Phosphodiester Functions Research Articles

61] Chemical synthesis of deoxyoligonucleotides by the modified triester method

Publisher Summary This chapter discusses the techniques for the chemical synthesis of deoxyoligonucleotides by the modified triester method. The basic principle of the triester method is to mask each internucleotidic phosphodiester function by a suitable protecting group during the course of the building sequence. As uncharged molecules, the phosphotriester intermediates are soluble in organic solvents and amenable to such conventional purification techniques of organic chemistry as silica-gel column chromatography. After building a desired sequence, all the protecting groups can be removed at the final step to give a deoxyoligonucleotide containing natural 3'→ 5'-phosphodiester bonds. The main advantages of this method are (1) opportunity for large-scale synthesis, (2) short time periods in the purification steps, and (3) high yields using stoichiometric amounts of the reactants. This is due to the absence of any endo-P-O' groups in the oligonucleotide chain thus avoiding chain scission and pyrophosphate formation. As any intermediate oligonucleotide so synthesized always contained a masked 3'-phosphate group, the original necessity of a phosphorylation step at each condensation stage was eliminated, thus simplifying the approach.