IGEM:British Columbia/2009/Notebook/Biosensor Sensitivity/2009/06/11

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Miniprep of RBS-RFP-Terminator, Assembly 1 (J23100 + GFP), pBAD reverse and Amp/Tet Construction Plasmid

  • Day cultures of 5.6mL media and cells innoculated from plates were started at 10:50
  • RBS-RFP-Terminator, Assembly 1 and pBAD reverse were all grown using LB-Amp
  • Amp/Tet Construction Plasmid was grown on LB-Tet
  • Three cultures of Assembly 1 were started because the colonies on the plates did not have a noticible green flouresence
  • Miniprepped using a Invitrogen ChargeSwitch Pro Filter Plasmid Miniprep kit after 7 hours culture
  • Did not observe many cells after being spun down in a Galaxy 16 centrifuge
  • Used 50uL of Elution Buffer instead of 100uL as wanted as high of a concentration as possible as the number of cells is less then usual (was not inncubated as long as usual)
  • Eric started overnight cultures of all 4 plasmids in case we need to do another miniprep tommorow

==Preparation of Miniprep ON Cultures)

  • Innoculated the following cultures of E. coli (done with Charles)
Plasmid/Part Strain of E. coli Antibiotic Description
pSB1At3 DB3.1 Tetracycline Tet construction plasmid
DB3.1 Ampicillin P_BAD Reverse
DB3.1 Ampicillin RBS-RFP-Terminator
DB3.1 Ampicillin Assembly 1
  • Took a colony and innoculated fresh 6 mL LB-antibiotic
  • Incubated at AMBL 37ºC incubator
  • Tomorrow: miniprep the cultures and make glycerol stocks of these.

Ampicillin - New stocks testing

  1. Made a new stock of Amp-100
    1. 20 mL sH2O + 2.0755 g ampicillin sodium salt
    2. Filter sterilized
  2. Stored in 2x 15 mL culture tubes (1000X stock)
  3. Tested by spread plating 20µL of stock