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

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(Difference between revisions)
 
Line 47: Line 47:
There are many colonies on the plate.<br>
There are many colonies on the plate.<br>
PCR colonies to test if they are correct.
PCR colonies to test if they are correct.
 +
 +
==='''2009.8.19'''===
 +
'''9:30'''<br>
 +
Mniprep the plasmids.<br>
 +
The general concentration of the plasmids are about 150ng/μL.
 +
'''11:00'''<br>
 +
Digest and PCR those plasmids to test if they are correct.
 +
The digestion system:<br>
 +
{|cellpadding=5
 +
|Total||10μL
 +
|-
 +
|Plasmids||6μL
 +
|-
 +
|EcoR1||1μL
 +
|-
 +
|Pst1||1μL
 +
|-
 +
|Buffer||2μL
 +
|}
 +
 +
The PCR system:<br>
 +
{|cellpadding=5
 +
|Total||10μL
 +
|-
 +
|Plasmids||1μL
 +
|-
 +
|For||1μL
 +
|-
 +
|Rev||1μL
 +
|-
 +
|Buffer||1μL
 +
|-
 +
|ddH2O||6μL
 +
|}
 +
'''12:00'''<br>
 +
Start to digest&PCR.
 +
'''16:30''' <br>
 +
Electrophoresis to test the digestion and PCR products.<br>
 +
All the clones have correct colonies.<br>
 +
 +
'''17:00'''<br>
 +
Induce the strain containing tetR and low-copy backbone by aTc.
 +
'''22:00'''<br>
 +
Using flow cytometry to test the induction results.<br>
 +
There are about 5 folds between the induced sample and the uninduced one.<br>
 +
 +
 +
 +
{{PKU_Beijing/Foot}}
{{PKU_Beijing/Foot}}
__NOTOC__
__NOTOC__

Latest revision as of 03:44, 22 October 2009
 
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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