IGEM:University of Illinois Urbana Champaign/2009/Notebook/Bioware 2010 Arsenic Bioremediation/2010/06/28

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6/28/10

11:00 am:

PCR of backbones:

10x Buffer 2 ul

dNTPs .5 ul

Template 1ul

Polymerase .2 ul

Primer F 1 ul

Primer R 1ul

diH2O 14.3 ul

Total: 20 ul

Template: pSB1A2, pSB1C3, pSB1T3, pSB1AC3-2, pSB1AK3 Psb1AT3.

Negative Controls:

1. no template, 1 ul water instead

2. For each sample, no polymerase, .2 water instead.

Program: iGEM 2Kb

11:35 am

Diluted primers VF2 and VR to 20 um,

PCR:

used 100ng template for each reaction

Program: iGEM 2Kb

BBa_K190015 BBa_K190033 BBa_K257108 Negative control
10X buffer 2.5 ul 2.5 ul 2.5 ul 2.5 ul
10mM dNTPs .5 ul .5 ul .5 ul .5 ul
Template .5 ul .25 ul 1 ul 5 ul
F Primer VF2 .625 ul .625 ul .625 ul .625 ul

1:15 PM

digestion for Steve

RFP <html>

XbaI 1 ul
PstI 1 ul
Buffer 2 5 ul
BSA .5 ul
H2O 41.5 ul
DNA 1 ul

</html>

Lock1

EcoRI 1 ul
SpeI 1 ul
Buffer 2 5 ul
BSA .5 ul
H2O 41.5 ul
DNA 1 ul

key1

XbaI 1 ul
PstI 1 ul
Buffer 2 5 ul
BSA .5 ul
H2O 41.3 ul
DNA 1.2 ul

pBAD

EcoRI 1 ul
SpeI 1 ul
Buffer 2 1 ul
BSA .5 ul
H2O 41 ul
DNA 1.5 ul

2:30 PM gel GRAB PICTURE FROM COMPUTER

lane sample amount
1 1 Kb Ladder 2 ul
2 BBa_K190015 10 ul + 2ul dye
3 BBa_K190033 10 ul + 2ul dye
4 BBa_K257108 10 ul + 2ul dye
5 Negative control 10 ul + 2ul dye
6 pBAD 10 ul + 2ul dye
7 Key 10 ul + 2ul dye
8 Lock 10 ul + 2ul dye
9 RFP 10 ul + 2ul dye
10 1 Kb Ladder 3 ul

3:30 PM GRAB PICTURE FROM COMPUTER

Gel of Backbones

Top half of gel

lane sample amount
1 1 Kb Ladder 3 ul
2 pSB1A2 5 ul +1 ul dye
3 PSB1A2 (-) 5 ul +1 ul dye
4 pSB1C3 5 ul +1 ul dye
5 pSB1C3 (-) 5 ul +1 ul dye
6 pSB1T3 5 ul +1 ul dye
7 pSB1T3 (-) 5 ul +1 ul dye
8 pSBAC3-2 5 ul +1 ul dye
9 pSBAC3-2 (-) 5 ul +1 ul dye
10 1 Kb Ladder 3 ul

Bottom half of gel

lane sample amount
1 1 Kb Ladder 3 ul
2 pSBAK3 5 ul +1 ul dye
3 pSBAK3(-) 5 ul +1 ul dye
4 pSBAT3 5 ul +1 ul dye
5 pSBAT3(-) 5 ul +1 ul dye
6 negative 5 ul +1 ul dye
7 1 kB ladder 3 ul