Improved chemistry for oligodeoxyribonucleotide synthesis substantially improves restriction enzyme cleavage of a synthetic 35mer

摘要

Two DNA duplexes of identical sequence and 35 nt in length were synthesized by an original and a highly improved version of phosphoramidite chemistry. By base composition analysis, DNA synthesized by improved chemistry (termed DMTS-imp) contained no detectable modified bases while DNA synthesized by the original chemistry (termed DMTS-std) had a large number of modifications. Under optimal reaction conditions, HhaI and RsaI cleaved the DMTS-std duplex to 76–77% completion and the DMTS-imp duplex to 96–99% completion. Restriction analysis and piperidine treatment yielded estimates of ～ 3.0% modified nucleotides in DMTS-std and ～1.0% in DMTS-imp. Overall, the improvements in chemistry increased the restriction efficiency of synthetic DNA up to 10-fold.