The CRISPR-Cas9 system has been adapted to upregulate any gene in its endogenous context, enabling overexpression experiments. The simplicity of programming this CRISPR activation (CRISPRa) system with small RNA guides is transformative for performing systematic gain-of-function studies. This exciting complement to loss-of-function studies also provides new tools to identify gene functions that might otherwise go undetected using a downregulation or knockout mechanism alone.
We will describe different CRISPRa technologies and provide experimental considerations for using either synthetic guide RNA (crRNA:tracrRNA) or lentiviral sgRNA, in either arrayed or pooled screening formats. These platforms can be applied in tandem to enable both rapid follow-up of screen hits and development of complementary assays for functional gene analysis.
In this webinar you will learn:
- The mechanisms of CRISPR activation
- How to set up CRISPRa experiments using predesigned guide RNA
- The relationship between baseline gene expression and activation levels
- The basics of using pooled lentiviral CRISPRa screening for target ID and validation
Senior Scientist at Dharmacon, a Horizon Discovery Group company
Žaklina Strezoska is a Senior Scientist at Dharmacon, a Horizon Discovery Group company. She joined Dharmacon in 2007 and currently leads projects in multiple research areas including RNA interference, CRISPR-Cas9 genome engineering, and CRISPRa transcriptional modulation with a focus on innovation and new product development. She has extensive experience in arrayed and pooled screening and hit identification and validation. Prior to joining Dharmacon, she worked at ALTANA Pharma where she gained experience in target identification, validation, and small molecule discovery. Žaklina received her B.S. and M.S. degrees in Molecular Biology from Belgrade University and her PhD. Degree in Molecular Genetics at the University of Illinois at Chicago. She completed a postdoctoral fellowship at Brown University.
Benedict Cross R&D Manager, Functional Genomic Screening, Horizon Discovery
Ben completed his PhD at the University of Manchester and trained as a post-doctoral scientist at the University of Cambridge studying reverse chemical genetic screening in the unfolded protein response. His research identified and defined the mechanism of a new tool compound against a key proteostasis kinase. Ben joined Horizon in 2013 to expand and advance Horizon's functional genomic screening capability and to lead a major research alliance in synthetic lethal target discovery. Ben led the development of pooled CRISPR-Cas9 screening at Horizon which was launched as a service in 2015. The addition of CRISPRi and CRISPRa followed in 2017 and Ben now manages the teams applying these functional genomic screening platforms in target discovery, validation and other research programs.