Team:KIT-Kyoto/Notebook/Labnote

From 2014.igem.org

(Difference between revisions)
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<!--Preparations for SDS-PAGE-->
<!--Preparations for SDS-PAGE-->
<strong>Preparations for SDS-PAGE</strong>
<strong>Preparations for SDS-PAGE</strong>
 +
<tr>
Add SDS sample buffer to protein extractions in a flask, incubate at 37℃ for 15 minutes and keep it at 4℃.
Add SDS sample buffer to protein extractions in a flask, incubate at 37℃ for 15 minutes and keep it at 4℃.
<!--Preparations for SDS-PAGE終わり-->
<!--Preparations for SDS-PAGE終わり-->
Line 826: Line 827:
<!--本文終わり-->
<!--本文終わり-->
 +
<!-- タイトル -->
 +
<h3>June 19, 2014</h3>
 +
<!-- タイトル終わり -->
 +
 +
<!-- 本文開始 -->
 +
<p class="sentence">
 +
<!-- 実験者名 -->
 +
KITOMI<br>
 +
<!-- 実験者名終わり -->
 +
<!--SDS-PAGE-->
 +
<strong>SDS-PAGE</strong>
 +
<table class="materials">
 +
<tr>
 +
<th rowspan="2">Host</th>
 +
<th colspan="2">Sample</th>
 +
<th rowspan="2">IPTG</th>
 +
</tr>
 +
<tr>
 +
<th>Vector</th>
 +
<th>Insert</th>
 +
</tr>
 +
<tr>
 +
<td>BL21(DE3)pLysS</td>
 +
<td colspan="2">pGEX6P-2</td>
 +
<td>induced</td>
 +
</tr>
 +
<tr>
 +
<td>BL21(DE3)pLysS</td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE1</td>
 +
<td>induced</td>
 +
</tr>
 +
<tr>
 +
<td>BL21(DE3)pLysS</td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTS2</td>
 +
<td>induced</td>
 +
<tr>
 +
<td>BL21(DE3)pLysS</td>
 +
<td colspan="2">pGEX6P-2</td>
 +
<td>non-induced</td>
 +
</tr>
 +
<tr>
 +
<td>BL21(DE3)pLysS</td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTS1</td>
 +
<td>non-induced</td>
 +
<tr>
 +
<td>BL21(DE3)pLysS</td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE2</td>
 +
<td>non-induced</td>
 +
</tr>
 +
</table>
 +
<!--SDS-PAGE終わり-->
 +
 +
 +
<br>
 +
</p>
 +
<!--本文終わり-->
 +
 +
<!-- タイトル -->
 +
<h3>June 21, 2014</h3>
 +
<!-- タイトル終わり -->
 +
 +
<!-- 本文開始 -->
 +
<p class="sentence">
 +
<!-- 実験者名 -->
 +
SOGA<br>
 +
<!-- 実験者名終わり -->
 +
<!--Miniprep-->
 +
<strong>Mineprep</strong>
 +
<table class="materials">
 +
<tr>
 +
<th rowspan="2">Host</th>
 +
<th colspan="2">Sample</th>
 +
</tr>
 +
<tr>
 +
<th>Vector</th>
 +
<th>Insert</th>
 +
</tr>
 +
<tr>
 +
<td>BL21(DE3)pLysS</td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMEST1</td>
 +
</tr>
 +
<tr>
 +
<td>BL21(DE3)pLysS</td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE2</td>
 +
</tr>
 +
</table>
 +
<!--Mineprep終わり-->
 +
<br>
 +
<!--PCR-->
 +
<strong>PCR</strong>
 +
<table class="materials">
 +
<tr>
 +
<th rowspan="2">Vector</th>
 +
<th colspan="2">Sample</th>
 +
<th rowspan="2">DNA Polymerase</th>
 +
</tr>
 +
<tr>
 +
<th>Insert</th>
 +
<th>Primer</th>
 +
</tr>
 +
<tr>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE1</td>
 +
<td>P-F1,P-R
 +
<td>KOD-FX-NEO</td>
 +
</tr>
 +
<tr>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE2</td>
 +
<td>P-F2,P-R</td>
 +
<td>KOD-FX-NEO</td>
 +
</tr>
 +
<tr>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE1</td>
 +
<td>P-F1,P-R</td>
 +
<td>KOD-FX-NEO</td>
 +
<tr>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE2</td>
 +
<td>P-F2,P-R</td>
 +
<td>KOD-FX-NEO</td>
 +
</tr>
 +
</table>
 +
<!--PCR終わり-->
 +
<br>
 +
<!--AGE-->
 +
<strong>AGE</strong>
 +
<table class="materials">
 +
<tr>
 +
<th rowspan="2">Lane</th>
 +
<th colspan="2">Sample/Marker</th>
 +
<th rowspan="2">Insert</th>
 +
<th rowspan="2">Primer</th>
 +
</tr>
 +
<tr>
 +
<th>Host</th>
 +
<th>Vector</th>
 +
</tr>
 +
<tr>
 +
<td>1</td>
 +
<td colspan="4">1kb Ladder</td>
 +
</tr>
 +
<tr>
 +
<td>2</td>
 +
<td>BY4247</td>
 +
<td colspan="3">p427-TEF</td>
 +
</tr>
 +
<tr>
 +
<td>3</td>
 +
<td></td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE1</td>
 +
<td>P-F1,P-R</td>
 +
<tr>
 +
<td>4</td>
 +
<td></td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE2</td>
 +
<td>P-F2,P-R</td>
 +
</tr>
 +
<tr>
 +
<td>5</td>
 +
<td></td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE1</td>
 +
<td>P-F1,P-R</td>
 +
</tr>
 +
<tr>
 +
<td>6</td>
 +
<td></td>
 +
<td>pGEX6P-2</td>
 +
<td>CuMTSE2</td>
 +
<td>P-F2,P-R</td>
 +
</tr>
 +
</table>
 +
<!--AGE終わり-->
 +
 +
<br>
 +
</p>
 +
<!--本文終わり-->

