Team:Warsaw/Calendar-Main/16 October 2009

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Creation of the cro-box testing devices

Kamil


Tasks:

  • Ligation of the device with two variants of the cro-box
  • Bacteria transformation
  • I14018 brick resurrection
  • R0010 brick multiplication

Methods:

  • The ligation mix was prepared as follows (same for both variants):

    20μl purified cro-box plasmid 
    20mu;l purified reporter device
    5μl ligation buffer (Fermentas)
    1μl T4 Ligase (Fermentas)
    4μl H2O
  • A fresh bach o chemocompetent bacteria was transformed with the ligation mixtures and incubated overnight on agarose plates supplemented with ampicillin.
  • A fresh bach o chemocompetent bacteria was transformed with the I14018 brick solution and incubated overnight on agarose plates supplemented with kanamycin.
  • Bacterial culture carrying the Roo10 brick was resuspended in 50ml of liquid LB medium suplemented with ampicillin and incubated overnight.


Mutagenesis of cI

Kuba

Task: Obtain a functional, termosensitive version of cI

Results: The colonies obtained from the mutagenesis of the bistable switch created three distinc populations, demonstrating the succes of mutagenesis, as well as suggesting, that more than one point mutation was obtained (later verified by sequencing; vide: https://2009.igem.org/Team:Warsaw/results#Experimental_results)

Switch-unset.jpg



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