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Revision as of 02:38, 5 August 2009
Lab-note on 24 July 2009
Experimenter | Kashima, Takashima, Hibino, Miyazawa
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Note-taker | Takashima
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Title
7/24 Miniprep Kit, Restriction Enzyme Digestion Electrophoresis
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Main Protocol
- Miniprep Kit
- Restriction Enzyme Digestion
- Electrophoresis
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Storage list
Sample Date | Sample Name | Enzyme | purification Kit | Product | Storage
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iGEM DNA parts kit | 3-O-2 | Eco R1 | | | 6F freezer
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iGEM DNA parts kit | 3-A-4 | Eco R1 | | | 6F freezer
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iGEM DNA parts kit | 3-O-2 | Eco R1 PST1 | | | 6F freezer
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iGEM DNA parts kit | 3-A-4 | Eco R1 PST1 | | | 6F freezer
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Miniprep
Protocol
- Cultured samples (090722 9:30 ~ 090723 16:50)
- Centrifuged 2mins at 15000g 4℃.
- Removed the top of layer.
- Centrifuged 2mins at 15000g 4℃.
- Removed the top of layer.
- Centrifuged 10mins at 14000g 4℃.
- Applied the top of layer to column.
- Centrifuged RT, 10sec and remove through.
- Added Buffer PB 500µl.
- Centrifuged RT, 10sec and remove through.
- Added Buffer PE 650µl.
- Centrifuged RT, 10sec and remove through.
- Centrifuged 1min at 17900g 4℃.
- Exchanged tube. (remove Ethanol)
- Added Milli Q 50µl RT 1min.
- Centrifuged 1min at 17900g 4℃.
- Collected through and measured concentration.
Results
Sample | conc./(ng/µl)
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3-O-2 | 283.5
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3-A-4 | 103.9
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Electrophoresis
Protocol
- Gel:Agarose
- Time:12:50-13:20
- Voltage:100 V
| 1 | 2 | 3 | 4
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Sample
| 3-O-2 (3 µl) | 3-A-4 (3 µl) | 3-O-2 (3 µl) | 3-A-4 (3 µl)
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enzyme
| Eco R1 (0.17 µl) | Eco R1 (0.17 µl) | Eco R1 (0.17 µl) | Eco R1 (0.17 µl)
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enzyme
| PST1 (0.3 µl) | PST1 (0.3 µl) | |
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Water / µl
| 1 | 1 | 1.4 | 1.4
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H buffer / µl
| 8.2 | 8.2 | 8.1 | 8.1
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Total / µl
| 12.7 | 12.7 | 12.9 | 12.9
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Incubated at 37 ℃ (090724 12:50 to 13:20 )
| 1 | 2 | 3 | 4 | 5
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Sample
| 3-O-2 | 3-A-4 | 3-O-2 | 3-A-4 | Marker
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volume / µl
| 10 | 10 | 13 | 13 | 2.5
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6 times loddingbuffer / µl
| 2 | 2 | 2.6 | 2.6 | 0.5
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Results
| 3-O-2 | 3-A-4
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Parts length / bp
| 2837 | 487
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Plasmid length / bp
| 3266 | 3266
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