Substitution of a borano (BH3-) group for nonbridging oxygen in the phosphate backbone of DNA results in a new class of isoelectronic and isoionic DNA analogues. An effective chemical method of synthesis of oligodeoxynucleoside boranophosphates (BH3--ODNs) on a solid phase has been developed via an H-phosphonate chain elongation approach followed by boronation. The boronation procedure involves the intermediate conversion of an H-phosphonate to a phosphite triester group by silylation and subsequent oxidation by a borane-amine complex. The efficiency of the boronation procedure to form BH3--ODNs is close to that of iodine oxidation to form phosphodiester ODNs. Oligothymidine boranophosphates of different lengths up to 12-mer have been readily synthesized, purified by HPLC and/or PAGE methods, and characterized by NMR spectroscopy and MS spectrometry. In physiologically relevant buffers the dodecathymidine boranophosphate hybridized with complementary dodecadeoxyadenylate and exhibited a cooperative melting transition (T-m = 14 degrees C). Studies of substrate properties showed that BH3--ODNs are readily 5＇-phosphorylated by T4 polynucleotide kinase. Boranophosphate analogues are much more resistant toward nuclease hydrolysis than phosphodiester ODNs, and more resistant to P-1 and S-1 nucleases and snake venom phosphodiesterase than phosphorothioate ODNs.