Revision as of 08:40, 2 October 2014

LabNote

June

June 10, 2014

OKAMOTO
Miniprep

Host Sample
Vector Insert
BL21(DE3) pGEX6P-2 CuMTSE1
BL21(DE3) pGEX6P-2 CuMTSE2
BL21(DE3) pGEX6P-2 CuMTS3
BL21(DE3) pGEX6P-2 CuMTS62

Restriction Digest
Host Sample Enzyme Buffer
Vector Insert
BL21(DE3)PlysS pGEX6P-2 CuMTSE1 EcoR1 Buffer H
BL21(DE3)PlysS pGEX6P-2 CuMTSE2 EcoR1 Buffer H
BL21(DE3)PlysS pGEX6P-2 CuMTS3 EcoR1 Buffer H
BL21(DE3)PlysS pGEX6P-2 CuMTSE62 EcoR1 Buffer H
BL21(DE3)PlysS pGEX6P-2 CuMTSE1 Sal1, Not1 Buffer H
BL21(DE3)PlysS pGEX6P-2 CuMTSE2 Sal1, Not1 Buffer H
BL21(DE3)PlysS pGEX6P-2 CuMTS3 Sal1, Not1 Buffer H
BL21(DE3)PlysS pGEX6P-2 CuMTSE62 Sal1, Not1 Buffer H

