“Parallel” and “Antiparallel Tail‐Clamps” Increase the Efficiency of Triplex Formation with Structured DNA and RNA Targets

Abstract

Sequence-specific triple-helix structures can be formed by parallel and antiparallel DNA clamps interacting with single-stranded DNA or RNA targets. Single-stranded nucleic acid molecules are known to adopt secondary structures that might interfere with intermolecular interactions. We demonstrate the correlation between a secondary structure involving the target--a stable stem predicted by in silico folding and experimentally confirmed by thermal stability and competition analyses--and an inhibitory effect on triplex formation. We overcame structural impediments by designing a new type of clamp: "tail-clamps". A combination of gel-shift, kinetic analysis, UV thermal melting and thermodynamic techniques was used to demonstrate that tail-clamps efficiently form triple helices with a structured target sequence. The performance of parallel and antiparallel tail-clamps was compared: antiparallel tail-clamps had higher binding efficiencies than parallel tail-clamps both with structured DNA and RNA targets. In addition, the reported triplex-stabilizing property of 8-aminopurine residues was confirmed for tail-clamps. Finally, we discuss the possible use of this improved triplex technology as a new tool for applications in molecular biology.

Design of oligonucleotide-capped mesoporous silica nanoparticles for the detection of miRNA-145 by duplex and triplex formation

“…Triplex can only be formed at specific polypurine-polypyrimidine sequences, which are widespread within the human genome, especially at promoter regions [10,11]. Previous works have reported the triplex-stabilizing properties of 8-aminopurines and the use of parallel and antiparallel tail-clamps to increase the efficiency of triplex formation with structured DNA and RNA targets [12,13]. This triplex affinity capture strategy has been successfully applied for the detection of Listeria mRNA and miRNA-145 using a SPR biosensor [14,15].…”

Strand Displacement of Double-Stranded DNA by Triplex-Forming Antiparallel Purine-Hairpins

“…However, in the biomedical field, antiparallel triplexes using duplex oligonucleotides are more promising than parallel structures, since the formation of antiparallel triplex is pH independent (Vasquez et al 1995;Faucon et al 1996;Mills et al 2002). In this regard, several groups described the ability of purine-hairpins to form an antiparallel triple helical structure with a polypyrimidine single-stranded targets (Wang 1994;Vo et al 1995;Mills et al 1999;Avino et al 2003;Nadal et al 2005). These triplex-forming antiparallel purine- Until now, this novel strategy for antiparallel pyr.pur*pur triplex formation is restricted to the binding of antiparallel purine-hairpins to single-stranded targets.…”

Parallel Clamps and Polypurine Hairpins (PPRH) for Gene Silencing and Triplex‐Affinity Capture: Design, Synthesis, and Use

“…This polypurine sequence contains three pyrimidine interruptions in each strand. , wedged (Rodriguez et al, 2015), and tail-clamps (Nadal et al, 2006;Nadal, Eritja, Esteve, & Plà, 2005). In all of these cases, a higher affinity polypyridine target and an increase in specificity have been observed.…”

“…). These include cyclic oligonucleotides (Grimau, Gargallo, Aviñó, & Eritja, ), wedged (Rodriguez et al., ), and tail‐clamps (Nadal et al., ; Nadal, Eritja, Esteve, & Plà, ). In all of these cases, a higher affinity polypyridine target and an increase in specificity have been observed.…”