Editing Plasmid MaxiPrep PlasmidSelect (section)

== <br>Remove Endotoxin and open-circle plasmid DNA from supercoiled plasmid using PlasmidSelect Xtra<br>  ==

The binding capacity of PlasmidSelect Xtra resin is 1 mg DNA per ml resin. 

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Lysate should be prepared using P1 + RNase, P2, P3;<br> 

Plasmid DNA should be purified using an anion-exchange column.<br> 

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1. Mix DNA in TE with&nbsp; 3.0M (NH<sub>4</sub>)<sub>2</sub>SO<sub>4</sub>, 100 mM Tris, 10 mM EDTA, pH7.0 to adjust to&nbsp; '''2.1 M''' (NH<sub>4</sub>)<sub>2</sub>SO<sub>4</sub>.<br> 

2. Load to column packed with PlasmidSelect Xtra resin, prequilibrated with '''2.0 M''' (NH<sub>4</sub>)<sub>2</sub>SO<sub>4</sub>, 100 mM Tris, and 10 mM EDTA, pH 7.5. 

3. Wash column with 2.0M (NH<sub>4</sub>)<sub>2</sub>SO<sub>4</sub>, 100 mM Tris, and 10 mM EDTA, pH 7.5. 

4. Elute supercoiled plasmid (SC pDNA) with '''1.7 M''' (NH<sub>4</sub>)<sub>2</sub>SO<sub>4</sub>, '''0.3 M NaCl,''' 100 mM Tris, 10 mM EDTA (pH7.5).<br> 

5. Clean column with water.<br> 

Endotoxin level should be less than 5 EU/mg.<br><br><br> 

[[Category:Protocols]]