Template:Kyoto/Notebook/Sep 25

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==Sep 25==
==Sep 25==
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===Electrophoresis===
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<!-- こっから -->
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<div class="experiment">
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<span class="author">nakamoto</span>
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{| class="wikitable"
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!Lane||Sample||Enzyme1||Enzyme2
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|-
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|1||100bp ladder||
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|-
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|2||Pcon-antisense (1C3-66E-fwd A2-662 rev)
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|-
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|3||DT (1C3-661-fwd 1C3 662 rev)
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|-
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|4||spinach-DT(1C3-661-fwd 1C3 662 rev)
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|-
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|5||Pcon-antisense-spinach-DT(1C3-66E fwd A2-661 rev)
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|-
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|6||100bp ladder
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|}
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[[File:igku_9251a.jpg]]<br>
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{| class="wikitable"
 +
!Lane||Sample||Enzyme1||Enzyme2
 +
|-
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|2||100bp ladder
 +
|-
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|3||Ptet (1C3 66E fwd-1C3 660 rev)
 +
|-
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|4||Pcon-attenuator(1C3-660 fwd A2-661 rev)
 +
|-
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|5||DT(1C3 662 fwd-1C3 661 rev)
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|-
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|6||Pcon-attenuator-aptamer-DT(1C3 66E fwd-A2-661-rev)
 +
|-
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|7||100bp ladder
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|}
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[[File:igku_9251b.jpg]]<br>
===Colony PCR===
===Colony PCR===
<div class="experiment">
<div class="experiment">

Revision as of 16:46, 26 September 2013

Contents

Sep 25

Electrophoresis

nakamoto

LaneSampleEnzyme1Enzyme2
1100bp ladder
2Pcon-antisense (1C3-66E-fwd A2-662 rev)
3DT (1C3-661-fwd 1C3 662 rev)
4spinach-DT(1C3-661-fwd 1C3 662 rev)
5Pcon-antisense-spinach-DT(1C3-66E fwd A2-661 rev)
6100bp ladder

Igku 9251a.jpg

LaneSampleEnzyme1Enzyme2
2100bp ladder
3Ptet (1C3 66E fwd-1C3 660 rev)
4Pcon-attenuator(1C3-660 fwd A2-661 rev)
5DT(1C3 662 fwd-1C3 661 rev)
6Pcon-attenuator-aptamer-DT(1C3 66E fwd-A2-661-rev)
7100bp ladder

Igku 9251b.jpg

Colony PCR

Nakamoto

Samplebase pair
9/23 Pcon-PT181 attenuator-aptamer12-1R-DT-(5~12)859
9/23 Pcon-PT181 attenuator-RBS-GFP-DT-(5~8)1527
9/23 Pcon-PT181 attenuator-pSB1C3-(5~8)601
PreDenatureDenatureAnnealingExtensioncycle
94 °C94 °C55 °C68 °C--
2 min30 s30 s1min5s30 cycles


Electrophoresis

no name

LaneSampleEnzyme1Enzyme2
1Pcon-PT181 attenuator-aptamer12-1R-DT-5----
2Pcon-PT181 attenuator-aptamer12-1R-DT-6----
3Pcon-PT181 attenuator-aptamer12-1R-DT-7----
4Pcon-PT181 attenuator-aptamer12-1R-DT-8----
5Pcon-PT181 attenuator-aptamer12-1R-DT-9----
6Pcon-PT181 attenuator-aptamer12-1R-DT-10----
7Pcon-PT181 attenuator-aptamer12-1R-DT-11----
8Pcon-PT181 attenuator-aptamer12-1R-DT-12----
91kbp ladder----
10Pcon-PT181 attenuator-RBS-GFP-DT-5----
11Pcon-PT181 attenuator-RBS-GFP-DT-6----
12Pcon-PT181 attenuator-RBS-GFP-DT-7----
13Pcon-PT181 attenuator-RBS-GFP-DT-8----
14Pcon-PT181 attenuator-pSB1C3-5----
15Pcon-PT181 attenuator-pSB1C3-6----
16Pcon-PT181 attenuator-pSB1C3-7----
17Pcon-PT181 attenuator-pSB1C3-8----

Igku 9252.jpg

Transformation

kojima and nakamoto

NameSampleCompetent CellsTotalPlate
Plac+tet aptamer12+Pcon tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-pT181attenuator-DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-Spinach-DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-pT181antisense+DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-pT181attenuator+tet aptamer12-DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon tetR-DT+Pcon-GFP-DT+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Pcon-pT181attenuator+RBS-GFP-DT1µL10µL11µL-
Pcon-pT181attenuator-DT+Pcon-pT181attenuator+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Ptet-pT181antisense+Spinach-DT+Pcon-RBS-GFP-DT+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Pcon-pT181antisense+Spinach-DT+Pcon-pT181attenuator+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Ptet-pT181antisense+Spinach-DT+Pcon-pT181attenuator+RBS-GFP-DT1µL10µL11µL-

