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Construction of rbs-TetR-mRFP-LVA-tripart (L173)
   
Ligation L173
| Digestion n°
| Name
| Digested by
| DNA concentration
|
| D112
| rbs-TetR
| EcoRI & SpeI (insert)
| ~20 µg/mL
|
| D187
| mRFP-LVA-tripart
| EcoRI & XbaI (vector)
| 16 µg/mL
|
Reaction mixtures
|
| Control
| Sample
|
| 3:1 (insert:vector) ratio
| 4:1 (insert:vector) ratio
|
| D112 (insert)
| 0 µL
| 1,4 µL
| 1,9 µL
|
| D187 (vector)
| 2,5 µL
| 2,5 µL
| 2,5 µL
|
| T4 DNA ligase 10X buffer
| 2 µL
| 2 µL
| 2 µL
|
| T4 DNA ligase
| 1 µL
| 1 µL
| 1 µL
|
| water
| 14,5 µL
| 13,1 µL
| 12,6 µL
|
- overnight reaction at about 16°C.
Construction of rbs-FlhDC-mRFP-LVA-tripart (L172)
Ligation L172
| Digestion n°
| Name
| Digested by
| DNA concentration
|
| D186
| rbs-FlhDC
| EcoRI & SpeI (insert)
| ~24 µg/mL
|
| D187
| mRFP-LVA-tripart
| EcoRI & XbaI (vector)
| 16 µg/mL
|
Reaction mixtures (3:1 (insert:vector) ratio)
|
| Control
| Sample
|
| D186 (insert)
| 0 µL
| 1 µL
|
| D187 (vector)
| 2,5 µL
| 2,5 µL
|
| T4 DNA ligase 10X buffer
| 2 µL
| 2 µL
|
| T4 DNA ligase
| 1 µL
| 1 µL
|
| water
| 14,5 µL
| 13,5 µL
|
- overnight reaction at about 16°C.
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