A rule of seven in Watson-Crick base-pairing of mismatched sequences

Abstract

Sequence recognition through base-pairing is essential for DNA repair and gene regulation, but the basic rules governing this process remain elusive. In particular, the kinetics of annealing between two imperfectly matched strands is not well characterized, despite its potential importance in nucleic acid–based biotechnologies and gene silencing. Here we use single-molecule fluorescence to visualize the multiple annealing and melting reactions of two untethered strands inside a porous vesicle, allowing us to precisely quantify the annealing and melting rates. The data as a function of mismatch position suggest that seven contiguous base pairs are needed for rapid annealing of DNA and RNA. This phenomenological rule of seven may underlie the requirement for seven nucleotides of complementarity to seed gene silencing by small noncoding RNA and may help guide performance improvement in DNA- and RNA-based bio- and nanotechnologies, in which off-target effects can be detrimental.

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Figure 1: Single-molecule porous-vesicle encapsulation assay.
Figure 2: The effect of terminal base-pair mismatch.
Figure 3: The effect of DNA mismatch position.
Figure 4: The effect of mismatch position on RNA.

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Acknowledgements

We thank B. Okumus, R. Clegg and Z. Bryant for critical suggestions. We acknowledge J. Chen, C. Joo and the members of N. Kim's group for discussion on microRNA. We thank current and past members of the Ha group for various suggestions. The project was supported by US National Institutes of Health grants GM074526 and GM065367, and US National Science Foundation grant 0822613 to T.H.; H.K. was supported in part by grant (KRF-2006-352-C00019) of the Korean Research Foundation.

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I.I.C. and T.H. designed the initial experiments. I.I.C. and H.K. performed the experiments and analyzed the data. I.I.C., H.K. and T.H. wrote the manuscript.

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Correspondence to Taekjip Ha.

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The authors declare no competing financial interests.

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Supplementary Figures 1–4 and Supplementary Table 1 (PDF 431 kb)

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Cisse, I., Kim, H. & Ha, T. A rule of seven in Watson-Crick base-pairing of mismatched sequences. Nat Struct Mol Biol 19, 623–627 (2012). https://doi.org/10.1038/nsmb.2294

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