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Team:DTU-Denmark/Notebook/25 August 2013
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Latest revision as of 12:05, 4 October 2013
25 August 2013
Contents |
Lab 208
Main purpose
- colony PCR
Who was in the lab
Kristian, Henrike
Procedure
Colony PCR
Performed colony PCR to confirm insert for HAO, AMO, cycAX and Nir transformants. Used Q5 premix with the following reaction mix:
| compound | amount |
|---|---|
| Q5 mix | 25 uL |
| FW primer | 3 uL |
| RV primer | 3 uL |
| template | 1 uL |
| MilliQ | 18 uL |
Template was made by resuspending 1 culture in 100uL MilliQ.
program:
| temperature | time | cycles |
|---|---|---|
| 98C | 10:00 | - |
| 98C | 0:10 | 36 |
| annealing temperature | 0:30 | 36 |
| 72C | 0:20 | 36 |
| 72C | 5:00 | - |
| 10C | hold | - |
details (primers, temp, expected fragment length):
- cycAX - FW_1, RV_2, 61C, 749bp
- HAO - FW_2, RV_3, 64C, 751bp
- AMO - FW_2, RV_3, 63C, 750bp
- Nir - FW_2, RV_3, 64C, 733bp
- Nir - FW_5, RV_6, 71C, 737bp
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