IGEM:UNAM-Genomics Mexico/2009/Notebook/H2 Pae/2011/07/30

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1. Continuing the plasmid's stability experiment. Dilutions and plaqued from the 74 hours culture and took of 100 colonies from 60 hours dilutions.


Plasmid stability

  • At 11:20 AM I took 1 ml of the culture which was, at this point, 774 hours old. I repeat the yesterday's procedure of:
1. Vortex the 1 ml culture.
2. Take 100μl of the previous culture and put them in 900 μL of liquid medium of antibiotic. This is the first dilution.
3. Vortex 30 times the dilution.
4. Repeat the steps 2 and 3 until reach the desired dilution.

I did it until reached the 5th dilution, because the dilutions of previous hours were too many to properly obtain 100 colonies. Then I plaqued 100 μL of the dilutions 3,4, and 5.

  • I was ready to took 100 colonies form the 48 hours, but in the corresponding petri dishes didt'n even grow 10 colonies and the petri dishes also were contaminated, so we don't have records from that time.
  • After I took 100 colonies of the plaqued dilutions 4 and 5, which came from the 60 hours old culture; I plaqued them twice in only two petri dishes, one containing gentamicin and the other one without antibiotic. The previous in order to see how many colonies conserved/lost the plasmid.

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