期刊
NATURE CHEMISTRY
卷 4, 期 3, 页码 208-214出版社
NATURE RESEARCH
DOI: 10.1038/NCHEM.1246
关键词
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资金
- Wyss Institute for Biologically Inspired Engineered faculty
- NIH [1DP2OD007292]
- NSF CAREER [CCF1054898]
- Office of Naval Research [N000141010827]
- Life Sciences Research Foundation
- Direct For Computer & Info Scie & Enginr
- Division of Computing and Communication Foundations [1054898] Funding Source: National Science Foundation
The specific hybridization of complementary sequences is an essential property of nucleic acids, enabling diverse biological and biotechnological reactions and functions. However, the specificity of nucleic acid hybridization is compromised for long strands, except near the melting temperature. Here, we analytically derived the thermodynamic properties of a hybridization probe that would enable near-optimal single-base discrimination and perform robustly across diverse temperature, salt and concentration conditions. We rationally designed 'toehold exchange' probes that approximate these properties, and comprehensively tested them against five different DNA targets and 55 spurious analogues with energetically representative single-base changes (replacements, deletions and insertions). These probes produced discrimination factors between 3 and 100(+) (median, 26). Without retuning, our probes function robustly from 10 degrees C to 37 degrees C, from 1 mM Mg2+ to 47 mM Mg2+, and with nucleic acid concentrations from 1 nM to 5 mu M. Experiments with RNA also showed effective single-base change discrimination.
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