Precise integration of inducible transcriptional elements (PrIITE) enables absolute control of gene expression

• Type: Article in International Scientific Journal
• Date: 2017

Authors

Pinto, R (Author) Other; Hansen, L (Author) Other; Hintze, J (Author) Other; Raquel Almeida (Author) FCUP; Larsen, S (Author) Other; Coskun, M (Author) Other; Davidsen, J (Author) Other; Mitchelmore, C (Author) Other; David L (Author) FMUP; Troelsen, JT (Author) Other; Bennett, EP (Author) Other

Journal

Title: Nucleic Acids Research

Vol. 45

ISSN: 0305-1048

Publisher: Oxford University Press

Indexing

ISI Web of Knowledge - 14 Citations

Scopus - 13 Citations

Other information

Authenticus ID: P-00M-QZZ

DOI: 10.1093/nar/gkx371

Abstract (EN): Tetracycline-based inducible systems provide powerful methods for functional studies where gene expression can be controlled. However, the lack of tight control of the inducible system, leading to leakiness and adverse effects caused by undesirable tetracycline dosage requirements, has proven to be a limitation. Here, we report that the combined use of genome editing tools and last generation Tet-On systems can resolve these issues. Our principle is based on precise integration of inducible transcriptional elements (coined PrIITE) targeted to: (i) exons of an endogenous gene of interest (GOI) and (ii) a safe harbor locus. Using PrIITE cells harboring a GFP reporter or CDX2 transcription factor, we demonstrate discrete inducibility of gene expression with complete abrogation of leakiness. CDX2 PrIITE cells generated by this approach uncovered novel CDX2 downstream effector genes. Our results provide a strategy for characterization of dose-dependent effector functions of essential genes that require absence of endogenous gene expression.

Language: English

Type (Professor's evaluation): Scientific

No. of pages: 15