title: Benchmarking of TALE- and CRISPR/dCas9-Based Transcriptional Regulators in Mammalian Cells for the Construction of Synthetic Genetic Circuits.
abstract: Transcriptional activator-like effector (TALE)- and CRISPR/Cas9-based designable recognition domains represent a technological breakthrough not only for genome editing but also for building designed genetic circuits. Both platforms are able to target rarely occurring DNA segments, even within complex genomes. TALE and dCas9 domains, genetically fused to transcriptional regulatory domains, can be used for the construction of engineered logic circuits. Here we benchmarked the performance of the two platforms, targeting the same DNA sequences, to compare their advantages for the construction of designed circuits in mammalian cells. Optimal targeting strands for repression and activation of dCas9-based designed transcription factors were identified; both platforms exhibited good orthogonality and were used to construct functionally complete NOR gates. Although the CRISPR/dCas9 system is clearly easier to construct, TALE-based activators were significantly stronger, and the TALE-based platform performed better, especially for the construction of layered circuits.
authors.affiliation: Department of Synthetic Biology and Immunology, National Institute of Chemistry , 1000 Ljubljana, Slovenia.|EN-FIST Centre of Excellence , 1000 Ljubljana, Slovenia.|Graduate School of Biomedicine, University of Ljubljana , 1000 Ljubljana, Slovenia.
authors.forename: Tina
authors.initials: T
authors.lastname: Lebar
authors.affiliation: Department of Synthetic Biology and Immunology, National Institute of Chemistry , 1000 Ljubljana, Slovenia.|EN-FIST Centre of Excellence , 1000 Ljubljana, Slovenia.
authors.forename: Roman
authors.initials: R
authors.lastname: Jerala
mesh_terms: D000818:Animals; D019985:Benchmarking; D016466:CHO Cells; D064113:CRISPR-Cas Systems; D003412:Cricetulus; D005786:Gene Expression Regulation; D053263:Gene Regulatory Networks; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D000072417:Protein Domains; D000094704:RNA, Guide, CRISPR-Cas Systems; D012097:Repressor Proteins / Q000235:genetics; D000071336:Transcription Activator-Like Effectors; D014157:Transcription Factors / Q000235:genetics
chemical_list: D000094704:RNA, Guide, CRISPR-Cas Systems; D012097:Repressor Proteins; D000071336:Transcription Activator-Like Effectors; D014157:Transcription Factors
keywords: CRISPR/Cas9; NOR gate; TAL effectors; synthetic circuits; transcriptional regulation