Team:PKU Beijing/Notebook/AND Gate 1/Input/TetR 2

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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.

Total50μL
Plasmids5μL
EcoR11μL
Pst11μL
Buffer2μL
ddH2O11μL

20:00
Electrophoresis to recycle the inserts.
The order of the samples: marker, digestion products, plasmids control.
Results:
PKU 20090816 Shan Shen 1.JPG
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:

Total10μL
Plasmids6μL
EcoR11μL
Pst11μL
Buffer2μL

The PCR system:

Total10μL
Plasmids1μL
For1μL
Rev1μL
Buffer1μL
ddH2O6μL

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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