green light receptor
NAME OF YOUR EXPERIMENT
Investigators:NAME
blue light receptor
Picking clones
Investigators: Sandra
Picking clones of transformed cells with LovTAp, NotGate, pSB1T3.
red light receptor
continuing with PCR of cph8
Investigators:Julia
2µl of the PCR reaction from 15th Aug. were loaded onto a gel.
But there was no band. Jakob is going to repeat the PCR.
3a-assembly with promotor+ribosome binding site infront of pcyA and ho1
Investigators:Julia
Digestion
- PR1 (pSB1C3)
- PR2 (pSB1C3)
- pcyA (pSB1T3)
- pcyATd (pSB1T3)
- pcyATb (pSB1T3)
- L23 (pSB1T3)
- vector (pSB1K3)
Amount of DNA and H20:
| Sample
| 500(ng)/DNA concentration (ng/μl)
| H20 μl
|
| pcyATd
| 2.8
| 35.2
|
| pcyaTb
| 2.9
| 35.1
|
| PR2
| 7.2
| 30.8
|
| PR1
| 6.1
| 31.9
|
| S22a
| 2,5
| 35.5
|
| pSB1K3
| 20
| 18
|
Enzymes necessary for digestion:
|
| PR1/PR2
| pcyA/S22a
| vector
|
| enzyme 1
| EcoRI
| XbaI
| EcoRI
|
| enzyme 2
| SpeI
| PstI
| PstI
|
-incubated for 1 hours at 37°C.
-heated for 20 minutes at 80°C
Ligation
Ligation of:
|
| PcyATd
| S22a (ho1)
|
| PR1
| 1 pTd
| 1 pTb
|
| PR2
| 2 pTd
| 2 pTb
|
PCR
Investigator: Jakob
PCR
| Name:
Jakob
| Date:
16.08.2011
|
| Continue from Experiment: 12.08.2011
Order primers
|
| Project Name:
Red light receptor, amplify Cph8
|
PCR-Mixture for one Reaction:
For a 50 µl reaction use
| 32,5µl
| H20
| Name
|
| 10µl
| 5x Phusion Buffer
| of Primer
|
| 2.5µl
| Primer fw
| P 58 (1:10)
|
| 2.5µl
| Primer dw
| P 59 (1:10)
|
| 1µl
| dNTPs
| of Template DNA
|
| 1µl
| DNA-Template
| JT122 5ng/µl
|
| 0.5 µl
| Phusion (add in the end)
|
|
The program we used:
| Temp.
| Time
| Nr. of cycles
|
| 98°
| 5´
| 1x
|
| 98°
| 30´´
| 10x
|
| 65°
| 30´´
|
| 72°
| 1´5´´
|
| 98°
| 30´´
| 20x
|
| 72°
| 1´5´´
|
| 72°
| 7´
| 1x
|
| 4°
| ∞
|
We got the information for the program and the primer sequences from the iGEM Team Uppsala Sweden.
Lysis cassette
NAME OF YOUR EXPERIMENT
Investigators:NAME
Precipitator
NAME OF YOUR EXPERIMENT
Investigators: NAME
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