An unbiased genome-wide analysis of zinc-finger nuclease specificity - PubMed (original) (raw)
. 2011 Aug 7;29(9):816-23.
doi: 10.1038/nbt.1948.
Angelo Lombardo, Anne Arens, Jeffrey C Miller, Pietro Genovese, Christine Kaeppel, Ali Nowrouzi, Cynthia C Bartholomae, Jianbin Wang, Geoffrey Friedman, Michael C Holmes, Philip D Gregory, Hanno Glimm, Manfred Schmidt, Luigi Naldini, Christof von Kalle
Affiliations
- PMID: 21822255
- DOI: 10.1038/nbt.1948
An unbiased genome-wide analysis of zinc-finger nuclease specificity
Richard Gabriel et al. Nat Biotechnol. 2011.
Abstract
Zinc-finger nucleases (ZFNs) allow gene editing in live cells by inducing a targeted DNA double-strand break (DSB) at a specific genomic locus. However, strategies for characterizing the genome-wide specificity of ZFNs remain limited. We show that nonhomologous end-joining captures integrase-defective lentiviral vectors at DSBs, tagging these transient events. Genome-wide integration site analysis mapped the actual in vivo cleavage activity of four ZFN pairs targeting CCR5 or IL2RG. Ranking loci with repeatedly detectable nuclease activity by deep-sequencing allowed us to monitor the degree of ZFN specificity in vivo at these positions. Cleavage required binding of ZFNs in specific spatial arrangements on DNA bearing high homology to the intended target site and only tolerated mismatches at individual positions of the ZFN binding sites. Whereas the consensus binding sequence derived in vivo closely matched that obtained in biochemical experiments, the ranking of in vivo cleavage sites could not be predicted in silico. Comprehensive mapping of ZFN activity in vivo will facilitate the broad application of these reagents in translational research.
Similar articles
- Histone deacetylase inhibition rescues gene knockout levels achieved with integrase-defective lentiviral vectors encoding zinc-finger nucleases.
Pelascini LP, Maggio I, Liu J, Holkers M, Cathomen T, Gonçalves MA. Pelascini LP, et al. Hum Gene Ther Methods. 2013 Dec;24(6):399-411. doi: 10.1089/hgtb.2013.107. Epub 2013 Oct 29. Hum Gene Ther Methods. 2013. PMID: 24059449 Free PMC article. - Creating designed zinc-finger nucleases with minimal cytotoxicity.
Ramalingam S, Kandavelou K, Rajenderan R, Chandrasegaran S. Ramalingam S, et al. J Mol Biol. 2011 Jan 21;405(3):630-41. doi: 10.1016/j.jmb.2010.10.043. Epub 2010 Nov 19. J Mol Biol. 2011. PMID: 21094162 Free PMC article. - Genome editing with CompoZr custom zinc finger nucleases (ZFNs).
Hansen K, Coussens MJ, Sago J, Subramanian S, Gjoka M, Briner D. Hansen K, et al. J Vis Exp. 2012 Jun 14;(64):e3304. doi: 10.3791/3304. J Vis Exp. 2012. PMID: 22732945 Free PMC article. - Genome engineering with zinc-finger nucleases.
Carroll D. Carroll D. Genetics. 2011 Aug;188(4):773-82. doi: 10.1534/genetics.111.131433. Genetics. 2011. PMID: 21828278 Free PMC article. Review. - Zinc Finger Nucleases: A new era for transgenic animals.
Swarthout JT, Raisinghani M, Cui X. Swarthout JT, et al. Ann Neurosci. 2011 Jan;18(1):25-8. doi: 10.5214/ans.0972.7531.1118109. Ann Neurosci. 2011. PMID: 25205916 Free PMC article. Review.
Cited by
- Advances in the labelling and selective manipulation of synapses.
Timalsina B, Lee S, Kaang BK. Timalsina B, et al. Nat Rev Neurosci. 2024 Oct;25(10):668-687. doi: 10.1038/s41583-024-00851-9. Epub 2024 Aug 22. Nat Rev Neurosci. 2024. PMID: 39174832 Review. - Beyond the promise: evaluating and mitigating off-target effects in CRISPR gene editing for safer therapeutics.
Lopes R, Prasad MK. Lopes R, et al. Front Bioeng Biotechnol. 2024 Jan 18;11:1339189. doi: 10.3389/fbioe.2023.1339189. eCollection 2023. Front Bioeng Biotechnol. 2024. PMID: 38390600 Free PMC article. Review. - Reshaping the Landscape of the Genome: Toolkits for Precise DNA Methylation Manipulation and Beyond.
Zhu C, Hao Z, Liu D. Zhu C, et al. JACS Au. 2023 Dec 21;4(1):40-57. doi: 10.1021/jacsau.3c00671. eCollection 2024 Jan 22. JACS Au. 2023. PMID: 38274248 Free PMC article. Review. - Site-specific genome editing in treatment of inherited diseases: possibility, progress, and perspectives.
Huang C, Li Q, Li J. Huang C, et al. Med Rev (2021). 2022 Nov 11;2(5):471-500. doi: 10.1515/mr-2022-0029. eCollection 2022 Oct. Med Rev (2021). 2022. PMID: 37724161 Free PMC article. Review. - Progress and Prospects of Gene Editing in Pluripotent Stem Cells.
Zhang Z, Bao X, Lin CP. Zhang Z, et al. Biomedicines. 2023 Aug 1;11(8):2168. doi: 10.3390/biomedicines11082168. Biomedicines. 2023. PMID: 37626665 Free PMC article. Review.
References
- Nature. 2005 Jun 2;435(7042):646-51 - PubMed
- Biochem Biophys Res Commun. 2005 Sep 9;334(4):1191-1197 - PubMed
- Methods Mol Biol. 2010;649:247-56 - PubMed
- Biotechnol Bioeng. 2010 May 1;106(1):97-105 - PubMed
- DNA Repair (Amst). 2007 Jun 1;6(6):781-8 - PubMed
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Other Literature Sources