Team:Goettingen/week11-3

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<h2><b>V07_13 </b></h2><br>
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<b>V07_13_1 1<sup>st</sup> round: Miniprep of clones 1-12 V07_12 </b><br>
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<li>Experiment: <br>Plasmids from the 1<sup>st</sup> mutation round were prepped using the peqGOLD Plasmid Miniprep Kit (PeqLab) following the user manual. DNA concentrations were determined for subsequent sequencing.</li>
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<li>Observations and results: <br>Both the liquid culture and the dilution plates were positive for bacterial growth. Plates showed the following results:<br>
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10<sup>4</sup> 10 clones<br>
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10<sup>5</sup> 2 clones<br>
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10<sup>6</sup> 0 clones<br>
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<b>V07_13_2 1<sup>st</sup> round: Miniprep of liquid culture V07_11</b><br>
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<li>Experiment: <br>We used 4 x 5 mL liquid culture split to 4 columns for plasmid preparation with the peqGOLD Plasmid Miniprep Kit (PeqLab).
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<a href="https://2012.igem.org/Team:Goettingen/Notebook">Back to overview</a><br>
<a href="https://2012.igem.org/Team:Goettingen/Notebook">Back to overview</a><br>
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Revision as of 17:35, 22 September 2012

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#3 Chemoreceptor Library - 11th Week

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V07_10


V07_10_1 1st round: Digestion DpnI/BsaI and clean-up
  • Experiment:
    The digestion and subsequent clean-up were carried out according to the protocol of week 10.
  • Observations and results:
    The corresponding gel showed a band of the expected size.

V07_10_2 1st round: Ligation
  • Experiment:
    The ligation was carried out according to the protocol of week 10 and incubated over night at 16 °C.


V07_11


V07_11_1 1st round: Ethanol precipitation of ligated mutated plasmids
  • Experiment:
    The ethanol precipitation was carried out according to protocol, this time with the right volumes!
  • Observations and results:
    The corresponding gel showed a band of the expected size.

V07_11_2 1st round: Transforamtion of electrocompetent cells wir the mutant plasmid mixture
  • Experiment:
    Electrocompetent cells were prepared according to protocol. We transformed 200 µL of cells with 10 µL of DNA according to the protocol of week 10. Dilution plates and liquid culture was incubated over night at 37 °C, 180 rpm.


V07_12


V07_12_1 1st round: Analysis of transformation V07_11
  • Experiment:
    Both the liquid culture and the dilution plates were checked for bacterial growth. 5 mL LB liquid cultures + CM were prepared for each clone on the plates for subsequent sequencing of the plasmids. The cultures were incubated over night at 37 °C and 180 rpm.
  • Observations and results:
    Both the liquid culture and the dilution plates were positive for bacterial growth. Plates showed the following results:
    104 10 clones
    105 2 clones
    106 0 clones

V07_12_2 1st round: Miniprep of liquid culture V07_11
  • Experiment:
    We used 4 x 5 mL liquid culture split to 4 columns for plasmid preparation with the peqGOLD Plasmid Miniprep Kit (PeqLab).


V07_13


V07_13_1 1st round: Miniprep of clones 1-12 V07_12
  • Experiment:
    Plasmids from the 1st mutation round were prepped using the peqGOLD Plasmid Miniprep Kit (PeqLab) following the user manual. DNA concentrations were determined for subsequent sequencing.
  • Observations and results:
    Both the liquid culture and the dilution plates were positive for bacterial growth. Plates showed the following results:
    104 10 clones
    105 2 clones
    106 0 clones

V07_13_2 1st round: Miniprep of liquid culture V07_11
  • Experiment:
    We used 4 x 5 mL liquid culture split to 4 columns for plasmid preparation with the peqGOLD Plasmid Miniprep Kit (PeqLab).


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