Notebook > AND Gate 1 > Input > aTc Sensor
2009.8.16
Test if the tetR promoter system works well in the low-copy backbone.
Get tetR+promoter+GRP from Wu Shuke.
16:40
Digest the promoter and reporter system.
| Total | 50μL
|
| Plasmids | 5μL
|
| EcoR1 | 1μL
|
| Pst1 | 1μL
|
| Buffer | 2μL
|
| ddH2O | 11μL
|
20:00
Electrophoresis to recycle the inserts.
The order of the samples: marker, digestion products, plasmids control.
Results:
Only the first one is correctly digested, recycle them.
2009.8.17
10:00
Link the inserts with vectors with has Kanamycin resistance.
17:50
Transformation.
19:30
Start to incubate.
2009.8.18
10:00
There are many colonies on the plate.
PCR colonies to test if they are correct.
2009.8.19
9:30
Mniprep the plasmids.
The general concentration of the plasmids are about 150ng/μL.
11:00
Digest and PCR those plasmids to test if they are correct.
The digestion system:
| Total | 10μL
|
| Plasmids | 6μL
|
| EcoR1 | 1μL
|
| Pst1 | 1μL
|
| Buffer | 2μL
|
The PCR system:
| Total | 10μL
|
| Plasmids | 1μL
|
| For | 1μL
|
| Rev | 1μL
|
| Buffer | 1μL
|
| ddH2O | 6μL
|
12:00
Start to digest&PCR.
16:30
Electrophoresis to test the digestion and PCR products.
All the clones have correct colonies.
17:00
Induce the strain containing tetR and low-copy backbone by aTc.
22:00
Using flow cytometry to test the induction results.
There are about 5 folds between the induced sample and the uninduced one.
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