Cross-Linked DNA: Site-Selective “Click” Ligation in Duplexes with Bis-Azides and Stability Changes Caused by Internal Cross-Links XiongHai SeelaFrank 2016 Heterodimeric interstrand cross-linked DNA was constructed by the “bis-click” reaction carried out on preformed oligonucleotide duplexes with the bis-azide <b>1</b>. For this, alkynylated 8-aza-7-deazapurine or corresponding 5-substituted pyrimidine nucleosides were synthesized. Cross-linking resulted in chemoselective formation of heterodimeric duplexes while homodimers were suppressed. For product identification, heterodimeric DNA was prepared by the “stepwise click” reaction, while noncomplementary homodimers were accessible by “bis-click” chemistry, unequivocally. Studies on duplex melting of complementary cross-linked duplexes (heterodimers) revealed significantly increased <i>T</i><sub>m</sub> values compared to the non-cross-linked congeners. The stability of this cross-linked DNA depends on the linker length and the site of modification. Cross-linked homodimers hybridized with single-stranded complementary oligonucleotides show much lower stability.