Tripler-forming oligonucleotides (TFOs) bind in the major groove to specific double-helical DNA sequences and have been shown to inhibit the function of targeted genes. Diaziridinylquinones are bifunctional alkylating agents that can form interstrand cross-links in DNA at 5＇-GNC sites. To demonstrate the feasibility of targeted interstrand cross-linking, a diaziridinylquinone-TFO conjugate was prepared from a diaziridinylquinone intermediate bearing an activated ester linker. The first demonstration of tripler-directed interstrand DNA cross-linking by a single targeted bifunctional alkylating agent is described. Up to 38% interstrand cross-linking was observed at pH 6.2.