ||
| + | - well 0: ladder<br> | ||
| + | - well 1: negative control of purification<br> | ||
| + | - well 2: purification product<br> | ||
Latest revision as of 17:52, 7 October 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: 5ng/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
