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Team:University of Lethbridge/Notebook/GeneralLabJuly
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-pLACI (BBa_R0011) | -pLACI (BBa_R0011) | ||
-pSTRONG (BBa_J23119) | -pSTRONG (BBa_J23119) | ||
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| + | loading dye [http://openwetware.org/wiki/Agarose_gel_loading_dye sizes] | ||
Revision as of 01:45, 15 July 2008
Contents |
July 1, 2008
Nathan Phillips, Andrew
Made 500 mL of LB agar + amp and 500 mL of Liquid LB
July 2, 2008
Nathan Puhl, Alix, Sebastian, Munima, Roxanne, Christa
Transformed BBa_J24679 (RBS + LacI), BBa_P0440 (RBS+TetR+T10+T12), leftover DNA from Double T (June 26, 2008), and 1 uL of pSB1A7 plasmid (June 18, 2008).
Protocol changes:
-3 uL of DNA -spin down 200 uL of cells and resuspend in 100 uL of LB; plate on LB + amp
July 3, 2008
Nathan Puhl, Munima, Christa, Alix, Roxanne, Sebastian
Checked transformation plates. Only the positive control (pSB1A7) had colonies (~1500) indicating that there is nothing wrong with the transformation protocol or cells so we must be having problems with the DNA extraction from the filter paper. Next week we will attempt various changes to the protocol to extract more DNA.
July 8, 2008
Munima, Christa, Nathan Puhl
Made 500mL of LB semi-solid media and poured 24 plates. Stored in the iGEM 4 C fridge
Nathan Puhl, Alix
Flourescent Reporter
Transformation from BioBricks LacI (BBA_J24679), TetR (BBa_P0440) and DT (BBa_B0015).
Protocol:
-punched out 2 spots of filter paper -15uL TE, for 30 min. @ 50 C -centrifuge 3 min. @ 15000 g -freeze 5 min. in -20 C freezer -heat shock 1 min. in 42 C water bath -centrifuge 3 min. -2 uL of plasmid into 25 uL DH5alpha -leave on ice for 30 min. (Left remaining DT, TetR, LacI to sit overnight at room temp. to possibly test for better DNA recovery) -put in 42 C water bath for 45 sec. -chill on ice for 2 min. -add 1 mL of SOC broth -incubate cells @ 37 C, 225 rpm for 60 min. -spin down 400 uL cells (1 min. 16000xG), remove 300 uL -resuspend -plate, incubate at 37 C overnight
Subcultured 3 biobricks (for flourescent reporter) glycerol stocked from last year into 5 mL liquid LB + Amp.
-RFP Sub. (BBa_I13507) -pLACI (BBa_R0011) -pSTRONG (BBa_J23119)
loading dye sizes
