Team:Paris Saclay/Notebook/August/28

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(Difference between revisions)
(Electrophoresis of 5 digestions of pPS5 by HindIII to check if the insert of CAD is in the right position)
(Electrophoresis of 5 digestions of pPS5 by HindIII to check if the insert of CAD is in the right position)
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[[File:280814 HV degisetion.jpg|300px]]
[[File:280814 HV degisetion.jpg|300px]]
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1-5: pPS5 (pJBEI+CAD) digested by HindIII
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6: pPS1 (pJBEI+Apra)
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7-8: ladder
====Electrophoresis of the same 5 PCR pooled====
====Electrophoresis of the same 5 PCR pooled====

Revision as of 10:32, 29 August 2014

Contents

Thursday 28th August

Lab Work

B - Construction of the fusion protein (color)

Transformation of DH5a by PSB1C3+chromoprotein

by Mélanie

--> a compléter

Addition of adenines at the ends of PCR fragment of chromoprotein

by Laetitia

components volumes
H2O 1 μl
Gotaq buffer 5X 1µl
dATP 1mM 2 µl
Chromoprotein gene fragment (30ng/µL) 5µL
Gotaq polymerase 1µl

1h- 70°C

Ligation of the PCR fragment of chromoprotein + AAA with pGEMTeasy

by Laetitia

components volumes
2X ligation buffer T4 DNA ligase 10 μl
pGEMTeasy 1µl
Ligation mix 7 µl
Ligase 2µL

4h - 16°C


D - lemon scent

Electrophoresis of 5 digestions of pPS5 by HindIII to check if the insert of CAD is in the right position

by Hoang Vu

280814 HV degisetion.jpg


1-5: pPS5 (pJBEI+CAD) digested by HindIII 6: pPS1 (pJBEI+Apra) 7-8: ladder

Electrophoresis of the same 5 PCR pooled

by Laetitia

Gel agarose 0,8% - 100V

The totality of the 5 PCR have been used. Then, we cut the lane corresponding of the Limonene synthase gene on a UV table.

280814 Laetitia verif decoup.jpg

Purification of the PCR product of Limonene synthase (BBa762100)

by Laetitia We used the kit PCR-Clean-UP to extract the DNA from the agarose gel. Elution volume : 15µl


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