June 11, 2014

KOBAYASHI
AGE

Lane Sample/Marker Enzyme
Vector Insert
1 λDNA-HindⅢ
2 pGEX6P-2 CuMTSE1 N.D
3 pGEX6P-2 CuMTSE1 EcoR1
4 pGEX6P-2 CuMTSE1 Sal1,Not1
5 pGEX6P-2 CuMTSE2 N.D
6 pGEX6P-2 CuMTSE2 EcoR1
7 pGEX6P-2 CuMTSE2 Sal1, Not1
8 pGEX6P-2 CuMTS3 N.D
9 pGEX6P-2 CuMTS3 EcoR1
10 pGEX6P-2 CuMTS62 N.D
11 pGEX6P-2 CuMTS62 EcoR1
12 pGEX6P-2 CuMTS62 Sal1,Not1

June 12, 2014

KOBAYASHI
Transformation

Host Sample
Vector Insert
BL21(DE3)pLysS pGEX6P-2 CuMTSE1
BL21(DE3)pLysS pGEX6P-2 CuMTSE2
BL21(DE3)pLysS pGEX6P-2 CuMTS3
BL21(DE3)pLysS pGEX6P-2 CuMTS62

June 13, 2014

KOBAYASHI
Colony Isolation
Select 6 colonies per plate from 4 plates from June 12th.

Host Sample Section
Vector Insert
BL21(DE3)pLysS pGEX6P-2 CuMTSE1 1-a,1-b
BL21(DE3)pLysS pGEX6P-2 CuMTSE2 2-a,2-b
BL21(DE3)pLysS pGEX6P-2 CuMTS3 3-a,3-b
BL21(DE3)pLysS pGEX6P-2 CuMTSE62 6-a,6-b

June 14, 2014

ONISHI
Miniprep

Host Sample
Vector Insert
1-a pGEX6P-2 CuMTSE1
1-b pGEX6P-2 CuMTSE1
2-a pGEX6P-2 CuMTSE2
2-b pGEX6P-2 CuMTSE2
3-a pGEX6P-2 CuMTS3
3-b pGEX6P-2 CuMTS3
6-a pGEX6P-2 CuMTS62
6-b pGEX6P-2 CuMTS62

Miniprep
Host Sample
Vector Insert
BL21(DE3) pGEX6P-2 CuMTSE1
BL21(DE3) pGEX6P-2 CuMTSE2
BL21(DE3) pGEX6P-2 CuMTS3
BL21(DE3) pGEX6P-2 CuMTS62

Restriction Digest
Host Sample Enzyme Buffer
Vector Insert
1-a pGEX6P-2 CuMTSE1 EcoR1 Buffer H
1-b pGEX6P-2 CuMTSE1 EcoR1 Buffer H
2-a pGEX6P-2 CuMTSE2 EcoR1 Buffer H
2-b pGEX6P-2 CuMTSE2 EcoR1 Buffer H
3-a pGEX6P-2 CuMTS3 EcoR1 Buffer H
3-b pGEX6P-2 CuMTS3 EcoR1 Buffer H
6-a pGEX6P-2 CuMTS62 EcoR1 Buffer H
6-b pGEX6P-2 CuMTS62 EcoR1 Buffer H
BL21(DE3) pGEX6P-2 CuMTSE1 EcoR1 Buffer H
BL21(DE3) pGEX6P-2 CuMTSE2 EcoR1 Buffer H
BL21(DE3) pGEX6P-2 CuMTS3 EcoR1 Buffer H
BL21(DE3) pGEX6P-2 CuMTS62 EcoR1 Buffer H

AGE
Lane Sample/Marker Insert Enzyme Buffer
Host Vector
1 1kb Ladder
2 λDNA-HindⅢ
3 1-a pGEX6P-2 CuMTSE1 EcoR1 Buffer H
4 1-b pGEX6P-2 CuMTSE1 EcoR1 Buffer H
5 2-a pGEX6P-2 CuMTSE2 EcoR1 Buffer H
6 2-b pGEX6P-2 CuMTSE2 EcoR1 Buffer H
7 3-a pGEX6P-2 CuMTS3 EcoR1 Buffer H
8 3-b pGEX6P-2 CuMTS3 EcoR1 Buffer H
9 6-a pGEX6P-2 CuMTS62 EcoR1 Buffer H
10 6-b pGEX6P-2 CuMTS62 EcoR1 Buffer H
11 BL21(DE3) pGEX6P-2 CuMTSE1 EcoR1 Buffer H
12 BL21(DE3) pGEX6P-2 CuMTSE2 EcoR1 Buffer H
13 BL21(DE3) pGEX6P-2 CuMTS3 EcoR1 Buffer H
14 BL21(DE3) pGEX6P-2 CuMTS62 EcoR1 Buffer H

