User:Mariana Ruiz Velasco L./Notebook/IGEM 2010/Wet lab journal/2010/05/18

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05/18/2010: Oops, change of plans

  • When I was just about to quantify the double terminator with the purified luciferase(pur.luc.), I realized that the band size of the second one was not the expected one (~1600 bp). In order to discard the fact that the band observed was that of the plasmid, but not the luciferase per se, I ran a gel of the pur.luc. and of the strain with the plasmid and luciferase. The results showed the plasmid and the luciferase, but the pur.luc. wasn't there.

  • In order to make sure that the next steps of the week plan are achieved successfully (ligation, transformation, induced mutation, etc), I decided to repeat the band purification.

  • I made a double restriction using ECOR1 and SPEI, and incubated ON at 37°C. The reaction (40μL) was done by duplicate using as DNA input the previously used strains of Arturo Zepeda. It was stored as Luciferasa restriccion ECO RI y SPE I para purificación (1-B).

-H2O ------> 6μl

-BSA ------> 1μl

-Buffer ---> 4μl

-ECO RI --> 2μl

-SPEI -----> 2μl

-DNA ------> 25μl