Template:Kyoto/Notebook/Sep 14

From 2013.igem.org

(Difference between revisions)
(Colony PCR)
(Liquid Culture)
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===Liquid Culture===
===Liquid Culture===
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<div class="experiment">
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<span class="author">No name</span>
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!Sample||medium
!Sample||medium
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|8/29 Pcon-RBS-GFP-DT-Pcon-RBS-luxR-DT-(1)||Plusgrow medium(+Amp)
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|9/13 Pλ-luxI(1)-1||Plusgrow medium(+Amp)
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|-
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|8/29 Pcon-RBS-GFP-DT-Pcon-RBS-luxR-DT-(2)||Plusgrow medium(+Amp)
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|8/29 Pcon-RBS-GFP-DT-Pcon-RBS-luxR-DT-(3)||Plusgrow medium(+Amp)
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|8/29 Pcon-RBS-GFP-DT-Pcon-RBS-luxR-DT-(4)||Plusgrow medium(+Amp)
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|8/29 Plux-RBS-GFP-DT-(1)||Plusgrow medium(+CP)
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|8/29 Plux-RBS-GFP-DT-(2)||Plusgrow medium(+CP)
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|8/29 Pbad/araC-RBS-RFP-DT-(1)||Plusgrow medium(+CP)
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|8/29 Pbad-araC-RBS-RFP-DT-(2)||Plusgrow medium(+CP)
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|RBS-lysis3-DT-(1)||Plusgrow medium(+CP)
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Revision as of 14:37, 25 September 2013

Contents

Sep 14

Restriction Enzyme Digestion

Hirano

9/14 pSB4K5SEcoRIXbaISpeIPstIBufferBBufferDBSAMilliQtotal
2cuts10µL1µL0µL1µL0µL3µL0µL3µL12µL30µL
NC0.5µL0µL0µL0µL0µL1µL0µL1µL7.5µL10µL
9/13 PlacXbaIPstIBufferHBSAMilliQtotal
1 cut9µL0µL1µL3µL3µL14µL30µL
NC1µL0µL0µL1µL1µL7µL10µL
9/14 PKaiBCEcoRIXbaISpeIPstIBufferBBSAMilliQtotal
2cuts11µL1µL0µL1µL0µL3µL3µL11µL30µL
NC4µL0µL0µL0µL0µL1µL1µL4µL10µL
9/14 RpaBEcoRIXbaISpeIPstIBufferDBSAMilliQtotal
2cuts13µL0µL1µL0µL1µL3µL3µL9µL30µL
NC2µL0µL0µL0µL0µL1µL1µL5µL10µL
8/17 RBS-GFP-DTEcoRIXbaISpeIPstIBufferDBSAMilliQtotal
1cut9µL0µL1µL0µL0µL3µL3µL14µL30µL
NC0.5µL0µL0µL0µL0µL1µL1µL7.5µL10µL
  • incubated 37°C 1hour

Liquid Culture

Hirano

Samplemedium
9/11 Plac(BBa-R0011)-(3)Plusgrow medium(+Amp)

PCR

No name

genome DNAMilliQBig Dye5x buffertemp(400mg)primertotal
Pcon-RBS-luxR-DT0.51.751.750.40.55.110.5
Pcon-RBS-luxR-DT0.51.751.750.40.55.110.5
Pcon-RBS-tetR-DT0.51.751.7530.5310.5
Pcon-RBS-tetR-DT0.51.751.7530.5310.5
Pcon-RBS-luxR-DT0.51.751.752.40.53.610.5
Plux-RBS-GFP-DT0.51.751.751.90.54.110.5
Plux-RBS-GFP-DT0.51.750.51.90.54.110.5
Pcon-RBS-GFP-DT0.51.750.51.20.54.810.5
RBS-lysis3-DT0.51.750.51.00.55.010.5
Pcon-RBS-GFP-DT0.51.750.51.40.54.610.5
Pcon-RBS-GFP-DT0.51.750.51.40.54.610.5
RBS-lysis3-DT0.51.751.750.40.55.110.5
Ptet-RBS-spinach-DT0.51.750.50.590.50.110.5
PreDenatureDenatureAnnealingExtensioncycle
96°C96°C50°C60°C--
2min10s5s4min30cycle

Restriction Enzyme Digestion

No name

9/14 Plac(1A2)EcoRIXbaISpeIPstIBufferBBSAMilliQtotal
2cuts23µL0µL0µL0.3µL0µL3µL3µL0µL30µL
9/14 RBS-GFP-DTEcoRIXbaISpeIPstIBufferHBSAMilliQtotal
2 cuts23µL1µL0µL0µL0µL3µL3µL0µL30µL

Ligation

Nakamoto and Hirano

stateVectorInserterLigation High ver.2
experiment9/13 Pcon-PT181attenuater(SpeI&PstI)1.79/13 RBS-lacZα-DT (XbaI & PstI)12.55
experiment9/13 pSB2C3 (XbaI & PstI)1.99/14 RpaB (XbaI & PstI)16.320.1
experiment9/13 pSB2C3 (XbaI & PstI)4.79/14 PkaiBC (EcoRI & SpeI)6.711.4
experiment9/14 RBS-GFP-DT (PstI&XbaI)0.79/14 PkaiBC (EcoRI & SpeI)2.63.3
experiment9/14 Plac (SpeI&PstI)2.49/14 RBS-lysis1-DT (XbaI & PstI)4.97.3
experiment9/14 Plac (SpeI&PstI)2.49/14 RBS-lysis2-DT (XbaI & PstI)7.09.4
experiment9/14 Plac (SpeI&PstI)2.49/14 RBS-lysis3-DT (XbaI & PstI)9.211.6

Samples were evaporeted used evaporator into about 7 µL

Colony PCR

No name

Samplebase pair
9/13 Plux+RBS-lysis1-DT (1)~(3)--
9/13 Plux+RBS-lysis2-DT (1)~(3)-
9/13 Plux+RBS-lysis3-DT (1)~(2)-
Samplebase pair
9/13 Pcon-RBS-luxR-DT+Plux-RBS-GFP-DT (1)~(4)--
9/13 Pcon-PT181attenuator+aptamer121R-DT (1)~(2)-
9/13 Plac+PT181attenuator (1)-
Samplebase pair
Dλ-lux(1) (1)~(2)--
NC --

Liquid Culture

No name

Samplemedium
9/13 Pλ-luxI(1)-1Plusgrow medium(+Amp)

Transformation

No Name

NameSampleCompetent CellsTotalPlate
8/27 pT181 attenuator-DT2µL20µL22µLCP
8/27 RBS-lysis3-DT2µL20µL22µLCP
8/28 RBS-lysis3-DT2µL20µL22µLCP
8/28 Pcon-RBS-GFP-DT-Pcon-RBS-LuxR-DT2µL20µL22µLAmp
8/28 RBS-lysis1-DT2µL20µL22µLCP
8/28 Plux-RBS-GFP-DT2µL20µL22µLCP
8/28 Spinach-DT2µL20µL22µLCP

Igku Aug29 Transformation1(N④-1).jpg
Igku Aug29 Transformation(N④-2).jpg
Igku Aug29 Transformation(N⑤).jpg
File:Igku Aug29 Transformation()after


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◎)