18 December 2008

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Date: 18 December 2008; Time: 0930; Agenda: Quantification of 24hr biofilm at 3 temperatures.

  • 5 ml from 10-ml overnight LB culture was put into 150 ml LB broth and incubated for 5hr at 37 C

Biofilm formation:

  • The procedure for biofilm growth is the same as that performed before, only that now instead of directly taking out the LB culture and then centrifuging this sample, we take out the LB culture and measure and transform it into the McFarland standard first BEFORE centrifuging. This makes sure that the M9 medium has the same starting cell number as the LB McFarland standard.
  • For LB McFarland standard:
    • Approximately 9X (0.3ml broth culture to 2.7ml LB broth) dilution of LB Broth required.
  • This LB McFarland standard was plated in order to verify that the McFarland standard was indeed obtained (to be counted the next day):
    • 10^-10 to 10^-20 dilutions plated.
  • For the M9 culture (to be counted the next day):
    • 10^-6 to 10-10 dilutions plated.
  • After both M9 and LB cultures are diluted 1:29 with the respective media, they are placed in the CBDs:
    • Columns 1-6: M9 Culture; Columns 7-12: LB Culture
  • The lids of the respective CBDs are then replaced and parafilmed.
  • The 3 CBDs are placed in the 3 incubators of 21deg, 30deg and 37 deg for 24hrs at 30rpm.