SYNTHESIS OF OLIGODEOXYNUCLEOTIDES CONTAINING 2-THIOPYRIMIDINE RESIDUES - A NEW PROTECTION SCHEME

A method is described for the incorporation of 2'-deoxy-2-thiouridine (d(S2)U) and 2'-deoxy-2-thiothymidine (d(S2)T) into oligodeoxynucleoti des at predetermined positions. This requires N-3 orO(4)-acylation of d(S2)U and d(S2)T with toluoyl chloride. These base-protected thiopyri midines are completely stable toward the aqueous iodine oxidation reag ent used in the phosphoramidite DNA synthesis method. The toluoyl prot ecting group is removed during the standard post-synthetic ammonia tre atment. This novel protection strategy allows d(S2)U and d(S2)T to be efficiently incorporated into oligodeoxynucleotides at predetermined s ites without the usual problem of desulfurization and decomposition. S everal 14-mers containing the Eco-Rl recognition site (dGGCGGAAXXCCGCC and dGGCGGAAXXCGCGG, where X represents dT, d(S2)U or d(S2)T) have be en synthesized and characterized by base composition, thermal denatura tion, CD spectroscopy and endonuclease substrate activity.