IGEM:IMPERIAL/2006/LabCalendar/2006-7-10

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Contents

John Sy

Culturing Bacteria

  • Combine together:
    • 10 mL growth medium
    • 200 uL DH5a E. coli gram negative bacteria
  • Allow to grow for a few hours until the optical density of the bacterial solution is between 0.8 and 1.0 as measured by the spectrophotometer
  • Cool the culture for a few minutes and place in an ice bath
  • Centrifuge the bacteria to obtain a bacterial pellet
  • Remove media and place on ice
  • Add 1 mL of 100 mM calcium chloride solution
  • Vortex and place back on ice bath
  • Transfer 1 mL into a microcentrifuge tube and place back on ice
  • Remove supernatant and resuspend in 100 uL of calcium chloride

Innoculation

  • Innoculate a colony of bacteria into 2 mL of ampicillin resistant growth medium

Transfering the part from the registry into the bacteria

  • We use part I13273 from well 4H (YFP Producer controleld by 3OC6HSL Receiver Device (LuxR based receiver controls production of YFP)
  • Pipette 15 uL of ultrapure water into the well and dilute
  • Remove and transfer into a microvial
  • Remove 5 uL of DNA solution and place into solution containing the bacteria
  • Leave solution for about 40 minutes
  • Heat shock the soution at 42C for 90 sec and place back on ice bath
  • Transfer the bacteria with the part onto an agar plate and spread out using a glass spreader
  • Incubate at 37C overnight
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