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
