DEVELOPMENTS OF THE METHOD FOR THE CHEMICAL SYNTHESIS OF RNA CONTAINING MODIFIED BASED" -DIRECTED TOWARD FULL AUTOMATION OF RNA CONTAINING MODIFIED BASES BY USE OF NEW SYNTHETIC UNITS-

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THE PURPOSE OF THIS PROJECT IS IS TO DEVELOPE NEW METHODS FOR THE RAPID SYNTHESIS OF RNA CONTAINING MODIFIED BASES.WE FIRST SUCCEEDED IN SYNTHESIZING THE 5'-TERMINAL OLIGORIBONUCLEOTIDES OF U1 RNA CONTAINING A UNIQUE STRUCTURE OF TRIMETHYLATED CAP AT ITS 5'-TERMINAL END.WE HAVE ALSO ESTABLISHED PHOSPHORAMIDITE BUILDING BLOCKS OF 6-THIOGUANOSINE,6-THIOINOSINE,AND 4-THIOURIDINE AS NEW PHOTOCROSSLINKING SITES.THESE MODIFIED RIBONUCLEOSIDES COULD BE INCORPORATED INTO RNA BY USING THESE SYNTHETIC UNITS.PARTICULARLY,4-THIOURIDINE 3', 5'-BISPHOSPHATE WAS CHEMICALY SYNTHESIZED AND USED FOR THE CHEMICAL SYNTHESIS OF THE 5'-TERMINAL BLOCK OF U1 RNA CONTAINING 4-THIOURIDINE AT THE 3'-TERMINAL SITE.2-AZIDODEOXYADENOSINE WAS SYNTHESIZED AS AN RIBONUCLEOSIDE MODEL AND INCORPORTED INTO DNA OLIGOMERS.THESE MODIFIED OLIGOMERS.HAVE PROVED TO HAVE A PHOTOCROSSLINKING ABILITY WITH TARGETED COMPLEMENTARY DNA STRANDS.IN ADDITION TO THESE STUDIES,WE HAVE SHOWED A NEW STRATEGY OF THE CHEMICAL SYNTHESIS OF OLIGODEOXYRIBONUCLEOTIDES WITHOUT USING BASE PROTECTING GROUPS BY THE PHOSPHONATE APPROACH USING A NEW CONDENSING REAGENT,BOMP.VERY PROMISING RESULTS WERE OBTAINED AT THIS DNA LEVEL.NO SIDE REACTIONS ON THE BASE MOITIES WERE OBSERVED UNDER THESE CONDITIONS.THEREFORE,WE ARE NOW EXTENDING THIS NEW STRATEGY TO THE CHEMICAL SYNTHESIS OF MODIFIED RNA,SINCE THIS APPROACH ENABLED US TO SYNTHESIZE OLIGODEOXYRIBONUCLEOTIDES CONTAINING 2-AZIDODEOXYADENOSINE AND N-ACETYLDEOXYCYTIDINE.