The synthesis and DNA binding properties of bis-PNA (peptide nucleic acid) are reported. Two PNA segments each of seven nucleobases in length were connected in a continuous synthesis via a flexible linker composed of three 8-amino-3,6-dioxaoctanoic acid units. The sequence of the first strand was TCTCTTT (C- to N-terminal), while the second strand was TTTCTCT or TTTJTJT, where J is pseudoisocytosine. These bis-PNAs form triple-stranded complexes of somewhat higher thermal stability than monomeric PNA with complementary oligonucleotides and the thermal melting transition shows very little hysteresis. When the J base is placed in the strand parallel to the DNA complement ('Hoogsteen strand'), the DNA binding was pH independent. The bis-PNAs were also superior to monomeric PNAs for targeting double-stranded DNA by strand invasion.