Abby V McGee, Yanjing V Liu, Audrey L Griffith, Zsofia M Szegletes, Bronte Wen, Carolyn Kraus, Nathan W Miller, Ryan J Steger, Berta Escude Velasco, Justin A Bosch, Jonathan D Zirin, Raghuvir Viswanatha, Erik J Sontheimer, Amy Goodale, Matthew A Greene, Thomas M Green, John G Doench
{"title":"Modular vector assembly enables rapid assessment of emerging CRISPR technologies.","authors":"Abby V McGee, Yanjing V Liu, Audrey L Griffith, Zsofia M Szegletes, Bronte Wen, Carolyn Kraus, Nathan W Miller, Ryan J Steger, Berta Escude Velasco, Justin A Bosch, Jonathan D Zirin, Raghuvir Viswanatha, Erik J Sontheimer, Amy Goodale, Matthew A Greene, Thomas M Green, John G Doench","doi":"10.1016/j.xgen.2024.100519","DOIUrl":null,"url":null,"abstract":"<p><p>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.</p>","PeriodicalId":72539,"journal":{"name":"Cell genomics","volume":null,"pages":null},"PeriodicalIF":11.1000,"publicationDate":"2024-03-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10943585/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Cell genomics","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1016/j.xgen.2024.100519","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"CELL BIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
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.