Team:Paris Saclay/Notebook/August/19

From 2014.igem.org

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(Digestion of PCR results)
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We transformed 100µL of competent bacteria and we added the totality of the ligation product (pGEMTeasy containing LS or PS or GS)
We transformed 100µL of competent bacteria and we added the totality of the ligation product (pGEMTeasy containing LS or PS or GS)
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====Preparation of petri dish====
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====Preparation of petri dishes====
''by Terry''
''by Terry''
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{| class="wikitable centre" width="50%"
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|+ 10 dishes with:
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|-
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! scope=col | Antibiotic
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! scope=col | Concentration
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|XGal
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|80μg/ml
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|-
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|IPTG
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|2*10<sup>-3</sup>
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|-
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|Ampicillin
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|10<sup>-3</sup>
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|}-->
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- 250mL LB+Agar
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'''4 petri dish using:'''
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-250 µL X-Gal(80mg/mL)
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- 250 µL ampi
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- 125 µL IPTG
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-->'''4 petri dish using''' :
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- 100mL LB+Agar
- 100mL LB+Agar

Revision as of 09:06, 22 August 2014

Contents

Tuesday 19th August

Lab work

B and D

Electrophoresis with the PpsI plasmide (obtain on thursday) + the chromoprotein PCR: we obtain results as expected

D - Lemon Scent

Digestion of PCR results

by Sean, Hoang Vu, Eugene

components volumes
PCR purified LS 15 μl
FastDigest green buffer 10X 2 μl
PacI 1 µl
H2O 2 µl
components volumes
PCR purified GS 15 μl
FastDigest green buffer 10X 2 μl
PacI 1 µl
H2O 2 µl
components volumes
PCR purified PS 15 μl
FastDigest green buffer 10X 2 μl
PacI 1 µl
H2O 2 µl


components volumes
Purified pPS1 30 μl
FastDigest green buffer 10X 4 μl
PacI 2 µl
H2O 4 µl

Purification of PCR products of limonen synthase

by Laetitia

We used the PCR products of limonen synthase prepared previously : August 18th

The five samples were pooled in one and then purified using this protocol : protocol

Bacterial transformation

by Laetitia

We used DH5alpha competent bacteria (prepared the 14/08) and they were transformed using the ligation product prepared previously August 18th using this protocol : protocol

We transformed 100µL of competent bacteria and we added the totality of the ligation product (pGEMTeasy containing LS or PS or GS)

Preparation of petri dishes

by Terry

10 dishes with:
Antibiotic Concentration
XGal 80μg/ml
IPTG 2*10-3
Ampicillin 10-3
-->

4 petri dish using:

- 100mL LB+Agar

- 100 µL ampi

Bacterial culture

by Laetitia

After the bacterial transformation, we spread for each strain (LS or PS or GS) :

-100µL on LB-Agar+Amp

-100µL on LB-Agar+Amp+IPTG+X-Gal

-200µL on LB-Agar+Amp+IPTG+X-Gal

-100µL of concentrated bacteria on LB-Agar+Amp+IPTG+X-Gal

37°C at night

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