Template:Kyoto/Notebook/Sep 2

From 2013.igem.org

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===Gel Extraction===
===Gel Extraction===
<div class="experiment">
<div class="experiment">
 +
<span class="author">No name</span>
 +
{| class="wikitable"
 +
!Lane||DNA||Enzyme
 +
|-
 +
|1||100bp ladder||-
 +
|-
 +
|2||rowspan="2"|pSB1C3(E+S)|rowspan="2"||EcoRI&SpeI
 +
|-
 +
|3
 +
|-
 +
|4||--||--
 +
|-
 +
|5||rowspan="2"|8/24 tetR aptamer 12_1R||rowspan="2"|EcoRI&SpeI
 +
|-
 +
|6
 +
|}
 +
[[File:igku_xxbeforexx.xxx]]<br>
 +
[[File:igku_xxafterxx.xxx]]<br>
 +
<span class="author">No name</span>
 +
{| class="wikitable"
 +
!Lane||DNA||Enzyme
 +
|-
 +
|1||100bp ladder||-
 +
|-
 +
|2||rowspan="2"|pT181 attenuator(2)(E+S)|rowspan="2"||EcoRI&SpeI
 +
|-
 +
|3
 +
|-
 +
|4||--||--
 +
|-
 +
|5||rowspan="2"|pT181 attenuator(2)(X+P)||rowspan="2"|EcoRI&SpeI
 +
|-
 +
|6
 +
|-
 +
|7||--||--
 +
|-
 +
|8||rowspan="2"|pT181 attenuator(2)(X+P)||rowspan="2"|EcoRI&SpeI
 +
|-
 +
|9
 +
|-
 +
|10||--||--
 +
|-
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|11||rowspan="2"|Spinach||rpwspan="2"|EcoRI&SpaI
 +
|-
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|12
 +
|}
 +
[[File:igku_xxbeforexx.xxx]]<br>
 +
[[File:igku_xxafterxx.xxx]]<br>
 +
{| class="wikitable"
 +
!Name||concentration[&micro;g/mL]||260/280||260/230
 +
|-
 +
|pSB1C3 (EcoRI&SpeI)||3.0||2.18||0.31
 +
|-
 +
|pSB1C3 (XbaI&PstI)||4.7||2.25||0.36
 +
|-
 +
|pT181 attenuator-(2)(EcoR&SpeI)||8.6||2.74||0.01
 +
|-
 +
|pT181 attenuator-(2) (XbaI&PstI)||16.5||2.46||0.03
 +
|-
 +
|Spinach (EcoRI&SpeI)||2.8||2.98||0.27
 +
|-
 +
|tetR aptamer12_1R (EcoRI&SpeI)||50.4||28.07||1.97
 +
|}
</div>
</div>

Revision as of 06:37, 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)|rowspan="2"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)|rowspan="2"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

EDTA Solution

Hirano

Colony PCR

Hirano

Master Plate

Hirano

Liquid Culture

Hirano

Colony PCR

Hirano

Transformation

Hirano
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