In this technical note, we have outlined a protocol for the chemical synthesis, deprotection, and purification of diphosphorylated RNA oligonucleotides. This method provides a straightforward option for direct chemical synthesis with the need for enzymatic intermediate steps. This procedure has been developed for the MerMade 12 Oligonucleotide synthesizer, but can be adapted for use with alternative machines. This method can be used for chemical synthesis of most 5'diphosphate RNA oligonucleotides, including those with modifications. As tributylammonium monophosphate is less reactive than tributylammonium pyrophosphate (used for triphosphorylation), the yield of diphosphorylated oligo may be lower. A minimal amount of monophosphate is usually seen on mass spec of triphosphorylated SLR14, while 10-20% is often seen with diphosphorylated SLR14.