DNA strands with appropriate sequences of dA and dT can form a stable duplex in which the two strands adopt a parallel (ps) instead of the conventional antiparallel (aps) orientation. Four 25-nt dA.dT-containing deoxyoligonucleotides (D1-4) were synthesized. D1 has the sequence 5'-dA10TA2T4A3TAT3-3'. Viewed with the same polarity, D2, D3, and D4 are the complement, inverted complement, and inverse of D1, respectively. The two combinations D1.D3 and D2.D4 form conventional antiparallel duplexes (aps-D1.D3, aps-D2.D4). D1.D2 and D3.D4, however, constitute stable parallel-stranded duplexes (ps-D1.D2, ps-D3.D4), as established by various criteria including the following: (i) The electrophoretic mobilities of ps-D1.D2 and ps-D3.D4 are similar to those of the antiparallel-stranded duplexes. (ii) The ultraviolet absorption and circular dichroism spectra of the ps duplexes are indicative of a base-paired structure, but differ systematically from those of the aps helices. (iii) Similar salt-dependent thermal transitions are observed for the four duplexes, but the melting temperatures of the ps molecules are lower by 13-18 degrees C.