W. Jiang and L. A. Marraffini, CRISPR-Cas: New Tools for Genetic Manipulations from Bacterial Immunity Systems, Annual Review of Microbiology, vol.69, issue.1, pp.209-228, 2015.
DOI : 10.1146/annurev-micro-091014-104441

M. Jinek, A Programmable Dual-RNA-Guided DNA Endonuclease in Adaptive Bacterial Immunity, Science, vol.274, issue.45, pp.816-821, 2012.
DOI : 10.1074/jbc.274.45.31896

S. H. Sternberg, S. Redding, M. Jinek, E. C. Greene, and J. A. Doudna, DNA interrogation by the CRISPR RNA-guided endonuclease Cas9, Nature, vol.18, issue.7490, pp.62-67, 2014.
DOI : 10.1261/rna.030882.111

F. J. Mojica, C. Díez-villaseñor, J. García-martínez, and C. Almendros, Short motif sequences determine the targets of the prokaryotic CRISPR defence system, Microbiology, vol.155, pp.733-740, 2009.

|. Doi, 10.1038/s41467-018-04209-5 ARTICLE, NATURE COMMUNICATIONS NATURE COMMUNICATIONS |, vol.9, 1912.
URL : https://hal.archives-ouvertes.fr/in2p3-00652853

E. Semenova, Interference by clustered regularly interspaced short palindromic repeat (CRISPR) RNA is governed by a seed sequence, Proc. Natl. Acad. Sci. USA, pp.10098-10103, 2011.
DOI : 10.1038/nsmb.2019

L. S. Qi, Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression, Cell, vol.152, issue.5, pp.1173-1183, 2013.
DOI : 10.1016/j.cell.2013.02.022

D. Bikard, Programmable repression and activation of bacterial gene expression using an engineered CRISPR-Cas system, Nucleic Acids Research, vol.109, issue.15, pp.7429-7437, 2013.
DOI : 10.1073/pnas.1109479109

J. M. Peters, A Comprehensive, CRISPR-based Functional Analysis of Essential Genes in Bacteria, Cell, vol.165, issue.6, pp.1493-1506, 2016.
DOI : 10.1016/j.cell.2016.05.003

X. Liu, Molecular Systems Biology, vol.13, issue.5, p.931, 2017.
DOI : 10.15252/msb.20167449

Y. Fu, High-frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells, Nature Biotechnology, vol.31, issue.9, pp.822-826, 2013.
DOI : 10.1021/bi00035a029

V. Pattanayak, High-throughput profiling of off-target DNA cleavage reveals RNA-programmed Cas9 nuclease specificity, Nature Biotechnology, vol.31, issue.9, pp.839-843, 2013.
DOI : 10.1021/ja057519l

P. D. Hsu, DNA targeting specificity of RNA-guided Cas9 nucleases, Nature Biotechnology, vol.49, issue.9, pp.827-832, 2013.
DOI : 10.1073/pnas.1019533108

S. H. Sternberg, B. Lafrance, M. Kaplan, and J. A. Doudna, Conformational control of DNA target cleavage by CRISPR???Cas9, Nature, vol.38, issue.7576, pp.110-113, 2015.
DOI : 10.1093/nar/gkq399

M. Jinek, Structures of Cas9 Endonucleases Reveal RNA-Mediated Conformational Activation, Science, vol.133, issue.2-3, p.1247997, 2014.
DOI : 10.1006/jsbi.2000.4350

C. Kuscu, S. Arslan, R. Singh, J. Thorpe, and M. Adli, Genome-wide analysis reveals characteristics of off-target sites bound by the Cas9 endonuclease, Nature Biotechnology, vol.32, issue.7, pp.677-683, 2014.
DOI : 10.1016/j.molcel.2010.05.004

X. Wu, Genome-wide binding of the CRISPR endonuclease Cas9 in mammalian cells, Nature Biotechnology, vol.480, issue.7, pp.670-676, 2014.
DOI : 10.1186/gb-2009-10-3-r25

D. Singh, S. H. Sternberg, J. Fei, J. A. Doudna, and T. Ha, Real-time observation of DNA recognition and rejection by the RNA-guided endonuclease Cas9, Nature Communications, vol.44, issue.1247, p.12778, 2016.
DOI : 10.1093/nar/gkw398
URL : http://www.nature.com/articles/ncomms12778.pdf

