From 2011.igem.org
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| - | <h2>27th of July</h2>
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| - | The strain of E. coli with the copy of super-folded GFP that was incubated the previous night has shown limited/no growth in the prepared LB broth. This could have been caused by:<br>
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| - | -No innoculation<br>
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| - | -Kanamycin concentration in LB broth was too high. We had used 85μg/ml.<br><br>
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| - | <b>New protocol:</b><br>
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| - | 50μg/ml is the recommended concentration. We will use 35-40μg/ml of kanamycin to speed up growth.</html>
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| - | <h2>28th of July</h2>
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| - | Transformation of cells with 6, 7 and 8:<br><br>
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| - | -Let competent cell strain 5α thaw for around 10 minutes on ice.<br>
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| - | -Add 2-3μl of DNA.<br>
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| - | -Leave on ice for 20-25 minutes.<br>
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| - | -Heat shock cells at 42°C for 45 seconds.<br>
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| - | -Leave on ice for 10 minutes.<br>
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| - | -Add 500μl of LB broth.<br>
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| - | -Incubate for 1 hour at 37°C.<br>
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| - | -Centrifuge for 1 minute.<br>
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| - | -Remove 100μl off the top of the eppendorf tube. Pour out the rest making sure that the pellet remains in the eppendorf tube.<br>
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| - | -Re-suspend the cells in the 100μl LB broth solution that was removed in the previous step.<br>
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| - | -Add 5μl on a chloramphenicol agar plate (concentration of 35μg/ml).<br>
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| - | -Add the rest of the sample to a second chloramphenicol agar plate.</html>
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Latest revision as of 14:26, 2 August 2011
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