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Nucleic acids are versatile scaffolds that accommodate a wide range of precisely defined operational characteristics. Rational design of sensing, molecular computing, nanotechnology, and other nucleic acid devices requires precise control over folding conformations in these macromolecules. Here, we report a new approach that empowers well-defined conformational transitions in DNA molecular devices. Specifically, we develop tools for precise folding of multiple DNA quadruplexes (i-motifs) within the same oligonucleotide strand. To accompli