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Template:Kyoto/Notebook/Sep 11
From 2013.igem.org
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!pKaiBC(200p)||10*buffer KOD plus||dNTPs||MgSO4||primer mix(F&R)||KOD plus||MilliQ||total | !pKaiBC(200p)||10*buffer KOD plus||dNTPs||MgSO4||primer mix(F&R)||KOD plus||MilliQ||total | ||
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| - | |0.5||2.5||2.5||1.5||1.5||0.5||16 | + | |0.5||2.5||2.5||1.5||1.5||0.5||16||25 |
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!pKaiBC(400p)||10*buffer KOD plus||dNTPs||MgSO4||primer mix(F&R)||KOD plus||MilliQ||total | !pKaiBC(400p)||10*buffer KOD plus||dNTPs||MgSO4||primer mix(F&R)||KOD plus||MilliQ||total | ||
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| - | |1||2.5||2.5||1.5||1.5||0.5||15.5 | + | |1||2.5||2.5||1.5||1.5||0.5||15.5||25 |
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!pKaiBC(800p)||10*buffer KOD plus||dNTPs||MgSO4||primer mix(F&R)||KOD plus||MilliQ||total | !pKaiBC(800p)||10*buffer KOD plus||dNTPs||MgSO4||primer mix(F&R)||KOD plus||MilliQ||total | ||
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| - | |2||2.5||2.5||1.5||1.5||0.5||14.5 | + | |2||2.5||2.5||1.5||1.5||0.5||14.5||25 |
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!pKaiBC(1ng)||10*buffer KOD plus||dNTPs||MgSO4||primer mix(F&R)||KOD plus||MilliQ||total | !pKaiBC(1ng)||10*buffer KOD plus||dNTPs||MgSO4||primer mix(F&R)||KOD plus||MilliQ||total | ||
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| - | |2.5||2.5||2.5||1.5||1.5||0.5||14 | + | |2.5||2.5||2.5||1.5||1.5||0.5||14||25 |
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===colony PCR=== | ===colony PCR=== | ||
<div class="experiment"> | <div class="experiment"> | ||
Revision as of 12:34, 24 September 2013
Contents |
Sep 11
PCR
| pKaiBC(200p) | 10*buffer KOD plus | dNTPs | MgSO4 | primer mix(F&R) | KOD plus | MilliQ | total |
|---|---|---|---|---|---|---|---|
| 0.5 | 2.5 | 2.5 | 1.5 | 1.5 | 0.5 | 16 | 25 |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 98°C | 60°C | 68°C | 30 |
| 2min | 10sec | 30sec | 30sec |
| pKaiBC(400p) | 10*buffer KOD plus | dNTPs | MgSO4 | primer mix(F&R) | KOD plus | MilliQ | total |
|---|---|---|---|---|---|---|---|
| 1 | 2.5 | 2.5 | 1.5 | 1.5 | 0.5 | 15.5 | 25 |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 98°C | 60°C | 68°C | 30 |
| 2min | 10sec | 30sec | 30sec |
| pKaiBC(800p) | 10*buffer KOD plus | dNTPs | MgSO4 | primer mix(F&R) | KOD plus | MilliQ | total |
|---|---|---|---|---|---|---|---|
| 2 | 2.5 | 2.5 | 1.5 | 1.5 | 0.5 | 14.5 | 25 |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 98°C | 60°C | 68°C | 30 |
| 2min | 10sec | 30sec | 30sec |
| pKaiBC(1ng) | 10*buffer KOD plus | dNTPs | MgSO4 | primer mix(F&R) | KOD plus | MilliQ | total |
|---|---|---|---|---|---|---|---|
| 2.5 | 2.5 | 2.5 | 1.5 | 1.5 | 0.5 | 14 | 25 |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 98°C | 60°C | 68°C | 30 |
| 2min | 10sec | 30sec | 30sec |
colony PCR
| Sample | base pair |
|---|---|
| 9/9Pcon-lacZα(p3B4k5) | 712bp |
| 9/9Pcon-spinach-DT(p3B4k5) | 605bp |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 94°C | 55°C | 68°C | -- |
| 5min | 30s | 30s | 42s |
| Sample | base pair |
|---|---|
| Pbad\araC-RBS-luxI-DT |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 94°C | 55°C | 68°C | -- |
| 5min | 30s | 30s | 2min |
| Sample | base pair |
|---|---|
| 9/9Pcon+RBS-luxI-DT | 1079bp |
| 9/10Pcon+sptamer12_1R-DT | 584bp |
| 9/10DT+Pcon-pT181 attenuater 1 | 781bp |
| 9/10Fusion1 attenuater(p3B1C3)-1 | 638bp |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 94°C | 55°C | 68°C | -- |
| 50sec | 30sec | 30sec | 1min |
Restriction Enzyme Digestion
| DNA | E | X | S | P | buffer | BSA | MilliQ | Total | ||
|---|---|---|---|---|---|---|---|---|---|---|
| （°°） µL | µL | µL | µL | µL | µL | µL | µL | µL | ||
| NC | µL | µL | µL | µL | µL | µL | µL | µL | µL |
