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Team:UNIPV-Pavia/Protocols/Precipitation
From 2008.igem.org
(Difference between revisions)
| Line 37: | Line 37: | ||
*'''absolute ethanol''' | *'''absolute ethanol''' | ||
*'''ethanol 70%''' | *'''ethanol 70%''' | ||
| + | *'''ddH2O''' | ||
*'''previously cut plasmid''' | *'''previously cut plasmid''' | ||
<br> | <br> | ||
| Line 43: | Line 44: | ||
*Freeze at -80°C for 30 min | *Freeze at -80°C for 30 min | ||
*Centrifuge at 13000 rpm, 4°C for 20 min | *Centrifuge at 13000 rpm, 4°C for 20 min | ||
| - | * | + | *Decant supernatant |
*Add 50 µl of 70% ethanol | *Add 50 µl of 70% ethanol | ||
*Centrifuge at 13000 rpm, 4°C for 20 min | *Centrifuge at 13000 rpm, 4°C for 20 min | ||
| - | * | + | *Remove all supernatant with a pipette |
| + | *Air dry pellet until ethanol is totally removed | ||
| + | *Elute with 5 µl ddH2O | ||
<br> | <br> | ||
Latest revision as of 13:30, 1 July 2008
 Home
|  The Team
|  The Project
|  Biological Safety
|  Parts Submitted to the Registry
|
|---|---|---|---|---|
 Dry Lab
|  Wet Lab
|  Modeling
|  Protocols
|  Activity Notebook
|
The protocols we used
- LB medium preparation
- Plasmid resuspension from IGEM paper spots
- Transformation
- Plasmid extraction
- BioBrick digestion with restriction enzymes
- DNA gel extraction
- DNA precipitation with sodium acetate
- Antarctic Phosphatase
- Ligation
- PCR
DNA precipitation with sodium acetate
(estimated time: 1 hour)
Materials needed:
- sodium acetate
- absolute ethanol
- ethanol 70%
- ddH2O
- previously cut plasmid
- Add 1/10 digestion volume of sodium acetate 3 M
- Add 2.5 digestion volume of absolute ethanol
- Freeze at -80°C for 30 min
- Centrifuge at 13000 rpm, 4°C for 20 min
- Decant supernatant
- Add 50 µl of 70% ethanol
- Centrifuge at 13000 rpm, 4°C for 20 min
- Remove all supernatant with a pipette
- Air dry pellet until ethanol is totally removed
- Elute with 5 µl ddH2O
