LAUSR

Click chemistry has become a widely used method to insert modifications into DNA. Due to its commercial availability, 5'-ethynyl-deoxyuridine (EdU) is commonly incorporated into the DNA for subsequent modification by click reaction. However, it is partially oxidized during deprotection during solid-phase synthesis, resulting in a ketone that is no longer accessible for click modification. To enable the high-fidelity solid-phase synthesis of EdU-containing DNA, this protocol describes a procedure to perform the click reaction on the solid phase before deprotection. Afterwards, the DNA can be deprotected and purified according to standard procedures, and the full modification of EdU with the azide of choice can be analyzed by HPLC and HPLC/MS.