Team:Paris Saclay/Notebook/August/27

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Contents

Wednesday 27th August

Lab Work

B - Construction of the fusion protein (color)

PCR clean-up of colonies containing pGEMTeasy+chromoprotein

by Sean

Three colonies were selected from the gel prepared earlier by Laetitia, namely #3,#4, and #6. The three gel samples underwent PCR clean-up.

Gel electrophoresis of PCR clean-up results containing pGEMTeasy+chromoprotein

by Sean

PCR clean-up was performed ealier today. 1µl of each result was used for the electrophoresis. Nothing appeared under UV.

Ligation

by Sean

Transformation of odor free E. coli with plasmids coding Fluo Protein

by Terry

Our fusion protein was not expressed in pGEMTeasy (no color in the culture), so, if our system do not work at all, I'm preparing FP (Fluorescent Protein) for our lemon.

6 plamids coding FP have been transformed in Odor Free :

  • pEYFP
  • pGFP
  • pESFP
  • pRFP+
  • pCFP+
  • pYFP+


D - lemon scent

Plasmide extraction

By Mélanie

Extraction of pPS5 with the phenol protocol (as previously described)

PCR of Limonene synthase (BBa762100)

by Mélanie

5 tubes to do a gradient

components volumes
H2O 29.75μl
Gotaq buffer 10µl
dNTP 10mM 1µl
iPS67 1µM
iPS66 1µM
DNA 1µl
MgCl2 4µl
Gotaq 1.25µl
PCR cycle
step temperature (°C) time
1 95 2min
2 95 30 sec
3 49-55 (gradient) 30sec
4 72 2min
5 72 5min

Electrophoresis of pPS3 and pPS4

by Terry

Result from yesterday 's extraction.

Alice, donne moi la photo.

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