CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells

Controllable gene delivery via vector-based systems remains a formidable challenge in mammalian syn-thetic biology and a desirable asset in gene therapy applications. Here, we introduce a methodology to control the copies and residence time of a gene prod-uct delivered in host human cells but also selectively disrupt fragments of the delivery vehicle. A crucial el-ement of the proposed system is the CRISPR protein Cas9. Upon delivery, Cas9 guided by a custom RNA sequence cleaves the delivery vector at strategically placed targets thereby inactivating a co-expressed gene of interest. Importantly, using experiments in human embryonic kidney cells, we show that spe-cific parameters of the system can be adjusted to fine-tune the delivery properties. We envision future applications in complex synthetic biology architec-tures, gene therapy and trace-free delivery