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Team:Heidelberg/Templates/Del week14 AF
From 2013.igem.org
Contents |
29-07-2013
Restriction digest of fragment FS_02 to FS_05; 11.2 kb; 27-07-2013 with BglII
File:20130729restrictiondigest 2log FS02-FS03 FS02-FS05 FS08-FS23besch.png
Test restriction digest (29.07); run at 100 V, 0.8 % gel (TAE)
Incubation at 37°C for about 3 hours
| what | µL |
|---|---|
| FS_02 to FS_05 (27-07-2013) | 15 |
| BglII | 1 |
| Buffer 3.1 | 2 |
| dd H2O | 2 |
Results:
- Test restriction digest worked out as predicted
30-07-2013
Amplification from FS_02 to FS_05; 11.2 kb
File:20130730 2log FS2-26.200nMor400nM65°Cconst FS2-26.200or400nM.68°Ctd 2xFS02-05200nM65const 2xFS02-05400nM65const.png
Amplification of Del A-G under different conditions and DelAF; run at 100 V 0.8% agarose gel (TAE)
2 reactions 200nM/400nM Primers
- Reaction
| what | µL 2nd PCR |
|---|---|
| D. acidovorans | 1 |
| FS_02 (1/10) | 5/2.5 |
| FS_05 (1/10) | 5/2.5 |
| Phusion Master Mix | 25 |
| DMSO | 2.5 |
| dd H2O | 11.5/16.5 |
- Conditions
| Biorad MyCycler* | ||
|---|---|---|
| Cycles-PCR | temperature [°C] | Time [s] |
| 1 | 98 | 10 |
| 12 | 98 | 1 |
| 68 ↓ 0.5 | 5 | |
| 72 | 3 min | |
| 18 | 98 | 1 |
| 66 | 5 | |
| 72 | 3 min | |
| 1 | 72 | 10 min |
| 1 | 4 | inf |
Results:
- Amplification did not work properly, as there were several unspecific bands. However we still had product.