L. A. Gilbert, Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation, Cell, vol.159, issue.3, pp.647-661, 2014.
DOI : 10.1016/j.cell.2014.09.029
URL : https://doi.org/10.1016/j.cell.2014.09.029

H. Zhao, ABSTRACT, Journal of Bacteriology, vol.198, issue.21, pp.2925-2935, 2016.
DOI : 10.1128/JB.00507-16
URL : https://hal.archives-ouvertes.fr/hal-00218335

I. Farasat, Efficient search, mapping, and optimization of multi-protein genetic systems in diverse bacteria, Molecular Systems Biology, vol.10, issue.6, p.731, 2014.
DOI : 10.15252/msb.20134955
URL : http://msb.embopress.org/content/msb/10/6/731.full.pdf

A. A. Nielsen and C. A. Voigt, Multi-input CRISPR/Cas genetic circuits that interface host regulatory networks, Molecular Systems Biology, vol.10, issue.11, p.763, 2014.
DOI : 10.15252/msb.20145735
URL : http://msb.embopress.org/content/msb/10/11/763.full.pdf

D. G. Gibson, Enzymatic assembly of DNA molecules up to several hundred kilobases, Nature Methods, vol.102, issue.5, pp.343-345, 2009.
DOI : 10.1038/nmeth.1318

C. Engler, R. Kandzia, and S. Marillonnet, A One Pot, One Step, Precision Cloning Method with High Throughput Capability, PLoS ONE, vol.103, issue.11, p.3647, 2008.
DOI : 10.1371/journal.pone.0003647.t002
URL : https://doi.org/10.1371/journal.pone.0003647

F. St-pierre, One-Step Cloning and Chromosomal Integration of DNA, ACS Synthetic Biology, vol.2, issue.9, pp.537-541, 2013.
DOI : 10.1021/sb400021j

S. K. Sharan, L. C. Thomason, S. G. Kuznetsov, and D. L. Court, Recombineering: a homologous recombination-based method of genetic engineering, Nature Protocols, vol.24, issue.2, pp.206-223, 2009.
DOI : 10.1128/JB.01695-07
URL : http://www.nature.com/nprot/journal/v4/n2/pdf/nprot.2008.227.pdf

M. Chaveroche, J. Ghigo, and C. Enfert, A rapid method for efficient gene replacement in the filamentous fungus Aspergillus nidulans, Nucleic Acids Research, vol.28, issue.22, p.97, 2000.
DOI : 10.1093/nar/28.22.e97

M. I. Love, W. Huber, and S. Anders, Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2, Genome Biology, vol.14, issue.12, p.550, 2014.
DOI : 10.1186/gb-2013-14-4-r36
URL : https://genomebiology.biomedcentral.com/track/pdf/10.1186/s13059-014-0550-8?site=genomebiology.biomedcentral.com

F. Pedregosa, Scikit-learn: machine learning in Python, J. Mach. Learn. Res, vol.12, pp.2825-2830, 2011.
URL : https://hal.archives-ouvertes.fr/hal-00650905

D. P. Kingma and J. Ba, Adam: a method for stochastic optimization, 2014.

T. D. Schmittgen and K. J. Livak, Analyzing real-time PCR data by the comparative CT method, Nature Protocols, vol.2, issue.6, pp.1101-1108, 2008.
DOI : 10.1593/neo.07916

J. E. Cronan, Improved plasmid-based system for fully regulated off-to-on gene expression in Escherichia coli: Application to production of toxic proteins, Plasmid, vol.69, issue.1, pp.81-89, 2013.
DOI : 10.1016/j.plasmid.2012.09.003

C. Cinesi, L. Aeschbach, B. Yang, and V. Dion, Contracting CAG/CTG repeats using the CRISPR-Cas9 nickase, Nature Communications, vol.17, p.13272, 2016.
DOI : 10.1093/hmg/ddn019
URL : http://www.nature.com/articles/ncomms13272.pdf

, consortium (ANR10-INBS-09-08) This work was supported by the European Research Council (ERC) under the Europe Union's Horizon 2020 research and innovation program (grant agreement No. [677823]); the French Government's Investissement d'Avenir program; Laboratoire d'Excellence 'Integrative Biology of Emerging Infectious Diseases' [ANR-10-LABX-62-IBEID]

L. C. Author and D. B. , designed the study and wrote the manuscript. L.C. performed the experiments. A.V. performed the mcherry fluorescence measurements