IGEM:University of Illinois Urbana Champaign/2009/Notebook/Bioware 2010 Arsenic Bioremediation/2010/07/26

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7/26/10

9:50 AM

Made glycerol stocks of 5 mL cultures

ArsR SDM1 ISB206

ArsR SDM2 ISB207

Miniprep (promega kit)

ArsR SDM1 184.9 ng/μL

ArsR SDM2 209.3 ng/μL

12:15 PM

PCR

Ingredient Amount Mastermix (4 reactions)
10X Buffer 2 μL 8 μL
10mM dNTPs .4 μL 1.6 μL
Template .9 μL (18ng) 3.6 μL
F Primer .4 μL added individually to each reaction
R Primer .4 μL added individually to each reaction
Polymerase .3 μL 1.2 μL
DMSO 1 μL 4 μL
H2O 14.6 μL 58.4 μL

Per Reaction: 19.2 μL mastermix, .4 μL F primer, .4 μL R primer.

Samples

LamBF1R

LamBF2R

ArsB

Negative control (no primers)

Primers

All primers at 10 micromolar

LamBF1

LamBF2

LamBR

ArsBF2

ArsBR2

Program

95°C 3 min
95°C 30 sec
53°C 30 sec
72°C 2 min
go to step 2 31X
72°C 5 min
hold at 4°C

3:30 PM

Gel

1% TAE, 175V

Lane Sample Amount
1 1 Kb Ladder 1 μL
2 ArsB 5μL sample + 1μL dye
3 LamBF1R 5μL sample + 1μL dye
3 LamBF2R 5μL sample + 1μL dye
4 Negative Control 5μL sample + 1μL dye

UIUC LamB.jpg