Synthesis and structural characterization of stable branched DNA g-quadruplexes using the trebler phosphoramidite

Guanine (G)-rich sequences can form a noncanonical four-stranded structure known as the G-quadruplex. G-quadruplex structures are interesting because of their potential biological properties and use in nanosciences. Here, we describe a method to prepare highly stable G-quadruplexes by linking four G-rich DNA strands to form a monomolecular G-quadruplex. In this method, one strand is synthesized first, and then a trebler molecule is added to simultaneously assemble the remaining three strands. This approach allows the introduction of specific modifications in only one of the strands. As a proof of concept, we prepared a quadruplex where one of the chains includes a change in polarity. A hybrid quadruplex is observed in ammonium acetate solutions, whereas in the presence of sodium or potassium, a parallel G-quadruplex structure is formed. In addition to the expected monomolecular quadruplexes, we observed the presence of dimeric G-quadruplex structures. We also applied the method to prepare G-quadruplexes containing a single 8-aminoguanine substitution and found that this single base stabilizes the G-quadruplex structure when located at an internal position.