Team:Paris Saclay/Notebook/August/28

From 2014.igem.org

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(Electrophoresis of 5 digestions of pPS5 by HindIII to check if the insert of CAD is in the right position)
(Lab Work)
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==Lab Work==
==Lab Work==
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===B - Construction of the fusion protein (color)===
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===Construction of the fusion protein (color)===
====Transformation of DH5a by PSB1C3+chromoprotein====
====Transformation of DH5a by PSB1C3+chromoprotein====
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===D - lemon scent===
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===lemon scent===
==== 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  ====

Revision as of 09:11, 14 October 2014

Contents

Thursday 28th August

Lab Work

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


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), our witness

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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