Abstract
A decanucleotide dC-C-C-T-C-G-A-G-G-G was synthesized by phosphotriester block condensation. Three protected blocks, dCCCTp, dCGAp and dGGG were prepared using N-acyl, 5'-O-monomethoxytrityldeoxynucleosides as the starting materials. The protected dGGG which served as the 3'-terminal block was synthesized by condensation of 3'-O-benzoyl-N-isobutyryldeoxyguanosine (5'-hydroxy component) with 5'-O-monomethoxytrityl-N-isobutyryldeoxyguanosine 3'-O-p-chlorophenyl phosphate (3'-O-phosphodie ster component) using mesitylenesulfonyl triazolide as the condensing reagent followed by removal of the 5'-monomethoxytrityl group for repeated condensation. The other two blocks were prepared by using the 3'-O-p-chlorophenyl phosphoranilidate of N, 5'-protected deoxynucleosides as the 3'-end unit (5'-hydroxy component). The phosphoranilidate of the fully protected trimers was removed by treatment with isoamyl nitrite for the condensation with the 5'-hydroxyl group of the growing chain. Fully protected nucleotides were isolated by chromatography on silica gel and the deblocked product was purified by ionexchange chromatography on DEAE-cellulose. The decanucleotide was characterized by mobility shift analysis and complete enzymatic digestions after labelling the 5'-end with [γ-32P] ATP using polynucleotide kinase.
