Objective Enhancers are cis-regulatory genomic regions that can modulate gene expression in a cell type-specific and time-controlled manner to regulate cellular behaviour. ChIP-seq and RNA-seq technologies have allowed genome-wide identification of potential enhancers based on the correlation among specific chromatin marks, transcription factor biding sites and gene expression. However, strategies to validate their function are still limited. Recently, it has been demonstrated that the CRISPR/Cas9 genome editing technology can be used to delete large genomic regions at high frequency by co-transfection of two guide RNAs (gRNAs) targeting collinear genomic sites and subsequent NHEJ-mediated repair. Nevertheless, this system is limited to individual tests. Here we aim to generate a novel lentiviral dual-gRNA expression system that enables large scale functional enhancer screening. The candidate will apply this dual-gRNA expression system to generate custom libraries that will allow the functional identification and subsequent validation of essential enhancers for the maintenance of mouse Embryonic Stem (mES) cell identity. This technology will provide a comprehensive view of the function of enhancers within their genomic context and will enable for the first time the performance of systematic, genetic forward enhancer screens. Fields of science medical and health sciencesmedical biotechnologygenetic engineeringgene therapynatural sciencesbiological sciencesdevelopmental biologymedical and health sciencesmedical biotechnologycells technologiesstem cellsmedical and health sciencesclinical medicineoncologynatural sciencesbiological sciencesgeneticsgenomes Keywords Stem Cells Enhancers CRISPR Cas9 Forward Genetic Screens Programme(s) H2020-EU.1.3. - EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions Main Programme H2020-EU.1.3.2. - Nurturing excellence by means of cross-border and cross-sector mobility Topic(s) MSCA-IF-2014-EF - Marie Skłodowska-Curie Individual Fellowships (IF-EF) Call for proposal H2020-MSCA-IF-2014 See other projects for this call Funding Scheme MSCA-IF - Marie Skłodowska-Curie Individual Fellowships (IF) Coordinator THE UNIVERSITY OF EDINBURGH Net EU contribution € 183 454,80 Address OLD COLLEGE, SOUTH BRIDGE EH8 9YL Edinburgh United Kingdom See on map Region Scotland Eastern Scotland Edinburgh Activity type Higher or Secondary Education Establishments Links Contact the organisation Opens in new window Website Opens in new window Participation in EU R&I programmes Opens in new window HORIZON collaboration network Opens in new window Total cost € 183 454,80