Developing predictive hybridization models for phosphorothioate oligonucleotides using high-resolution melting

• Published in: PloS one Vol. 17; no. 5; p. e0268575
• Main Authors: Wang, Siyuan S., Xiong, Erhu, Bhadra, Sanchita, Ellington, Andrew D.
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 18.05.2022; Public Library of Science (PLoS)
• Subjects: Biology and Life Sciences; Circuit design; Datasets; Deoxyribonucleic acid; Design; DNA; DNA - genetics; Experimentation; Experiments; Helices; High resolution; Hybridization; Mathematical models; Melting; Modelling; Nucleic Acid Conformation; Nucleic Acid Hybridization; Nucleic acids; Oligonucleotides; Parameters; Phosphorothioate; Phosphorothioate Oligonucleotides; Physical Sciences; Prediction models; Regression analysis; Research and Analysis Methods; Stability; Standard deviation; United States; United States > US
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0268575

The ability to predict nucleic acid hybridization energies has been greatly enabling for many applications, but predictive models require painstaking experimentation, which may limit expansion to non-natural nucleic acid analogues and chemistries. We have assessed the utility of dye-based, high-resolution melting (HRM) as an alternative to UV-Vis determinations of hyperchromicity in order to more quickly acquire parameters for duplex stability prediction. The HRM-derived model for phosphodiester (PO) DNA can make comparable predictions to previously established models. Using HRM, it proved possible to develop predictive models for DNA duplexes containing phosphorothioate (PS) linkages, and we found that hybridization stability could be predicted as a function of sequence and backbone composition for a variety of duplexes, including PS:PS, PS:PO, and partially modified backbones. Individual phosphorothioate modifications destabilize helices by around 0.12 kcal/mol on average. Finally, we applied these models to the design of a catalytic hairpin assembly circuit, an enzyme-free amplification method used for nucleic acid-based molecular detection. Changes in PS circuit behavior were consistent with model predictions, further supporting the addition of HRM modeling and parameters for PS oligonucleotides to the rational design of nucleic acid hybridization.