User:Mariana Ruiz Velasco L./Notebook/IGEM 2010/Wet lab journal/2010/09/24

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Hot and cold!

  • I ran a restriction to check whether the mutated red luciferase with mutated RBS restriction was present. The last two lanes indicate that the product has the expected size.

Restr mutluz.JPG

Click beetle luciferases assay

  • Today, I also used the cultures grown overnight to check whether the luciferases showed any brightness. I followed the assay mentioned by Edinburgh's team for two cultures of green emitting luciferase and two cultures of red. To that, I added 1μL of 100mM luciferin and waited in the dark room for ~30min taking pictures with a SLR camera with high ISO (~400), for about 30secs to 1 min exposure. I observed nothing :'(.

  • After the initial disappointment, I talk to Cambridge team and they help us while trying to understand the causes why the assay was a complete disaster. I will continue to check the mutated luciferase from firefly along with the pT7 luciferase.