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. 2022 Mar 30;13:867195. doi: 10.3389/fimmu.2022.867195

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Copyright © 2022 Moraes, Trentini, Fousteris, Eto, Chudzinski-Tavassi, Leite and Kanno

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Schematic representation of the pKLM-CRISPR vector. The pKLM-CRISPR vector, contains the expression cassette for codon optimized cas9 gene and a cassette for expression of sgRNA, both under regulation of pUV15tetO, a tetracycline-inducible promoter. The repressor tetR is expressed constitutively by the pimyc promoter. Transcriptional terminators (T) were introduced after each expression cassette. The restriction sites used for cloning are also indicated. The two BbsI sites were introduced to facilitate the cloning of the crRNA sequences upstream of tracrRNA. The vectors also contain a kanamycin resistance marker (KanR); origin of replication in E. coli (oriC) and mycobacteria (oriM).