Template:Kyoto/Notebook/Sep 2

From 2013.igem.org

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===EDTA Solution===
 
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<div class="experiment">
 
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<span class="author">Hirano</span>
 
===Colony PCR===
===Colony PCR===
<div class="experiment">
<div class="experiment">
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<span class="author">Hirano</span>
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<span class="author">No name</span>
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{| class="wikitable"
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!|Sample||base pair
 +
|-
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|8/27 pT181 attenuator-DT-1||738
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|-
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|8/27 pT181 attenuator-DT-2||738
 +
|-
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|8/27 pT181 attenuator-DT-3||738
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|-
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|8/27 TetR-aptamer 12_1R-DT-1||521
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|-
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|8/27 TetR-aptamer 12_1R-DT-2||521
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|-
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|8/27 pT181 antisense-DT-1||542
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|}
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{| class="wikitable"
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!PreDenature||Denature||Annealing||Extension||cycle
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|-
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|94&deg;C||94&deg;C||55&deg;C||68&deg;C||--
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|-
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|5min||30s||30s||30s||30cycles
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|}
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[[File:igku_Aug28electrophoresis3?.png]]
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{| class="wikitable"
 +
!|Sample||base pair
 +
|-
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|8/27 P&lambda;-luxI-1||???
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|-
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|8/27 P&lambda;-luxI||???
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|}
 +
{| class="wikitable"
 +
!PreDenature||Denature||Annealing||Extension||cycle
 +
|-
 +
|94&deg;C||94&deg;C||55&deg;C||68&deg;C||--
 +
|-
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|5min||30s||30s||30min||3cycles
 +
|}
 +
[[File:igku_Aug19electrophoresis1.png]]
 +
</div>
===Master Plate===
===Master Plate===

Revision as of 06:40, 23 September 2013

Contents

Sep 2

Liquid Culture

Hirano

Samplemedium
9/1 entA-(Master Plate)-14mL SOB(+Km)

37°C

Colony PCR

Tatsui

Samplebase pair
9/1 RBS-lysis2-DT-(1)985
Ptet-RBS-lacZα-DT-(1)765
Plac-RBS-lacZα-DT-(1)765
Plac-RBS-lacZα-DT-(2)765
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s1min30cycles

File:Igku sep2electrophoresis1.png

Restriction Enzyme Digestion

Nakamoto

pSB1C3-(1)EcoRISpeIXbaIPstIBufferBBufferDBSAMilliQtotal
2 cuts(E+S)7µL1µL1µL0µL0µL3µL0µL0.3µL17.7µL30µL
NC(E+S)0.3µL0µL0µL0µL0µL1µL0µL0.1µL8.6µL10µL
2 cut(X+P)s7µL0µL0µL1µL1µL0µL3µL0.3µL17.7µL30µL
NC(X+P)0.3µL0µL0µL0µL0µL0µL1µL0.1µL8.6µL10µL
8/21 tRMA-spinach(1)EcoRISpeIBufferBBSAMilliQtotal
2 cuts8µL1.0µL1.0µL3µL0.3µL17.6µL30µL
NC0.5µL0µL0µL1µL0.1µL8.4µL10µL
8/21 tetR aptamer12_1R-(1)EcoRISpeIBufferBBSAMilliQtotal
2 cuts6µL1.0µL1.0µL3µL0.3µL18.7µL30µL
NC0.4µL0µL0µL1µL0.1µL8.5µL10µL
8/21 pT181 attenuator-(2)EcoRISpeIXbaIPstIBufferBBufferDBSAMilliQtotal
2 cuts(E+S)8µL1µL1µL0µL0µL3µL0µL0.3µL16.3µL30µL
NC(E+S)0.5µL0µL0µL0µL0µL1µL0µL0.1µL8.4µL10µL
2 cut(X+P)s8µL0µL0µL1µL1µL0µL3µL0.3µL16.3µL30µL
NC(X+P)0.5µL0µL0µL0µL0µL0µL1µL0.1µL8.4µL10µL

Liquid Culture

Kojima

Samplemedium
tRNA-Spinach-1Plusgrow medium(+CP)
tetR aptamaer12_1R-1Plusgrow medium(+CP)
pT181 attenuator-1Plusgrow medium(+CP)　 = pT181 antisense-1Plusgrow medium(+CP)

Master Plate

Kojima

NumberUse LB plate(+CP)
1tRNA Spinach-1
2tetR aptamaer12_1R
3tetR aptamaer12_P
4tetR aptamaer12_1M
5pT181 attenuator-2
6Fusion1 attenuator-1
7Fusion3m2 attenuator-1
8pT181 antisense-1
9Fusion1 antisense-1
10Fuaion6 antisense-1

Gel Extraction

No name

LaneDNAEnzyme
1100bp ladder-
2pSB1C3(E+S)EcoRI&SpeI
3
4----
58/24 tetR aptamer 12_1REcoRI&SpeI
6

File:Igku xxbeforexx.xxx
File:Igku xxafterxx.xxx
No name

LaneDNAEnzyme
1100bp ladder-
2pT181 attenuator(2)(E+S)EcoRI&SpeI
3
4----
5pT181 attenuator(2)(X+P)EcoRI&SpeI
6
7----
8pT181 attenuator(2)(X+P)EcoRI&SpeI
9
10----
11SpinachEcoRI&SpaI
12

File:Igku xxbeforexx.xxx
File:Igku xxafterxx.xxx

Nameconcentration[µg/mL]260/280260/230
pSB1C3 (EcoRI&SpeI)3.02.180.31
pSB1C3 (XbaI&PstI)4.72.250.36
pT181 attenuator-(2)(EcoR&SpeI)8.62.740.01
pT181 attenuator-(2) (XbaI&PstI)16.52.460.03
Spinach (EcoRI&SpeI)2.82.980.27
tetR aptamer12_1R (EcoRI&SpeI)50.428.071.97

Colony PCR

No name

Samplebase pair
8/27 pT181 attenuator-DT-1738
8/27 pT181 attenuator-DT-2738
8/27 pT181 attenuator-DT-3738
8/27 TetR-aptamer 12_1R-DT-1521
8/27 TetR-aptamer 12_1R-DT-2521
8/27 pT181 antisense-DT-1542
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s30s30cycles

File:Igku Aug28electrophoresis3?.png

Samplebase pair
8/27 Pλ-luxI-1???
8/27 Pλ-luxI???
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s30min3cycles

File:Igku Aug19electrophoresis1.png

Master Plate

Hirano

Liquid Culture

Hirano

Colony PCR

Hirano

Transformation

Hirano
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