Cotterell, JamesVila-Cejudo, MartaBatlle Morera, LauraSharpe, James2020-10-082020-10-082020Cotterell J, Vila-Cejudo M, Batlle-Morera L, Sharpe J. Endogenous CRISPR/Cas9 arrays for scalable whole-organism lineage tracing. Development. 2020 May 12; 147(9): dev184481. DOI: 10.1242/dev.1844810950-1991http://hdl.handle.net/10230/45429The past decade has seen a renewed appreciation of the central importance of cellular lineages to many questions in biology (especially organogenesis, stem cells and tumor biology). This has been driven in part by a renaissance in genetic clonal-labeling techniques. Recent approaches are based on accelerated mutation of DNA sequences, which can then be sequenced from individual cells to re-create a 'phylogenetic' tree of cell lineage. However, current approaches depend on making transgenic alterations to the genome in question, which limit their application. Here, we introduce a new method that completely avoids the need for prior genetic engineering, by identifying endogenous CRISPR/Cas9 target arrays suitable for lineage analysis. In both mouse and zebrafish, we identify the highest quality compact arrays as judged by equal base composition, 5' G sequence, minimal likelihood of residing in the functional genome, minimal off targets and ease of amplification. We validate multiple high-quality endogenous CRISPR/Cas9 arrays, demonstrating their utility for lineage tracing. Our pragmatically scalable technique thus can produce deep and broad lineages in vivo, while removing the dependence on genetic engineering.application/pdfeng© 2020. Published by The Company of Biologists Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributedLlinatge cel·lularMorfogènesiCèl·lules mareSeqüència de nucleòtidsGenomesinfo:eu-repo/semantics/articlehttp://dx.doi.org/10.1242/dev.184481info:eu-repo/semantics/openAccess