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Template:Kyoto/Notebook/Sep 20
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(→Electrophoresis) |
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==Sep 20== | ==Sep 20== | ||
===Ligation=== | ===Ligation=== | ||
| - | |||
| - | |||
<div class="experiment"> | <div class="experiment"> | ||
| - | <span class="author">Okazaki,Kanaoka,Tatsui | + | <span class="author">Okazaki, Kanaoka, Tatsui and Hirano</span> |
{| class="wikitable" | {| class="wikitable" | ||
!state||colspan="2"|Vector||colspan="2"|Inserter||Ligation High ver.2 | !state||colspan="2"|Vector||colspan="2"|Inserter||Ligation High ver.2 | ||
|- | |- | ||
| - | |experiment||9/18 Plac( | + | |experiment||9/18 Plac (SpeI & PstI)||3.0 µL||9/18 RBS-lysis3-DT (XbaI & PstI)||12.4 µL||3.5 µL |
|- | |- | ||
| - | |experiment||9/12 Ptet- | + | |experiment||9/12 Ptet-pT181 antisense||1.0 µL||9/18 spinach-DT||11.9 µL||3.5 µL |
|- | |- | ||
| - | |experiment||9/13 Pcon-RBS-luxR-DT||2.1µL||9/10 Plux-RBS-GFP-DT||5.6µL||3.5 µL | + | |experiment||9/13 Pcon-RBS-luxR-DT||2.1 µL||9/10 Plux-RBS-GFP-DT||5.6 µL||3.5 µL |
|- | |- | ||
| - | |experiment||9/13 Pcon- | + | |experiment||9/13 Pcon-pT181 attenuator (SpeI & PstI)||2.0 µL||9/18 aptamer 12-1R-DT (XbaI & PstI)||6.3 µL||3.5 µL |
|- | |- | ||
| - | |experiment||9/18 | + | |experiment||9/18 Plac ②(SpeI & PstI)||3.0 µL||9/18 aptamer 12-1R-DT(XbaI & PstI)||4.5µL||x3.5 µL |
|- | |- | ||
|} | |} | ||
</div> | </div> | ||
| - | |||
===Transformation=== | ===Transformation=== | ||
| Line 28: | Line 25: | ||
!Name||Sample||Competent Cells||Plate | !Name||Sample||Competent Cells||Plate | ||
|- | |- | ||
| - | |9/17 | + | |9/17 pSB1C3-Pcon-pT181 attenuator-DT||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/17 | + | |9/17 pSB1C3-Pcon-aptamer 12_1R-DT||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/17 | + | |9/17 pSB1C3-Pcon-antisense-spinach-DT||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/17 | + | |9/17 pSB1C3-Pcon-RBS-tetR-DT x2||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/14 | + | |9/14 pSB1C3-PkaiBC||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/14 | + | |9/14 pSB1C3-RpaB||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/16 | + | |9/16 pSB1C3-pT181 attenuator(EcoRI&SpeI) ||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/17 | + | |9/17 pSB1C3-pT181 attenuator(XbaI&PstI) ||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/ | + | |9/17 pSB1C3-Pcon-spinach-DT||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/14 RBS-GFP-DT-PkaiBC||2µL||20µL||CP | + | |9/14 RBS-GFP-DT-PkaiBC||2 µL||20 µL||CP |
