IGEM:PKU Beijing/2009/Notebook/AND Gate 1/Core/2009/07/21/Shan Shen

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00:40
Digest SupD and terminator plasmids.
System of SupD:

Total 20μL
Plasmids 6μL
EcoR1 1μL
Spe1 1μL
Buffer 2μL
ddH2O 10μL

System of terminator 1-23L:

Total 20μL
Plasmids 6μL
EcoR1 1μL
Xba1 1μL
Buffer 2μL
ddH2O 10μL

1:00
Start to digest.

10:00
Electrophoresis to test the digestion products.
The order and the amount of the samples: marker 5μL, SupD digestion, 1-23L plasmids, 1-23L digestion.
PKU 20090721 Shan Shen 3.JPG
Results:
The digestions are successful.

11:10
Recycle the fragments of SupD.

13:10
Ligation of the vectors of 1-23L and inserts of SupD.
System of ligation: 10μL

Control system 1:7 system
Vectors 1μL 1μL
Inserts 0μL 7μL
Buffer 1μL 1μL
Ligase 1μL 1μL
ddH2O 7μL 0μL

13:10
Ligation starts.

14:21
Miniprep bi-stable 1,2,3,SupD1 and 1-9H.
Concentrations:

Number of the plasmids Concentration(ng/μL)
Bi-stable 1 160
Bi-stable 2 200
Bi-stable 3 250
SupD1 250
1-9H 110

17:20
Transform the ligation products of SupD+1-23L backbone.