The aim of these steps is to bind nucleic acids to the columns and subsequently destroy DNA in order to keep only RNAs.
Material and reagents :
2 X NucleoSpin® RNA Plant Column (light bue ring);
2 X Collection tube 2 ml;
Ethanol 70%;
MDB Buffer;
rDNAse mixture (=rDNAse + buffer) = same stock for all groups.
Procedure :
Add 350 μL ethanol (70 %) to the homogenized lysate and mix by pipetting up and down (5 times).
For each preparation take one NucleoSpin® RNA Plant Column (light blue ring) placed in a Collection Tube and load the lysate. Centrifuge for 30 s at 11,000 x g. Place the column in a new Collection Tube (2 mL).
Add 350 μL of the MDB (Membrane Desalting Buffer) solution and centrifuge at 11,000 x g for 1 min to dry the membrane. Salt removal will make the following rDNase digest much more effective.
Apply 95 μL DNase reaction mixture directly onto the center of the silica membrane of the column.