IGEM:University of Illinois Urbana Champaign/2009/Notebook/Bioware 2010 Arsenic Bioremediation/2010/07/09

UIUC - Arsenic Collection System!

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7/9/10 Colony Boil 50 ul H2O, 3 colonies. gave 25 ng/ul DNA. PCR Ingredient Amount Mastermix (9 reactions) 10X Buffer 2 ul 18 ul 10 mM dNTPs .4 ul 3.6 ul F primer .4 ul added individually to each reaction R primer .4 ul added individually to each reaction Template 4 ul (100ng) 36 ul Polymerase .2 ul 1.8 ul H2O 12.6 ul 113.4 ul Per Reaction: 19.2 ul mastermix, .4 ul F Primer, .4 ul R primer Primers pArsR F1 pArsR F2 pArsR R1 pArsR R2 ArsR F RFC10 EcoRI ArsR R RFC10 ArsR R fusion BSAI ArsR F Fusion BSAI LamB F1 RFC10 XbaI LamB F2 RFC10 XbaI LamB R RFC10 SpeI Samples LamBF1R LamBF2R ArsR Fusion ArsR pArsR F1R1 pArsR F1R2 pArsR F2R1 pArsR F2R2 Negative control (no primers) Program 95°C 3 min 95°C 30 sec 53°C 30 sec 72°C 90 sec go to step 2 31X 72°C 5 min hold at 4°C Meagan did transformation of yesterday's ligation at 3:55 PM, incubated for an hour at 37°C and plated 200 ul on Tet plate at 4:55 PM.