Applied λDNA-HindⅢ to compare and contrast AGE of July 14th and June 11th.


June 16, 2014

YAMAJI
Preculture

Host Sample
Vector Insert
BL21(DE3)pLysS pGEX6P-2 CuMTSE1
BL21(DE3)pLysS pGEX6P-2 CuMTSE2
BL21(DE3)pLysS pGEX6P-2 CuMTS3
BL21(DE3)pLysS pGEX6P-2 CuMTS62
BL21(DE3)pLysS pGEX6P-2

Main Culture
Host Sample IPTG
Vector Insert
BL21(DE3)pLysS pGEX6P-2 induced
BL21(DE3)pLysS pGEX6P-2 CuMTSE1 induced
BL21(DE3)pLysS pGEX6P-2 CuMTS2 induced
BL21(DE3)pLysS pGEX6P-2 non-induced
BL21(DE3)pLysS pGEX6P-2 CuMTS1 non-induced
BL21(DE3)pLysS pGEX6P-2 CuMTSE2 non-induced

Protein Extraction(E.coli)
Host Sample IPTG
Vector Insert
BL21(DE3)pLysS pGEX6P-2 induced
BL21(DE3)pLysS pGEX6P-2 CuMTSE1 induced
BL21(DE3)pLysS pGEX6P-2 CuMTS2 induced
BL21(DE3)pLysS pGEX6P-2 non-induced
BL21(DE3)pLysS pGEX6P-2 CuMTS1 non-induced
BL21(DE3)pLysS pGEX6P-2 CuMTSE2 non-induced

Preparations for SDS-PAGE Add SDS sample buffer to protein extractions in a flask, incubate at 37℃ for 15 minutes and keep it at 4℃.

June 19, 2014

KITOMI
SDS-PAGE

Host Sample IPTG
Vector Insert
BL21(DE3)pLysS pGEX6P-2 induced
BL21(DE3)pLysS pGEX6P-2 CuMTSE1 induced
BL21(DE3)pLysS pGEX6P-2 CuMTS2 induced
BL21(DE3)pLysS pGEX6P-2 non-induced
BL21(DE3)pLysS pGEX6P-2 CuMTS1 non-induced
BL21(DE3)pLysS pGEX6P-2 CuMTSE2 non-induced

June 21, 2014

SOGA
Mineprep

Host Sample
Vector Insert
BL21(DE3)pLysS pGEX6P-2 CuMEST1
BL21(DE3)pLysS pGEX6P-2 CuMTSE2

PCR
Vector Sample DNA Polymerase
Insert Primer
pGEX6P-2 CuMTSE1 P-F1,P-R KOD-FX-NEO
pGEX6P-2 CuMTSE2 P-F2,P-R KOD-FX-NEO
pGEX6P-2 CuMTSE1 P-F1,P-R KOD-FX-NEO
pGEX6P-2 CuMTSE2 P-F2,P-R KOD-FX-NEO

AGE
Lane Sample/Marker Insert Primer
Host Vector
1 1kb Ladder
2 BY4247 p427-TEF
3 pGEX6P-2 CuMTSE1 P-F1,P-R
4 pGEX6P-2 CuMTSE2 P-F2,P-R
5 pGEX6P-2 CuMTSE1 P-F1,P-R
6 pGEX6P-2 CuMTSE2 P-F2,P-R

July

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