CHEMICAL SYNTHESIS AND CHARACTERIZATION OF BRANCHED OLIGODEOXYRIBONUCLEOTIDES (BDNA) FOR USE AS SIGNAL AMPLIFIERS IN NUCLEIC-ACID QUANTIFICATION ASSAYS

The divergent synthesis of bDNA structures is described. This new type of branched DNA contains one unique oligonucleotide, the primary sequ ence, covalently attached through a comb-like branching network to man y identical copies of a different oligonucleotide, the secondary seque nce. The bDNA comb molecules were assembled on a solid support using p arameters optimized for bDNA synthesis. The chemistry was used to synt hesize bDNA comb molecules containing 15 secondary sequences. The bDNA comb molecules were elaborated by enzymatic ligation into branched am plification multimers, large bDNA molecules (a total of 1068 nt) conta ining an average of 36 repeated DNA oligomer sequences, each capable o f hybridizing specifically to an alkaline phosphatase-labeled oligonuc leotide. The bDNA comb molecules were characterized by electrophoretic methods and by controlled cleavage at periodate-cleavable moieties in corporated during synthesis. The branched amplification multimers have been used as signal amplifiers in nucleic acid quantification assays for detection of viral infection. It is possible to detect as few as 5 0 molecules with bDNA technology.