Template:Kyoto/Notebook/Sep 17

From 2013.igem.org

(Difference between revisions)
(→Electrophoresis)
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===Ligation===
===Ligation===
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<div class="experiment">
<div class="experiment">
<span class="author">No Name</span>
<span class="author">No Name</span>
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!state||colspan="2"|Vector||colspan="2"|Inserter||Ligation High ver.2
!state||colspan="2"|Vector||colspan="2"|Inserter||Ligation High ver.2
|-
|-
-
|experiment||8/28 Plux-(SpeI&PstI)||5.0&micro;g||9/13 RBS-lysis1-DT-(XbaI&PstI)||3.5&micro;g||3.5&micro;g
+
|experiment||8/28 Plux (SpeI & PstI)||5.0 &micro;L||9/13 RBS-lysis1-DT (XbaI & PstI)||3.5 &micro;L||3.5 &micro;L
|-
|-
-
|experiment||8/28 Plux-(SpeI&PstI)||5.0micro;g||9/13 RBS-lysis2-DT-(XbaI&PstI)||3.6&micro;g||3.5&micro;g
+
|experiment||8/28 Plux (SpeI & PstI)||5.0 &micro;L||9/13 RBS-lysis2-DT (XbaI & PstI)||3.6 &micro;L||3.5 &micro;L
|-
|-
-
|experiment||8/28 Plux-(SpeI&PstI)||11.0&micro;g||9/13 RBS-lysis3-DT-(XbaI&PstI)||9.5&micro;g||3.5&micro;g
+
|experiment||8/28 Plux (SpeI & PstI)||11.0 &micro;L||9/13 RBS-lysis3-DT (XbaI & PstI)||9.5 &micro;L||3.5 &micro;L
|-
|-
-
|experiment||9/14 Plac-(SpeI&PstI)||2.4&micro;g||9/16 PT181-attenuator-DT-(XbaI&PstI)||4.8&micro;g||3.5&micro;g
+
|experiment||9/14 Plac (SpeI & PstI)||2.4 &micro;L||9/16 pT181 attenuator-DT (XbaI & PstI)||4.8 &micro;L||3.5 &micro;L
|-
|-
-
|experiment||9/14 Plac-(SpeI&PstI)||2.4&micro;g||9/3 PT181-antisense-(XbaI&PstI)||2.0&micro;g||2.2&micro;g
+
|experiment||9/14 Plac (SpeI & PstI)||2.4 &micro;L||9/3 pT181 antisense (XbaI & PstI)||2.0 &micro;L||2.2 &micro;L
|-
|-
-
|experiment||9/14 Plac-(SpeI&PstI)||2.4&micro;g||9/10 aptamer12-1R-DT-(XbaI&PstI)||1.8&micro;g||2.1&micro;g
+
|experiment||9/14 Plac (SpeI & PstI)||2.4 &micro;L||9/10 aptamer 12_1R-DT (XbaI & PstI)||1.8 &micro;L||2.1 &micro;L
|-
|-
-
|experiment||9/13 pSB1C3-(XbaI&PstI)||2.0&micro;g||9/15 Pcon-PT181 antisense-spinach-DT-(XbaI&PstI)||12.0&micro;g||3.5&micro;g
+
|experiment||9/13 pSB1C3 (XbaI & PstI)||2.0 &micro;L||9/15 Pcon-PT181 antisense-Spinach-DT (XbaI & PstI)||12.0 &micro;L||3.5 &micro;L
|-
|-
-
|experiment||9/16 pSB1C3-(EcoRI&SpeI)||1.0&micro;g||9/15 Pcon-spinach-DT-(EcoRI&SpeI)||4.1&micro;g||2.0&micro;g
+
|experiment||9/16 pSB1C3 (EcoRI & SpeI)||1.0 &micro;L||9/15 Pcon-Spinach-DT (EcoRI & SpeI)||4.1 &micro;L||2.0 &micro;L
|-
|-
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|experiment||9/16 pSB1C3-(EcoRI&SpeI)||1.0&micro;g||9/15 Pcon-RBS-tetR-DT-(EcoRI&SpeI)||2.0&micro;g||1.5&micro;g
+
|experiment||9/16 pSB1C3 (EcoRI & SpeI)||1.0 &micro;L||9/15 Pcon-RBS-tetR-DT (EcoRI & SpeI)||2.0 &micro;L||1.5 &micro;L
|-
|-
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|experiment||9/16 pSB1C3-(EcoRI&SpeI)||1.0&micro;g||9/15 Pcon-PT181-attenuator-DT-(EcoRI&SpeI)||2.4&micro;g||1.7&micro;g
+
|experiment||9/16 pSB1C3 (EcoRI & SpeI)||1.0 &micro;L||9/15 Pcon-PT181 attenuator-DT (EcoRI & SpeI)||2.4 &micro;L||1.7 &micro;L
|-
|-
-
|experiment||9/16 pSB1C3-(EcoRI&SpeI)||1.0&micro;g||9/15 Pcon-aptamer12-112-DT-(EcoRI&SpeI)||17.0&micro;g||1.5&micro;g
+
|experiment||9/16 pSB1C3 (EcoRI & SpeI)||1.0 &micro;L||9/15 Pcon-aptamer 12_1R-DT (EcoRI & SpeI)||17.0 &micro;L||1.5 &micro;L
|-
|-
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|experiment||9/16 pSB1C3-(XbaI&PstI)||1.0&micro;g||9/15 Pcon-aptamer12-112-DT-(EcoRI&SpeI)||4.4&micro;g||3.5&micro;g
+
|experiment||9/16 pSB1C3 (XbaI & PstI)||1.0 &micro;L||9/15 Pcon-aptamer 12_1R-DT (EcoRI & SpeI)||4.4 &micro;L||3.5 &micro;L
|}
|}
</div>
</div>
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===Gel Extraction===
===Gel Extraction===

