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  • Title: Design and Application of DNA Modification-Specific Transcription-Activator-Like Effectors.
    Author: Buchmuller B, Muñoz-López Á, Gieß M, Summerer D.
    Journal: Methods Mol Biol; 2021; 2198():381-399. PubMed ID: 32822046.
    Abstract:
    Transcription-activator like effectors (TALEs) are DNA-binding proteins used for genome targeting. TALEs contain a central domain of concatenated repeats, of which each selectively recognizes one nucleobase at the DNA major groove. Based on this simple and predictable interaction with little context dependence, TALEs offer programmable targeting of user-defined DNA sequences. Since many epigenetic DNA modifications protrude into the DNA major groove, natural and engineered TALE repeats can provide "epigenetic" selectivity, making TALEs a flexible platform to design probes for the analysis of epigenetic DNA modifications. Here, we describe guidelines for the design of TALE proteins with selectivity for epigenetic cytosine 5-modifications, the validation of their interaction with modified DNA nucleobases, and their employment in affinity enrichment assays. These techniques enable quantification of epigenetic nucleobases in user-defined genomic DNA sequences with nucleotide and strand resolution.
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