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From 2011.igem.org

Contents 1 Baptiste, Hovannes & Ouriel 1.1 First conclusions on our methods 2 Kevin 2.1 Following step for characterisation of tetO array

Baptiste, Hovannes & Ouriel

Microscopy experiments of today.

As we begin to master totally the Nikon microscope we chose to run three settings for the 3610 gfp-/3610 gfp+ experiments. The aim is to see which way is the best to plate them on our slides and more important at which growth state. We start for the same overnight culture and then create three microscopic slides from our two 3610 strains:

1. One directly from the overnight culture in stationnary phase.
2. One diluting until the OD reached 0.1. We then wait for the OD to grow to 0.8 (roughly 2 hours) and we concentrate it as usual (4000 rpm during 10 min and then put 200 microliters of LB on the pellet).
3. One by diluting 1/500 the overnight culture. We then wait for the OD to grow to 0.8 (roughly 4 hours) and we concentrate it as usual (4000 rpm during 10 min and then put 200 microliters of LB on the pellet).

First conclusions on our methods

Despite what we thought, the autofocus programme on the Nikon is not perfect. We need to adjust it a little bit to our needs. It gave us slightly blurry image for the second serie of slide (dilution to an OD of 0.1) and catastrophic results for the stationnary phase slide. Fortunately, the stationnary phase means that bacteria do not grow and we were able to interpret the results even missing over an hour of images. Our slide for the 1/500 dilution was very faulty. We could not see anything clearly and we will have to retry it tomorrow. Letting the drop dry or not is the primary issue here. We will try both ways tomorrow to see which seems better.

Kevin

Following step for characterisation of tetO array

Double transformation works ! Tomorrow microscopy.