Modular vector assembly enables rapid assessment of emerging CRISPR technologies.

Autor:	McGee AV; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Liu YV; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Griffith AL; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Szegletes ZM; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Wen B; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Kraus C; RNA Therapeutics Institute, University of Massachusetts Chan Medical School, Worcester, MA 01605, USA., Miller NW; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Steger RJ; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Velasco BE; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Bosch JA; Department of Genetics, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA., Zirin JD; Department of Genetics, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA., Viswanatha R; Department of Genetics, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA., Sontheimer EJ; RNA Therapeutics Institute, University of Massachusetts Chan Medical School, Worcester, MA 01605, USA.; Program in Molecular Medicine, University of Massachusetts Chan Medical School, Worcester, MA 01605, USA.; Li Weibo Institute for Rare Diseases Research, University of Massachusetts Chan Medical School, Worcester, MA 01605, USA., Goodale A; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Greene MA; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Green TM; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA., Doench JG; Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
Jazyk:	angličtina
Zdroj:	BioRxiv : the preprint server for biology [bioRxiv] 2023 Oct 27. Date of Electronic Publication: 2023 Oct 27.
DOI:	10.1101/2023.10.25.564061
Abstrakt:	The diversity of CRISPR systems, coupled with scientific ingenuity, has led to an explosion of applications; however, to test newly-described innovations in their model systems, researchers typically embark on cumbersome, one-off cloning projects to generate custom reagents that are optimized for their biological questions. Here, we leverage Golden Gate cloning to create the Fragmid toolkit, a modular set of CRISPR cassettes and delivery technologies, along with a web portal, resulting in a combinatorial platform that enables scalable vector assembly within days. We further demonstrate that multiple CRISPR technologies can be assessed in parallel in a pooled screening format using this resource, enabling the rapid optimization of both novel technologies and cellular models. These results establish Fragmid as a robust system for the rapid design of CRISPR vectors, and we anticipate that this assembly approach will be broadly useful for systematic development, comparison, and dissemination of CRISPR technologies. Competing Interests: COMPETING INTERESTS EJS is a cofounder, scientific advisor, and equity holder of Intellia Therapeutics, and a member of the scientific advisory board of Tessera Therapeutics. JGD consults for Microsoft Research, BioNTech, and Pfizer. JGD consults for and has equity in Tango Therapeutics. JGD serves as a paid scientific advisor to the Laboratory for Genomics Research, funded in part by GSK, and the Innovative Genomics Institute, funded in part by Apple Tree Partners. JGD receives funding support from the Functional Genomics Consortium: Abbvie, Bristol Myers Squibb, Janssen, Merck, and Vir Biotechnology. JGD’s interests are reviewed and managed by the Broad Institute in accordance with its conflict of interest policies.
Databáze:	MEDLINE
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