期刊
NATURE COMMUNICATIONS
卷 11, 期 1, 页码 -出版社
NATURE PUBLISHING GROUP
DOI: 10.1038/s41467-020-17725-0
关键词
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资金
- Burroughs Wellcome Fund (Career Award at the Scientific Interface)
- DARPA [BrdiN66001-17-2-4055]
- NIH [UC4DK116255, R01 DK113597, HL095722]
Genetically fusing protein domains to Cas9 has yielded several transformative technologies; however, the genetic modifications are limited to natural polypeptide chains at the Cas9 termini, which excludes a diverse array of molecules useful for gene editing. Here, we report chemical modifications that allow site-specific and multiple-site conjugation of a wide assortment of molecules on both the termini and internal sites of Cas9, creating a platform for endowing Cas9 with diverse functions. Using this platform, Cas9 can be modified to more precisely incorporate exogenously supplied single-stranded oligonucleotide donor (ssODN) at the DNA break site. We demonstrate that the multiple-site conjugation of ssODN to Cas9 significantly increases the efficiency of precision genome editing, and such a platform is compatible with ssODNs of diverse lengths. By leveraging the conjugation platform, we successfully engineer INS-1E, a beta-cell line, to repurpose the insulin secretion machinery, which enables the glucose-dependent secretion of protective immunomodulatory factor interleukin-10.
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