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From 2011.igem.org

Cyrille

Going on with the QCM experiments

The transformation have worked pretty well. The negative control before digestion has some clone, and the negative control after digestion has none. On all the plate, we have a important number of colony, more important than in any negatives control. This indicate that the PCR may have worked.

On the selected clones, the last two (11, and 12) comes from very small colonies on the plate. The hypothesis is that they might be parasite, or clones grown with a non metilated plasmid. Anyway, the yield obtained after the miniprep is poor, and they may come from parasite.

The miniprep was done over 12 selected clones. The culture was grown overday, started at 9:30 and centrifugated at 15:00. The yields are around 500ng/µL depending on the the tubes. The concentration was measured with the Tecan machine.

1µg of each of the 12 clones (except for the last two, from which we took the maximum quantity that is to say 8 µL) was digested using EcoRI-Fast digest in green digestion buffer, for 15 min at 37°C. The result was loaded on the gel, and let run for 20 min at 50V and then at 100V for the rest. Here is the result of yhe gel:

Kevin