SYNTHESIS AND PROPERTIES OF COVALENTLY CL OSED DNA DUPLEXES CONTAINING PYROPHOSPHATE AND SUBSTITUTED PYROPHOSPHATE INTERNUCLEOTIDE GROUPS

A new method for the efficient synthesis of covalently closed DNA dupl exes (DNA dumbbells) and the introduction of pyrophosphate and substit uted pyrophosphate internucleotide groups into their structure is prop osed. The method is based on chemical ligation in DNA duplexes that ar e formed by a polynucleotide the ends of which are brought together du e to the introduction of the minihairpin structure [GRAPHICS] DNA dumb bells containing a pyrophosphate (substituted pyrophosphate) group res ult from the interaction as being between the 3'-terminal phosphate (m ethylphosphate) group of the polynucleotide and the 5'-terminal phosph ate group of deoxyguanosine of the minihairpin sequence, which flanks the polynucleotide from the 5' end. 1-Ethyl-3-(3'-dimethylaminopropyl) carbodiimide was used as a condensing agent. The yield of covalently c losed 42-mer DNA duplexes containing a pyrophosphate group was 98%, th at of duplexes with a substituted pyrophosphate group was 25%. The rea ctivity of the substituted pyrophosphate group incorporated into DNA d umbbells was studied. It is shown that the group efficiently interacts with nucleophiles in an aqueous medium at pH 8.0.