Revision as of 13:46, 24 September 2013

Contents

Sep 17

Liquid Culture

Tatsui and Hirano

Samplemedium
Spinach-DTLB medium(+CP)
Pcon-Spinach-DTLB medium(+Amp)　
Pcon-pT181 antisense-Spinach-DTLB medium(+Amp)　

PCR

Hirano

Pcon-tetR aptamer-DT(VF)BigDye5x bufferprimer(VF)MilliQtotal
1.30.51.750.50.4510.5
Pcon-tet ap-DT(VR)BigDye5x bufferprimer(VR)MilliQtotal
1.30.51.750.50.4510.5
PreDenatureDenatureAnnealingExtensioncycle
96 °C96 °C50 °C60 °C--
2 min10 sec5 sec4 min30 cycles

Electrophoresis

No name

LaneSampleEnzyme1Enzyme2
11kbp ladder----
2Pcon-RBS-tetR-DTXbaIPstI
3Pcon-RBS-tetR-DT----
4pSB4K5XbaIPstI
5pSB4K5----
6pSB1C3EcoRISpeI
7pSB1C3EcoRIPstI
8pSB1C3----
9pT181 attenuatorEcoRISpeI
10pT181 attenuator----
111kbp ladder----

File:Igku xxxxxx.xxx

Ligation

No Name

stateVectorInserterLigation High ver.2
experiment8/28 Plux (SpeI & PstI)5.0 µL9/13 RBS-lysis1-DT (XbaI & PstI)3.5 µL3.5 µL
experiment8/28 Plux (SpeI & PstI)5.0 µL9/13 RBS-lysis2-DT (XbaI & PstI)3.6 µL3.5 µL
experiment8/28 Plux (SpeI & PstI)11.0 µL9/13 RBS-lysis3-DT (XbaI & PstI)9.5 µL3.5 µL
experiment9/14 Plac (SpeI & PstI)2.4 µL9/16 pT181 attenuator-DT (XbaI & PstI)4.8 µL3.5 µL
experiment9/14 Plac (SpeI & PstI)2.4 µL9/3 pT181 antisense (XbaI & PstI)2.0 µL2.2 µL
experiment9/14 Plac (SpeI & PstI)2.4 µL9/10 aptamer 12_1R-DT (XbaI & PstI)1.8 µL2.1 µL
experiment9/13 pSB1C3 (XbaI & PstI)2.0 µL9/15 Pcon-PT181 antisense-Spinach-DT (XbaI & PstI)12.0 µL3.5 µL
experiment9/16 pSB1C3 (EcoRI & SpeI)1.0 µL9/15 Pcon-Spinach-DT (EcoRI & SpeI)4.1 µL2.0 µL
experiment9/16 pSB1C3 (EcoRI & SpeI)1.0 µL9/15 Pcon-RBS-tetR-DT (EcoRI & SpeI)2.0 µL1.5 µL
experiment9/16 pSB1C3 (EcoRI & SpeI)1.0 µL9/15 Pcon-PT181 attenuator-DT (EcoRI & SpeI)2.4 µL1.7 µL
experiment9/16 pSB1C3 (EcoRI & SpeI)1.0 µL9/15 Pcon-aptamer 12_1R-DT (EcoRI & SpeI)17.0 µL1.5 µL
experiment9/16 pSB1C3 (XbaI & PstI)1.0 µL9/15 Pcon-aptamer 12_1R-DT (EcoRI & SpeI)4.4 µL3.5 µL

Gel Extraction

No Name

LaneDNAEnzyme
11kbpladder--
2Pt181 attenuator -(2)EcoRI+SpeI
3
4

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

Nameconcentration[µg/mL]260/280260/230
PT181-attenuator EcoI+SpeI9.21.150.12
LaneDNAEnzyme
11kbpladder--
2pSB1C3EcoI+SpeI
3
4

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

Nameconcentration[µg/mL]260/280260/230
pSB1C310.01.840.40
LaneDNAEnzyme
11kbpladder--
2pSB4K5XbaI+PstI
3
4

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

Nameconcentration[µg/mL]260/280260/230
pSB4K57.61.810.32
LaneDNAEnzyme
11kbpladder--
2Pcon-RBS-tetR-DTXbaI+PstI
3
4

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

Nameconcentration[µg/mL]260/280260/230
Pcon-RBS-tetR-DT8.02.401.14
LaneDNAEnzyme
11kbpladder--
2pSB1C3EcoRI+PstI
3
4

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

Nameconcentration[µg/mL]260/280260/230
pSB1C39.12.170.35

Mutation PCR

Kinose

pC KaiABC(100pg)Prime STAR(2x)Primer(KaiA-Bsai MuT-Fwd)Primer(KaiA-Bsai MuT-Rev)MilliQtotal
1.025.01.01.022.050.0
1.525.01.01.021.550.0
2.025.01.01.021.050.0
PreDenatureDenatureAnnealingExtensioncycle
96°C98°C65°C72°C--
5m10s15s40s30cycles
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