Team:Weimar-Heidelberg Arts/Project/Bacteria Game/Wetlab

From 2010.igem.org

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<p><a href="https://2010.igem.org/Team:Weimar-Heidelberg_Arts/Project/Bacteria_Game">Return to the main Bacteria Game page</a>
<h2> <span class="mw-headline" id="Wetlab_Notebook"> Wetlab Notebook </span></h2>
<h2> <span class="mw-headline" id="Wetlab_Notebook"> Wetlab Notebook </span></h2>
<ul><li> usage of <a href="https://2009.igem.org/Team:Cambridge" class="external text" rel="nofollow">E. chromi principle</a> (pigmented <i>E. coli</i> cells)  
<ul><li> usage of <a href="https://2009.igem.org/Team:Cambridge" class="external text" rel="nofollow">E. chromi principle</a> (pigmented <i>E. coli</i> cells)  

Latest revision as of 17:31, 27 October 2010

Return to the main Bacteria Game page

Wetlab Notebook

  • usage of E. chromi principle (pigmented E. coli cells)
    • as a colorful crowd
      • plasmids available in iGEM Spring 2010 DNA Distribution (see table 1)
  • testing of a synthetic E.coli predator-prey system (Song et al., 2009), (Ballagaddé et al., 2008)
    • for final assault on the swarming plate and under the microscope
      • kindly provided by R. Smith (see table 2)
  • wildtype E. coli
    • for the game-kit
      • offered by A. Kern

30/09/2010

  • transformation of TOP10 cells with plasmids (see table 1) out of the registry following standard recommendations
  • over-night growth on selective LB agar plates (10 g Tryptone, 5 g Yeast extract, 10 g NaCl and 15 g Agar ad 1 L ddH2O + required antibiotics)
plasmid Part pigment color backbone registry location
table 1: used iGEM constructs
pLA01 BBa_K274110 red pSB1A2 2010 Kit Plate 3, 6J
pLA02 BBa_K274210 orange pSB1A2 2010 Kit Plate 3, 6N
pLA03 BBa_K274002 purple pSB1T3 2010 Kit Plate 3, 12B
pLA04 BBa_K274003 dark green pSB1K3 2010 Kit Plate 3, 20H
pLA05 BBa_K274004 light green pSB1K3 2010 Kit Plate 3, 20J
  • glycerol stocks (prepared directly after arrived) from predator and prey cells (see table 2) were plated on selective LB agar plates (s. a.)
sample name/function cell strain plasmid marker
table 2: used predator-prey system, gift from R. Smith
1a predator MG1655 ptetLuxRLasI-luxCcdA(SC101), placCcdBs-tetGFPuv(LVA) Cm, Kan
3a prey MG1655 pLasRLuxI-luxCcdBs, ptet-mCherry(ColE1) Cm, Kan


01/10/2010

  • inoculation of day cultures with colonies from the previous day
    • in 5 mL LB + required antibiotic
  • preparation of TB soft agar plates (10 g Tryptone, 5 g NaCl and 3 g Agar ad 1 L ddH2O + required antibiotics swarm plates) for swarming
    • inoculation by carefully pipetting 3 μL of liquid culture into the solidified agar
    • incubation of swarm plates at room temperature for 20 h
      • sufficient humidity was ensured by petri dishes filled with water
    • a photo was taken (by Canon EOS 5D Mark II) every 30 sec. for 6 h
  • inoculation of over-night cultures from the day cultures

02/10/2010

  • preparation and conduction of microscopy experiments with predator-prey system following liquid-phase protocols
    • snapshots were taken every second for one minute at beginning and at the very end of the experiments
    • time-lapse took 3 h with a picture each 20 seconds using given filters for GFPuv and mCherry

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