Team:Calgary/Notebook/Protocols/Process11
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(Difference between revisions)
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<ol> | <ol> | ||
<li> Place gel on the UV box</li> | <li> Place gel on the UV box</li> | ||
- | <li> Carefully extract the fragment suspended in the gel/li> | + | <li> Carefully extract the fragment suspended in the gel </li> |
<li> Mass gel fragments </li> | <li> Mass gel fragments </li> | ||
<li> Place fragment into a 1.5 mL tube and add 4 µL of H2O </li> | <li> Place fragment into a 1.5 mL tube and add 4 µL of H2O </li> |
Revision as of 22:37, 28 September 2011
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Plasmid Extraction
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Gel extraction
This protocol is utilized in accordance to the manufacturer's protocol from Omega E.Z.N.A (EaZy Nucleic Acid Isolation)
- Place gel on the UV box
- Carefully extract the fragment suspended in the gel
- Mass gel fragments
- Place fragment into a 1.5 mL tube and add 4 µL of H2O
- Volume of water added to volume of gel is 200% however if fragment it small 1 mL of water will suffice
- Remove H2O
- Add equal amounts of H2O and Binding Buffer (XP2) to the gel
- Incubate mixture at 55 degrees for 7 mins
- Mix with vortex for 2 mins
- Place in the HiBind DNA Mini Column in the 2 mL tube
- Add 700 µL at 10,000xg for 1 min
- Discard liquid
- Add 300 µL Binding Buffer (XP2) into the HiBind DNA Mini Column and spin down at 10,000xg for 1 min
- Discard liquid
- Wash the column with 700 µL of SPW buffer with added ethanol and spin down at 10,000xg for 1 min
- Discard liquid
- Wash the column with 700 µL of SPW buffer again and spin down at 10,000xg for 1 min
- Discard the liquid
- Spin down the column at 13,000xg for 1 min to dry the column
- Elute in 50 µL of H2O and wait 1 min
- Spin down the column at 13,000xg for 1 min to dry the column
- Use a spectrophotometer to measure the concentration and the purity of your plasmid
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