03/06/13
- Colony PCR: for Rene's project
Colony PCR
- Streaked single colonies on gridded plates yesterday
- Used bit of streak for PCR today
- All parts are on pSB1A3-based vectors (http://partsregistry.org/Part:pSB1A3)
- Use primers VF2 and VR to amplify inserts
Reagent
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Volume
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Expected: Primers add ~250bp 1-3. K084007 (835 bp) = 1085 4-6. R0071 (53 bp) = 303 7. K081016 (735 bp) = 985
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10 μL/lane; 1% agarose; Ladder
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Bacteria from tip |
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10 μM VF2 primer |
1.0
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10 μM VR primer |
1.0
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2x GoTaq Green |
12.5
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dH2O |
10.5
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25.0
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PCR
- 95°C, 3 min
- [95°C, 15 sec; 57°C 15 sec; 72 °C 60 sec] x35
- 72°C, 3 min
- 4°C, ∞
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