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Team:Heidelberg/Tyrocidine week18 biobrickmod
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Contents |
RFC10 Standardization of Tyc Modules
new Primer for B1 and a new fw Primer for TycC5
Amplification Tyc B1 and Tyc C5
29-08-2013
- Reaction I
| what | µl |
|---|---|
| Bacillus | 1 |
| AT09/11 | 2 |
| AT10/06 | 2 |
| Phusion Flash 2x Master Mix | 10 |
| DMSO | 1 |
| ddH20 | 4 |
- Conditions II
| Cycles | temperature [°C] | Time [min:s] |
|---|---|---|
| 1 | 98 | 2:00 |
| 14 | 98 | 0:01 |
| 65 ↓0.5 | 0:05 | |
| 72 | 0:55 | |
| 16 | 98 | 0:01 |
| 68 | 0:05 | |
| 72 | 0:55 | |
| 1 | 72 | 5:00 |
| 1 | 10 | inf |
Results of Gel extraction Tyc B1 and Tyc C5
| Sample | concentration ng/µl | Adsorption 260 | Adsorption 280 |
|---|---|---|---|
| H2O | 0.1 | 0.002 | 0.015 |
| B1 | 43.5 | 0.871 | 0.461 |
| C5 | 23.9 | 0.477 | 0.296 |
Restriction Digest
Tyc ADeltaCom, B1, C5, C6 (AT_01 to AT02, AT_09 to AT_10, AT_11 to AT_06, AT_07 to AT_08; xx kb; xx kb ; xx kb; xx kb) with EcoRI-HF and PstI
| what | µl |
|---|---|
| Tyc ADeltaCom, B1, C5, C6 | 20 |
| EcoRI-HF | 1 |
| PstI-HF | 1 |
| Buffer CutSmart | 5 |
| dd H2O | 24 |
Inkubate 37°C 2h
Restriction Digest of pSB1C3 (xx kb) with EcoRI-HF and PstI
| what | µl |
|---|---|
| pSB1C3 | 3 |
| EcoRI-HF | 0.5 |
| PstI-HF | 0.5 |
| Buffer CutSmart | 5 |
| dd H2O | 41 |
Inkubate 37°C 2h
Nucleotide Removal and Concentration Measurement of digested pSB1C3, Tyc ADeltaCom, B1, C5, C6
Concentration measurements showed that DNA was lost with Nucleotide Removal.
Reamplification and RFC10_Modules#Restriction_Digestas above