|- | |- | ||
| - | |Plux-RBS-lysis3-DT||2µL||20µL||CP | + | |Plux-RBS-lysis3-DT||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/20 Ptet- | + | |9/20 Ptet-pT181 antisense-spinach-DT||2 µL||20 µL||CP |
|- | |- | ||
| - | |9/20 Pcon-spinach-DT( | + | |9/20 Pcon-spinach-DT(pSB4K5) ||2 µL||20 µL||Kan |
|- | |- | ||
| - | |9/20 Pcon- | + | |9/20 Pcon-pT181 antisense-spinach-DT(pSB4K5) ||2 µL||20 µL||Kan |
|- | |- | ||
| - | |9/20 KaiABC||2µL||20µL|| | + | |9/20 KaiABC||2 µL||20 µL||Kan |
|- | |- | ||
| - | |9/20 KaiABC||2µL||20µL|| | + | |9/20 KaiABC||2 µL||20 µL||Kan |
|- | |- | ||
| - | |9/20 Plac-RBS-lysis3-DT||2µL||20µL||Amp | + | |9/20 Plac-RBS-lysis3-DT||2 µL||20 µL||Amp |
|- | |- | ||
| - | |9/20 Plac- | + | |9/20 Plac-aptamer 12_1R||2 µL||20 µL||Amp |
|- | |- | ||
| - | |9/20 Plux-RBS-GFP-DT-Pcon-RBS-luxR-DT||2µL||20µL||Amp | + | |9/20 Plux-RBS-GFP-DT-Pcon-RBS-luxR-DT||2 µL||20 µL||Amp |
|- | |- | ||
| - | |Plac- | + | |Plac-aptamer12_1R-DT||2 µL||20 µL||Amp |
|- | |- | ||
| - | |9/20 Plac- | + | |9/20 Plac-aptamer12_1R-DT||2 µL||20 µL||Amp |
|- | |- | ||
| - | |9/20Pcon-pT181attenuator-aptamer-DT||2µL||20µL||Amp | + | |9/20Pcon-pT181attenuator-aptamer-DT||2 µL||20 µL||Amp |
|- | |- | ||
| - | |9/17 Plac-pT181-attenuatorx2||2µL||20µL||Amp | + | |9/17 Plac-pT181-attenuatorx2||2 µL||20 µL||Amp |
|- | |- | ||
| - | |9/15 Pcon-RBS-lacZα-DT||2µL||20µL||Amp | + | |9/15 Pcon-RBS-lacZα-DT||2 µL||20 µL||Amp |
|- | |- | ||
| - | |9/14 Pcon-pT181attenuator-RBS-lacZα-DT||2µL||20µL||Amp | + | |9/14 Pcon-pT181attenuator-RBS-lacZα-DT||2 µL||20 µL||Amp |
|- | |- | ||
| - | |PUC19||2µL||20µL||Amp | + | |PUC19||2 µL||20 µL||Amp |
| - | + | ||
| - | + | ||
|} | |} | ||
</div> | </div> | ||
===PCR=== | ===PCR=== | ||
| - | |||
<div class="experiment"> | <div class="experiment"> | ||
<span class="author">Tatsui</span> | <span class="author">Tatsui</span> | ||
| Line 91: | Line 85: | ||
!Sample||base pair | !Sample||base pair | ||
|- | |- | ||
| - | | | + | |pT181 attenuator(pSB1C3)||-- |
|- | |- | ||
|Pcon-RBS-lacZα-DT||-- | |Pcon-RBS-lacZα-DT||-- | ||
| Line 98: | Line 92: | ||
!PreDenature||Denature||Annealing||Extension||cycle | !PreDenature||Denature||Annealing||Extension||cycle | ||
|- | |- | ||
| - | |94°C||94°C||55°C||68°C||-- | + | |94 °C||94 °C||55 °C||68 °C||-- |
|- | |- | ||
| - | | | + | |5 min||30 s||30 s||3 min||30 cycles |
|} | |} | ||
</div> | </div> | ||
| Line 112: | Line 106: | ||
|1||Pcon-RBS-lacZα-DT||-- | |1||Pcon-RBS-lacZα-DT||-- | ||
|- | |- | ||
| - | |2|| 1kb ladder||-- | + | |2||1kb ladder||-- |
