"
Team:Freiburg/Notebook/9 September
From 2011.igem.org
SophieCramer (Talk | contribs) (→blue light receptor) |
SophieCramer (Talk | contribs) (→blue light receptor) |
||
| Line 19: | Line 19: | ||
==<span style="color:blue;">blue light receptor</span>== | ==<span style="color:blue;">blue light receptor</span>== | ||
| - | + | ===PCR=== | |
| Line 95: | Line 95: | ||
stored in blue light box | stored in blue light box | ||
| - | + | ||
| + | |||
| + | ===Digestion=== | ||
Revision as of 12:20, 10 September 2011
Contact 
Contents |
Commons
Testdigests
Investigators: Sophie
Testdigest of our various minipreps
green light receptor
NAME OF YOUR EXPERIMENT
Investigators:NAME
blue light receptor
PCR
| Name: Sophie
| Date: 9.9.11 |
| Continue from Experiment: Gibson (Date): 9.9.11
(Name): Sophie, Sandra | |
| Project Name: | |
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 | P97 |
| 2.5µl | Primer dw | P100 |
| 1µl | dNTPs | of Template DNA |
| 1µl | DNA-Template | G-♥-NOT and G-♥-NOT 50 |
| 0.5 µl | Phusion (add in the end) |
What program do you use?
First 25 cycles: touchdown 69°C -0.4°C
next 5 cycles: touchdown 72°C -0.2°C
To confirm the PCR-Product has the correct size, load 2 µl of the sample onto an agarose-gel.
How did you label the PCR-Product, where is it stored and what do you do next?
Labeled: G-♥-NOT inserts a,b
stored in blue light box
Digestion
| Name: Sophie | Date: 9.9.11 |
| Continue from Date 9.9.11 Name: Sophie
Experiment: PCR | |
| Project Name: ♥-NOT in PR-vector | |
Procedure
- add H2O (38μl-DNA )
- 5 μl NEB4 buffer (stored at iGEM’s, -20°C)
- 5 μl 10x BSA (used 1:10 diluted sample stored at iGEM’s, -20°C)
- DNA (500 ng) (Vector:Insert ratio 1:3 in following Ligation)
- 1 μl restriction enzymes (stored at iGEM’s, -20°C)
- heat for 1-2 hours 37°C (6 hours if time)
- heat for 20 minutes 80°C (inactivation of enzymes)
- keep at 4°C if you cannot continue
Measured DNA-concentration with Nanodrop to calculate the volume of DNA to do the digestion:
| Sample Name | DNA concentration (μg/μl) |
| G-♥-NOT a,b | ~140 |
Restriction enzymes you need to cut the vector, insert1 and insert 2:
| Components | | Insert(μl) | |
| DNA (500ng) | |||
| BSA (10x) (5μl) | |||
| NEB4 Buffer (5μl) | |||
| Enzyme 1 (1μl) | Spe I | Nhe I | |
| Enzyme 2 (1μl) | Pst I | Pst I | |
| H2O (38 μl- DNA) | |||
| In total 50 μl | |||
Documentation:
Why are you doing this experiment? Where are the samples stored? Antibiotica resistance, vector used etc.
| The LOV-Repressor protein needs to be expressible
Samples stored in blue light box Vector used: all PR-vectors (D39- D44) |
red light receptor
NAME OF YOUR EXPERIMENT
Investigators:NAME
Lysis cassette
NAME OF YOUR EXPERIMENT
Investigators:NAME
Precipitator
NAME OF YOUR EXPERIMENT
Investigators: NAME
