Team:Newcastle/Slide Preparation


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Slide Preparation

  1. Grow overnight culture in 5 ml of LB broth, with appropriate antibiotics if required, at 37°C.
  2. Dilute the overnight culture to a ratio of 1:100 (overnight culture:LB with appropriate antibiotic ). Incubate at 37°C for 1 hour.
  3. Set up the following broths with different concentrations of IPTG in 5 ml of LB broth:
    • 2mM IPTG
    • 1mM IPTG
    • 0.2mM IPTG
    • 0.02mM IPTG
    • Broth only
  4. Innoculate the broth containing the different concentrations of IPTG with the appropriate cultures and incubate at 37°C for two hours.
  5. Prepare the slides by pippeting 500 µl of 1.2% agarose. Gently place the coverslip over the slide.
  6. Wait for about 5 minutes for the agarose to harden. Remove the coverslip by gently sliding the coverslip horizontally from the slide. This is to ensure that agarose layer remains undisturbed.
  7. Transfer 200 µl of each sample to microfuge tubes and label accordingly. Add 0.5 µl of membrane dye to each sample.
  8. Place 0.5 µl of each sample in each well of the slide and label accordingly. Place a coverslip on top of the slide.
  9. The slide is now ready for microscopy.

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