Team:Calgary/Notebook/Protocols/Process11

From 2011.igem.org

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TITLE=Plasmid Extraction|
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TITLE=Gel Extraction|
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<a name="GE"></a>
 
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<br /><br /> <h2 style="color:#0066CC">Gel extraction</h2>
 
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<p>This protocol is utilized in accordance to the manufacturer's protocol from Omega E.Z.N.A (EaZy Nucleic Acid Isolation)</p>
<p>This protocol is utilized in accordance to the manufacturer's protocol from Omega E.Z.N.A (EaZy Nucleic Acid Isolation)</p>
<ol>
<ol>
<li> Place gel on the UV box</li>
<li> Place gel on the UV box</li>
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<li> Carefully extract the fragment suspended in the gel/li>
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<li> Carefully extract the fragment suspended in the gel </li>
<li> Mass gel fragments </li>
<li> Mass gel fragments </li>
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<li> Place fragment into a 1.5 mL tube and add 4 &micro;L of H2O </li>
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<li> Place fragment into a 1.5 mL tube and add 4 &micro;L of H<sub>2</sub>O </li>
<li> Volume of water added to volume of gel is 200% however if fragment it small 1 mL of water will suffice </li>
<li> Volume of water added to volume of gel is 200% however if fragment it small 1 mL of water will suffice </li>
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<li> Remove H2O </li>
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<li> Remove H<sub>2</sub>O</li>
<li> Add equal amounts of H2O and Binding Buffer (XP2) to the gel</li>
<li> Add equal amounts of H2O and Binding Buffer (XP2) to the gel</li>
<li> Incubate mixture at 55 degrees for 7 mins </li>
<li> Incubate mixture at 55 degrees for 7 mins </li>
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<li> Discard the liquid </li>
<li> Discard the liquid </li>
<li> Spin down the column at 13,000xg for 1 min to dry the column </li>
<li> Spin down the column at 13,000xg for 1 min to dry the column </li>
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<li> Elute in 50 &micro;L of H2O and wait 1 min </li>
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<li> Elute in 50 &micro;L of H<sub>2</sub>O and wait 1 min </li>
<li> Spin down the column at 13,000xg for 1 min to dry the column </li>
<li> Spin down the column at 13,000xg for 1 min to dry the column </li>
<li> Use a spectrophotometer to measure the concentration and the purity of your plasmid</li>
<li> Use a spectrophotometer to measure the concentration and the purity of your plasmid</li>

Latest revision as of 04:24, 29 September 2011


Gel Extraction

This protocol is utilized in accordance to the manufacturer's protocol from Omega E.Z.N.A (EaZy Nucleic Acid Isolation)

  1. Place gel on the UV box
  2. Carefully extract the fragment suspended in the gel
  3. Mass gel fragments
  4. Place fragment into a 1.5 mL tube and add 4 µL of H2O
  5. Volume of water added to volume of gel is 200% however if fragment it small 1 mL of water will suffice
  6. Remove H2O
  7. Add equal amounts of H2O and Binding Buffer (XP2) to the gel
  8. Incubate mixture at 55 degrees for 7 mins
  9. Mix with vortex for 2 mins
  10. Place in the HiBind DNA Mini Column in the 2 mL tube
  11. Add 700 µL at 10,000xg for 1 min
  12. Discard liquid
  13. Add 300 µL Binding Buffer (XP2) into the HiBind DNA Mini Column and spin down at 10,000xg for 1 min
  14. Discard liquid
  15. Wash the column with 700 µL of SPW buffer with added ethanol and spin down at 10,000xg for 1 min
  16. Discard liquid
  17. Wash the column with 700 µL of SPW buffer again and spin down at 10,000xg for 1 min
  18. Discard the liquid
  19. Spin down the column at 13,000xg for 1 min to dry the column
  20. Elute in 50 µL of H2O and wait 1 min
  21. Spin down the column at 13,000xg for 1 min to dry the column
  22. Use a spectrophotometer to measure the concentration and the purity of your plasmid

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