2'-endo locked or frozen (S-type)/3'-endo locked or frozen (N-type) nucleoside analogues were synthesized. Conformational analysis based on (3)J(HH) and NOE measurements is presented which is further confirmed by X-ray crystal structural studies. 2'-5' isoDNA oligonucleotides (ON) were synthesized using these modified nucleoside analogues and UV-T-m studies of the resultant 2'-5' isoDNA : RNA duplexes reflect the site-and sequence-dependent effects and confirm that the S-type sugar conformations were preferred over the N-type sugar geometry in such duplexes.