Team:Paris Saclay/Notebook/August/26

From 2014.igem.org

(Difference between revisions)
(→Lab work)
(→Streaking of colonies containing BBa_K762100+pGEMTeasy)
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===D - Lemon Scent===
===D - Lemon Scent===
-
==Streaking of colonies containing BBa_K762100+pGEMTeasy==
+
====Streaking of colonies transformed by BBa_K762100+pGEMTeasy (LS) ====
Due to yesterday's strange results, we conduct 30 streakings in a petri dish: 28 white colonies and 2 blue colonies.
Due to yesterday's strange results, we conduct 30 streakings in a petri dish: 28 white colonies and 2 blue colonies.
 +
====Digestion of pGEMTeasy+ LS by SacI ====
 +
 +
The goal is to check if the limonen synthase gene is really inside the plasmid. We used the two samples named " LS2 " and " LS3 " which are normally pGEMTeasy+LS .
 +
 +
{| class="wikitable centre" width="50%"
 +
|+
 +
|-
 +
! scope=col | component
 +
! scope=col | volume
 +
|-
 +
|Plasmid
 +
|20 μl
 +
|-
 +
|Fast Digest buffer 10X
 +
|3μl
 +
|-
 +
|SacI
 +
|1μl
 +
|-
 +
|H20
 +
|6μl
 +
|}
 +
 +
====Electrophoresis of the digestion products by SacI  ====
 +
 +
T : control negatif without enzyme
 +
 +
{| class="wikitable centre" width="50%"
 +
|+
 +
|-
 +
! scope=col | component
 +
! scope=col | volume
 +
|-
 +
|blue
 +
|2 μl
 +
|-
 +
|Plasmid
 +
|2 μl
 +
|-
 +
|H20
 +
|8μl
 +
|}
 +
 +
D : sample digested by SacI
 +
 +
 +
L - TLS2 - DLS2 - TLS3 - DLS3
{{Team:Paris_Saclay/notebook_footer}}
{{Team:Paris_Saclay/notebook_footer}}

Revision as of 14:09, 26 August 2014

Contents

Tuesday 26th August

Lab work

plasmid extraction : pGEMTeasy + chromoprotein

by Laetitia

We used the bacteria containing pGEMTeasy + chromoprotein launched by Laetitia monday the 25th for the plasmid extraction, 6 independent cultures which come from the 6 stocks made yesterday.

To extract the plasmid, we used the plasmid DNA purification kit (Macherey-Nagel). Protocol for low copy plasmid.

Digestion of pGEMTeasy+chromoproteinby XbaI and PstI

by Laetitia

The goal is to check the presence of the insert inside the plasmid. We digest the 6 samples of purified plasmid.

component volume
Plasmid 10 μl
Fast Digest buffer 10X 1,5 μl
XbaI 0.5μl
PstI 0.5μl
H20 2,5 μl

1h at 37°C

Electrophoresis of the digestion product of pGEMTeasy+chromoprotein

by Laetitia

T : control negatif without enzyme

component volume
blue 2 μl
Plasmid 2 μl
H20 8μl


D : sample digested by XbaI and PstI

D1 - D2 - D3 - D4 - D5 - D6 - T - L


D - Lemon Scent

Streaking of colonies transformed by BBa_K762100+pGEMTeasy (LS)

Due to yesterday's strange results, we conduct 30 streakings in a petri dish: 28 white colonies and 2 blue colonies.

Digestion of pGEMTeasy+ LS by SacI

The goal is to check if the limonen synthase gene is really inside the plasmid. We used the two samples named " LS2 " and " LS3 " which are normally pGEMTeasy+LS .

component volume
Plasmid 20 μl
Fast Digest buffer 10X 3μl
SacI 1μl
H20 6μl

Electrophoresis of the digestion products by SacI

T : control negatif without enzyme

component volume
blue 2 μl
Plasmid 2 μl
H20 8μl

D : sample digested by SacI


L - TLS2 - DLS2 - TLS3 - DLS3

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