IGEM:Paris Liliane Bettencourt/Notebook/Paris 2010/2010/07/25: Difference between revisions

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*Insert your text here
<font color=red>Antoine</font>
 
<br><u>Minipreps</u>
*B0015 272.5 ng/μL
*J23110 991.9 ng/μL
 
<br><u>Restriction digest</u>
* B0015 cut by X+P: 25 μL miniprep (22/07) + 1 μL each enzyme + 0.3 μL BSA + 3 μL Buffer 3
* P1003 cut by S+P: 6 μL miniprep (22/07) + 1 μL each enzyme + 0.1 μL BSA + 1 μL Buffer 2 + 0.9 H20
* J23110 cut by S+P: 16 μL miniprep (22/07) + 1 μL each enzyme + 0.2 μL BSA + 2 μL Buffer 2
* C0040 cut by X+P: 9 μL miniprep (22/07) + 1 μL each enzyme + 0.15 μL BSA + 1.5 μL Buffer 3 + 2.35 H20
* C0012 cut by X+P: 19 μL miniprep (22/07) + 1 μL each enzyme + 0.25 μL BSA + 2.5 μL Buffer 3 + 1.25 H2O
 
Inactivation 20 min 65°C and storage at -20°C for J23110 digest product.
 
 
<br><u>Electrophoresis</u>
Gel 2%


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Revision as of 06:24, 5 August 2010

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Antoine


Minipreps

  • B0015 272.5 ng/μL
  • J23110 991.9 ng/μL


Restriction digest

  • B0015 cut by X+P: 25 μL miniprep (22/07) + 1 μL each enzyme + 0.3 μL BSA + 3 μL Buffer 3
  • P1003 cut by S+P: 6 μL miniprep (22/07) + 1 μL each enzyme + 0.1 μL BSA + 1 μL Buffer 2 + 0.9 H20
  • J23110 cut by S+P: 16 μL miniprep (22/07) + 1 μL each enzyme + 0.2 μL BSA + 2 μL Buffer 2
  • C0040 cut by X+P: 9 μL miniprep (22/07) + 1 μL each enzyme + 0.15 μL BSA + 1.5 μL Buffer 3 + 2.35 H20
  • C0012 cut by X+P: 19 μL miniprep (22/07) + 1 μL each enzyme + 0.25 μL BSA + 2.5 μL Buffer 3 + 1.25 H2O

Inactivation 20 min 65°C and storage at -20°C for J23110 digest product.



Electrophoresis Gel 2%

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Raphaël
Glycerol 15% 2 glycerols per culture, one stored at -20°C and the other at -80°C

  • TOP10 pSB1A2 weak RBS -TetA(C) - Amp
  • TOP10 pSB1A2 med.RBS -TetA(C) - Amp
  • TOP10 pSB1A2 weak RBS -LuxR - Amp
  • TOP10 pSB1A2 med.RBS -LuxR - Amp
  • TOP10 pSB1A2 stand.RBS -LuxR - Amp


Minipreps : 4 minipreps of a 10ml culture per biobrick, eluted with 30μL buffer and collected into 1 tube for a total volume of around 120μL:

  • weak RBS -TetA(C) - Amp
  • med.RBS -TetA(C) - Amp
  • weak RBS -LuxR - Amp
  • med.RBS -LuxR - Amp
  • stand.RBS -LuxR - Amp


Nanodrop to determine the amount of DNA in the miniprep samples:

Sample name 260/280 260/230 ng/μL
weak RBS -TetA(C) 1.87 2.31 186.5
med.RBS -TetA(C) 1.86 2.31 185.1
weak RBS -LuxR 1.84 2.38 159.8
med.RBS -LuxR 1.84 2.36 175.6
stand.RBS -LuxR 1.85 2.35 191.7



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