Team:Paris Liliane Bettencourt/Notebook/2011/08/26/

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Cyrille

Biobricking of YFP-TetR

The miniprep of 8 clones from the quick change mutagenesis transformation where minipreped. As there are no sequencing facilities in the week-end, a digestion test will be carried on the miniprep.

500 ng for each miniprep was digested by PstI in 10µL.

A gel was runned with the digestion. The worrect bands will be extracted.

ladder-1-2-3-4-5-6-7

firts cutting attempt.jpg

Preparation of the GFP amber

A gel was runned from the different miniprep found in the frigde to identofy the good ones. The gel is the following.

ladder - GFP T7t 1 - GFP T7t 2 -GFP T7t 3 -GFP T7t 4 - GFPmut3b

GFP T7t 12-6 seems to have the correct size. GFPmut3b as well.

Danyel

Two clones of the strain S79 (holding pt7 + RBS + GFP + Ter) were transformed last night into BL21 strains. The strain transformed with clone number 3 was already fluorescent without IPTG. Both plates showed strongly fluorescent colonies with IPTG. The functionality of the S79 construct is thus verified. One more transformation was done today with the clone for which a glycerol has already been made.

Camille

I discovered (Thanks to Danyel) I paid attention to the mistakes I did yesterday. Indeed I digested all my inserts with Ecor1 and Pst1, when it was needed, for the integration in the S46 and S58 a digestion in Ecor1 and Spe1. Therefore I used 3 clons of the only ligation that should be ok (pSB1C3 + pHyperspank) and launched it in liquid culture.

I then digested the digestion product from the other day(pHyperspank in Ecor1 and Pst1) and digested it with Spe1. I then purified it with a PCR purification kit. I couldn't measure the concentration in DNA because of the TECAN was already used. I used the gel extraction of the digested S46 to do the ligation and transformed it then. Babak did my minipreps of the strain containing pHyperspank in pSB1C3: