A method for solid-phase synthesis of stereodefined PS-oligos via an oxathiaphospholane approach using pure P-diastereomers of nucleoside oxathiaphospholane monomers is described. The oxathiaphospholane monomers are synthesized by phosphitylation of 5'-O-DMTr-N-protected deoxyribonucleosides with 2-chloro-spiro-4,4-pentamethylene-1,3,2-oxathiaphospholane followed by sulfurization. The procedure is general and may be applied to other analogs, depending on the aldehyde (or mercaptoalcohol) used. Starting from an 18O-labeled mercaptoalcohol, the corresponding 18O-labeled phosphitylating reagent and nucleoside monomers can be obtained and used for synthesis of labeled stereodefined PS-oligos, which are useful for studying mechanisms of enzymatic reactions. Details are provided for chromatographic separation of the 5'-O-DMTr-N-protected-deoxyribonucleoside-3'-O-(2-thio-spiro-4,4-pentamethylene-1,3,2-oxathiaphospholane)s into their P-diastereomers, and for manual solid-phase synthesis of PS-oligos. Oxidation of 5'-O-DMTr-N-protected-deoxyribonucleoside-3'-O-(2-thio-spiro-4,4-pentamethylene-1,3,2-oxathiaphospholane)s with selenium dioxide yields their 2-oxo-analogs, which are suitable either for elongation of stereodefined PS-oligos with segments consisting of unmodified nucleotide units possessing phosphate internucleotide linkages, or for generating isotopomeric 18O-labeled PO-oligos of predetermined P-chirality.