Aleksandra
Gel extraction, elution in 35µl warm water
- Ps/sRBS/LuxR/attC/term/attC
- Pm/sRBS/LuxR/attC/term/attc
- Ps/mRBS/LuxR/attC/term/attc
- Ps/wRBS/LuxR/attC/term/attc
- RBS/LuxI
PCR kit purification, elution in 35µl warm water
- wRBS/LacZα
- stRBS/lacZα
- pLux
- Pm/wRBS/LuxR
- Pm/mRBS/LuxR
Nanodrop
Sample name |
260/280 |
260/230 |
ng/µl
|
|
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|
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Ps/sRBS/LuxR/attC/term/attC |
1.86 |
0.18 |
63.2
|
Pm/sRBS/LuxR/attC/term/attc |
1.82 |
0.36 |
75.2
|
Ps/mRBS/LuxR/attC/term/attc |
1.74 |
0.43 |
27.8
|
Ps/wRBS/LuxR/attC/term/attc |
1.83 |
0.42 |
33.3
|
RBS/LuxI |
1.67 |
0.03 |
8.2
|
wRBS/LacZα |
1.83 |
1.99 |
28.3
|
stRBS/lacZα |
1.77 |
1.18 |
42.7
|
pLux |
1.76 |
1.51 |
56.1
|
Pm/wRBS/LuxR |
1.56 |
0.76 |
42.5
|
Pm/mRBS/LuxR |
1.75 |
1.76 |
35.5
|
Ligation for 45mins at 25°C
- pLux+LuxI : 1µl + 18µl + 2.2 buffer + 0.5µl ligase
- Pm/wRBS+attC/t/attC/mRFP : 0.5µl + 5µl + 0.5µl boffer + 0.5µl ligase
- Pm/mRBS+attC/t/attC/mRFP : 0.5µl + 5µl + 0.5µl boffer + 0.5µl ligase
Controls :
- pLux : 1µl + 18µl water + 2.2 buffer + 0.5µl ligase
- Pm/wRBS : 0.5µl + 5µl water + 0.5µl boffer + 0.5µl ligase
- Pm/mRBS : 0.5µl + 5µl water + 0.5µl boffer + 0.5µl ligase
Transformation of DH5α cells prepares by last year's iGEM team (P1)
With today's ligations:
- pLux+LuxI
- Pm/wRBS+attC/t/attC/mRFP
- Pm/mRBS+attC/t/attC/mRFP
Controls :
- pLux
- Pm/wRBS
- Pm/mRBS
- positive control (pUC19)
Double transformation to verify the phenotype (RFP) when the integrase is activated
- pSUlib (pBAD/IntI, Kan) and pSB1A2 (Ps/sRBS/attC/term/attC/mRFP, Amp)
- pSUlib (pBAD/IntI, Kan) and pSB1A2 (Ps/mRBS/attC/term/attC/mRFP, Amp)
- pSUlib (pBAD/IntI, Kan) and pSB1A2 (Ps/wRBS/attC/term/attC/mRFP, Amp)
- pSUlib (pBAD/IntI, Kan) and pSB1A2 (Pm/sRBS/attC/term/attC/mRFP, Amp)
Overnight cultures
- wRBS/TetA(C)
- mRBS/TetA(C)
- stRBS/TetA(C)
- wRBS/LacZα
- sRBS/LacZα
- RBS/LuxI
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