Synthesis of comb-shaped DNA using a non-nucleosidic branching phosphoramidite

Branched DNAs (bDNAs) having comb-like structures have found wide utility in molecular diagnostics and DNA nanotechnology. bDNAs can be generated either by designing and assembling linear DNA molecules into rigid non-covalent structures or by using an orthogonally protected branching unit to synthesize covalently linked structures. Despite the advantages of the covalently linked structures, use of this motif has been hampered by the challenging synthesis of appropriately protected branching monomers. We report the facile synthesis of a branching monomer having orthogonal DMT and Lev protecting groups using readily available δ-velarolactone and 1,3-diaminopropan-2-ol. Using this branching monomer, a comb-shaped bDNA was synthesized having three different DNA arms. The synthesis and hybridization capability of the bDNA was assessed by fluorescence microscopy using fluorescently labeled complementary and mismatched DNA probes. Convenient access to an orthogonally protected branching monomer is anticipated to accelerate applications of bDNAs in applications including diagnostics, biosensing, gene-profiling, DNA computing, multicolor imaging, and nanotechnology.