|- | |- | ||
| - | |3|| | + | |3||pSB1C3-pT181attenuator ||-- |
|} | |} | ||
</div> | </div> | ||
Latest revision as of 11:39, 24 September 2013
Contents |
Sep 20
Ligation
| state | Vector | Inserter | Ligation High ver.2 | ||
|---|---|---|---|---|---|
| experiment | 9/18 Plac (SpeI & PstI) | 3.0 µL | 9/18 RBS-lysis3-DT (XbaI & PstI) | 12.4 µL | 3.5 µL |
| experiment | 9/12 Ptet-pT181 antisense | 1.0 µL | 9/18 spinach-DT | 11.9 µL | 3.5 µL |
| experiment | 9/13 Pcon-RBS-luxR-DT | 2.1 µL | 9/10 Plux-RBS-GFP-DT | 5.6 µL | 3.5 µL |
| experiment | 9/13 Pcon-pT181 attenuator (SpeI & PstI) | 2.0 µL | 9/18 aptamer 12-1R-DT (XbaI & PstI) | 6.3 µL | 3.5 µL |
| experiment | 9/18 Plac ②(SpeI & PstI) | 3.0 µL | 9/18 aptamer 12-1R-DT(XbaI & PstI) | 4.5µL | x3.5 µL |
Transformation
| Name | Sample | Competent Cells | Plate |
|---|---|---|---|
| 9/17 pSB1C3-Pcon-pT181 attenuator-DT | 2 µL | 20 µL | CP |
| 9/17 pSB1C3-Pcon-aptamer 12_1R-DT | 2 µL | 20 µL | CP |
| 9/17 pSB1C3-Pcon-antisense-spinach-DT | 2 µL | 20 µL | CP |
| 9/17 pSB1C3-Pcon-RBS-tetR-DT x2 | 2 µL | 20 µL | CP |
| 9/14 pSB1C3-PkaiBC | 2 µL | 20 µL | CP |
| 9/14 pSB1C3-RpaB | 2 µL | 20 µL | CP |
| 9/16 pSB1C3-pT181 attenuator(EcoRI&SpeI) | 2 µL | 20 µL | CP |
| 9/17 pSB1C3-pT181 attenuator(XbaI&PstI) | 2 µL | 20 µL | CP |
| 9/17 pSB1C3-Pcon-spinach-DT | 2 µL | 20 µL | CP |
| 9/14 RBS-GFP-DT-PkaiBC | 2 µL | 20 µL | CP |
| Plux-RBS-lysis3-DT | 2 µL | 20 µL | CP |
| 9/20 Ptet-pT181 antisense-spinach-DT | 2 µL | 20 µL | CP |
| 9/20 Pcon-spinach-DT(pSB4K5) | 2 µL | 20 µL | Kan |
| 9/20 Pcon-pT181 antisense-spinach-DT(pSB4K5) | 2 µL | 20 µL | Kan |
| 9/20 KaiABC | 2 µL | 20 µL | Kan |
| 9/20 KaiABC | 2 µL | 20 µL | Kan |
| 9/20 Plac-RBS-lysis3-DT | 2 µL | 20 µL | Amp |
| 9/20 Plac-aptamer 12_1R | 2 µL | 20 µL | Amp |
| 9/20 Plux-RBS-GFP-DT-Pcon-RBS-luxR-DT | 2 µL | 20 µL | Amp |
| Plac-aptamer12_1R-DT | 2 µL | 20 µL | Amp |
| 9/20 Plac-aptamer12_1R-DT | 2 µL | 20 µL | Amp |
| 9/20Pcon-pT181attenuator-aptamer-DT | 2 µL | 20 µL | Amp |
| 9/17 Plac-pT181-attenuatorx2 | 2 µL | 20 µL | Amp |
| 9/15 Pcon-RBS-lacZα-DT | 2 µL | 20 µL | Amp |
| 9/14 Pcon-pT181attenuator-RBS-lacZα-DT | 2 µL | 20 µL | Amp |
| PUC19 | 2 µL | 20 µL | Amp |
PCR
| Sample | base pair |
|---|---|
| pT181 attenuator(pSB1C3) | -- |
| Pcon-RBS-lacZα-DT | -- |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94 °C | 94 °C | 55 °C | 68 °C | -- |
| 5 min | 30 s | 30 s | 3 min | 30 cycles |
Electrophoresis
| Lane | Sample | Enzyme |
|---|---|---|
| 1 | Pcon-RBS-lacZα-DT | -- |
| 2 | 1kb ladder | -- |
| 3 | pSB1C3-pT181attenuator | -- |
