IGEM:Peking University/2008/Experiment/PKU 0807018 G1 1
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PKU_080718_G1_1
- Enzyme cutting-pADH-lac2(PCR product)
 
- System for double ligation
 
 
 
 
 
 
 
 
 
| Volume (μL) | |
| Promoter | 20 | 
| BamHI | 1 | 
| SacI | 1 | 
| 0.5×K Buffer | 2.5 | 
| ddH2O | 25.5 | 
- System for single ligation
 
 
 
 
 
 
 
 
 
| Volume (μL) | |
| Promoter | 20 | 
| XhoI | 2 | 
| SacI | 2 | 
| 1×M Buffer | 5 | 
| ddH2O | 21 | 
- 'PCR- pGREG504-pADH-(lac1/lacS/tet)-50ul
- Using program lac12MQT
 

- Gel running and extraction-promoter(PCR product)
 
- Enzyme cutting-AID
 

- lanes: intact AID(control)|λmarker|Promoter(single)|promoter(double)|AID|AID
 
 
 
 
 
 
- Enzyme cutting-pADH-lac1/lacS/tet
 
- System for double ligation
 
 
 
 
 
 
 
 
 
| Volume (μL) | ||
| Promoter | 20 | |
| BamHI | 1 | ×3 | 
| SacI | 1 | |
| 0.5×K Buffer | 2.5 | |
| ddH2O | 25.5 | 
- System for single ligation
 
 
 
 
 
 
 
 
 
| Volume (μL) | ||
| Promoter | 20 | |
| XhoI | 2 | ×3 | 
| SacI | 2 | |
| 1×M Buffer | 5 | |
| ddH2O | 21 | 
- Single Ligation-pGREG504-pAID-lac2
 
| Volume (μL) | |
| lac2 | 4 | 
| pGREG504 | 1 | 
| ligation buffer | 5 | 
- Double Ligation-pGREG504-pAID-lac2-AID
 
| Volume (μL) | |
| lac2 | 3 | 
| AID | 3 | 
| pGREG504 | 1 | 
| ligation buffer | 7 | 
- Control C1
 
| Volume (μL) | |
| lac2 | 4 | 
| ddH2O | 4 | 
| ligation buffer | 5 | 
- Control C2
 
| Volume (μL) | |
| pGREG504 | 1 | 
| ddH2O | 4 | 
| ligation buffer | 7 | 
- Control C3
 
| Volume (μL) | |
| lac2 | 3 | 
| ddH2O | 4 | 
| ligation buffer | 7 | 
- Control C4
 
| Volume (μL) | |
| AID | 3 | 
| ddH2O | 4 | 
| ligation buffer | 7 | 
- Control C5
 
| Volume (μL) | |
| AID | 3 | 
| lac2 | 3 | 
| ddH2O | 1 | 
| ligation buffer | 7 | 
- Gel running-testing-pADH-lac1/S/tet
- Gel running-testing-digestion-lac2/AID
 
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