Home Why is it the eluted plasmid cannot be cut by restriction enzymes properly?

It Is Possible That Salt Residue In Buffers Or Ethanol Residue In WS Buffer Is Not Removed Completely And Thus Affects The Downstream Reaction. In Case Of Salt Residue, Wash The Column Twice With WS Buffer. In Case Of Ethanol Residue, After Washing With WS Buffer, Make Sure That The Flow-Through Is Discarded And Centrifuge The Column At Full Speed For 3 Minutes. If Necessary, To Centrifuge For A Few Minutes More To Ensure Complete Removal Of Ethanol.

Another Reason Is That Plasmid Is Denatured. Denaturation Happens If Incubation In MX2 Buffer Has Gone For Too Long Time. This Can Be Visualized During Electrophoresis That A Band Migrates Faster Than The Supercoiled Form. After MX2 Buffer Is Added, Do NOT Incubate For More Than 5 Minutes.