Abstract
We present a robust strategy to covalently link proteins and DNA using HUH-endonuclease domains as fusion partners (HUH-tags). We show that HUH-tags react robustly with specific sequences of unmodified single-stranded DNA, and we have identified five tags that react orthogonally with distinct DNA sequences. We demonstrate the versatility of HUH-tags as fusion partners in Cas9-mediated gene editing and the construction of doubly DNA-tethered proteins for single-molecule studies. Finally we demonstrate application to cellular imaging in live and fixed cells.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 7030-7035 |
| Number of pages | 6 |
| Journal | Journal of the American Chemical Society |
| Volume | 139 |
| Issue number | 20 |
| DOIs | |
| State | Published - May 24 2017 |
Bibliographical note
Funding Information:This study was supported by an NIH NIGMS R35 grant to W.R.G. (1R35GM119483), funds to W.R.G. from the Pew Biomedical Scholar Program (29634), and UMN startup funds. We would like to thank Peter Gordon for the kind gift of inducible GFP cell lines. We also would like to thank Eric Hendrickson and Brian Ruis for helpful discussions about the Cas9 experiments. Typhoon FLA9500 images were collected at the University of Minnesota Imaging Center.
Publisher Copyright:
© 2017 American Chemical Society.
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Mark A Sanders (Program Director) & Guillermo Marques (Scientific Director)
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