NameSampleCompetent CellsTotalPlate
aptamer12-DT(EcoRI+Xbal)+Pcon-pT181attenuator(EcoRI+Spel)2µL20µL22µL-
Ptet(Spel+Pstl)+RBS-GFP-DT(Xbal+Pstl))2µL20µL22µL-

Ligation

nakamoto

stateVectorInserterLigation High ver.2
experiment9/25 aptamer12_1R-DT(EcoRI&XbaI)4.5 µLPcon-pT181 attenuator(EcoRI&SpeI)7.7 µL6.1 µL
experiment9/17 pSB1C3(EcoRI&SpeI)2.0 µL9/3 pT181 attenuator(EcoRI&SpeI)14 µL8 µL
experiment9/25 Pcon-pT181 attenuator(EcoRI&XbaI)4.5 µL9/25 RBS-GFP-DT(EcoRI&SpeI)19.3 µL11.9 µL
experiment9/25 Pcon-RBS-GFP-DT(EcoRI&XbaI)4.2 µLPcon-RBS-tetR-DT(EcoRI&SpeI)20 µL12.1 µL

Digestion

Liquid Culture

Kojima

Samplemedium
DT -
pT181attenuator -
Pcon-RBS-tetR-DT -

Gel Extraction

Kojima

LaneDNAEnzyme
1100bp ladder-
2Pcon-pT181antisense
3
4
5-
6DT
7
8
9-
10Spinach-DT
11
12

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

Nameconcentration[µg/mL]260/280260/230
（´ω｀）
（ΦωΦ^）


LaneDNAEnzyme
1
2
3
4
5
6

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

Nameconcentration[µg/mL]260/280260/230
（´ω｀）
（ΦωΦ^）


LaneDNAEnzyme
1
2
3
4
5
6

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

Nameconcentration[µg/mL]260/280260/230
（´ω｀）
（ΦωΦ^）


LaneDNAEnzyme
1
2
3
4
5
6

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

Nameconcentration[µg/mL]260/280260/230
（´ω｀）
（ΦωΦ^）


LaneDNAEnzyme
1
2
3
4
5
6

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

Nameconcentration[µg/mL]260/280260/230
（´ω｀）
（ΦωΦ^）


Nakamoto

LaneDNAEnzyme
1
2
3
4
5
6

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

Nameconcentration[µg/mL]260/280260/230
（´ω｀）
（ΦωΦ^）

Restriction Enzyme Digestion

Tatsui

DNArestricion enzymerestriction enzymeBufferBSAMilliQtotal
2 cuts（°°） µL µL µL µL µL µL µL
NC µL µL µL µL µL µL µL

Liquid Culture

No name

Samplemedium
Pcon-pT181anntisense-spinach-DT(4k5)-

Ligation

Nakamoto and Tatsui

stateVectorInserterLigation High ver.2
experimentnamexx µLnamexx µLxx µL
NCnamexx µLnamexx µLxx µL

PCR

No name

genome DNAKOD plus10x bufferdNTPMgSO4SasA_fwd primerSasA_rev primerMilliQtotal
0.52.52.51.50.750.75 25
PreDenatureDenatureAnnealingExtensioncycle
　°C　°C　°C　°C--
（◎Д◎）　　　　

Transformation

Tatsui

NameSampleCompetent CellsTotalPlate
aptamer12-1R-DTµLµLµLCP
pSB1C3(EcoRI+Spel)+pT181attenuator(EcoRI+Spal)µLµLµLCP
Pcon-pT181attenuator(EcoRI+Xbal)+RBS-GFP-DT(EcoRI+Spel)µLµLµLAmp
Pcon-RBS-GFP-DT(EcoRI+Xbal)+Pcon-RBS-tetR-DT(EcoRI+Spel)µLµLµLAmp
pSB1C3(EcoRI+Spel)+Pcon-pT181attenuator-aptamer12-1R-DT(EcoRI+Spel)µLµLµLCP

Electrophoresis

Hirano

LaneSampleEnzyme1Enzyme2
1100bp ladder
2DT(2-1)

File:Igku xxxxxx.xxx

精製？

Restriction Enzyme Digestion

DNArestricion enzymerestriction enzymeBufferBSAMilliQtotal
2 cuts（°°） µL µL µL µL µL µL µL
NC µL µL µL µL µL µL µL

液体培養

プレーティング

Colony PCR

No name

Samplebase pair
(｀・ω・´)
PreDenatureDenatureAnnealingExtensioncycle
　°C　°C　°C　°C--
（◎Д◎）　　　　

RNA抽出

cDNA合成

Electrophoresis

Nakamoto

LaneSampleEnzyme1Enzyme2
1100bp ladder
2Pcon-RBS-GFP-DT(RNA)

File:Igku xxxxxx.xxx

Ligation

じっけんしゃ

stateVectorInserterLigation High ver.2
experimentnamexx µLnamexx µLxx µL
NCnamexx µLnamexx µLxx µL
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