Editing In vitro capped mRNA (section)

== RNA Purification with MegaClear Kit ==

1. Add 700 μl of Binding Solution Concentrate to the poly(A) tailing reaction (200 μl) and mix thoroughly by vortex.

2. Add 500 μl of 100% ethanol and mix thoroughly by vortex.

3. Insert a Filter Cartridge into a Collection Tube and add 700 μl of the RNA sample into the Filter Cartridge

4. Centrifuge at ~16,000 x g for 30 seconds. Decant the flow-through liquid and return the Filter Cartridge to the Collection Tube.<br> Note: Centrifugation in this and all subsequent steps is performed at room temperature.

5. Add the remaining RNA sample into the same Filter Cartridge and centrifuge at ~16,000 x g for 30 seconds. Decant the flow-through liquid and return the Filter Cartridge to the Collection Tube.

6. Add 500 μl of alcohol diluted Wash Solution to each Filter Cartridge and centrifuge at ~16,000 x g for 30 seconds. Decant the flow-through <br>liquid and return the Filter Cartridge to the Collection Tube.

7. Add another 500 μl of alcohol diluted Wash Solution to each Filter Cartridge and centrifuge at ~16,000 x g for 30<br>seconds. Decant the flow-through liquid and return the Filter Cartridge to the Collection Tube.

8. Dry the Filter Cartridge by centrifugation at ~16,000 x g for 1 minute.

9. Transfer the Filter Cartridge into a new Collection Tube. 

Add 50 μl of Elution Solution directly onto the center of the filter inside the Filter Cartridge. Close the cap of the tube<br>immediately.

10. Heat the tube at 65 °C for 10 minutes on a clean heat block or in a clean water bath.<br>11. Centrifuge the tube at ~16,000 x g for 1 minute to elute RNA.