CRISPR-Based Self-Cleaving Mechanism for Controllable Gene Delivery in Human Cells




Journal Title

Journal ISSN

Volume Title


Oxford University Press


Controllable gene delivery via vector-based systems remains a formidable challenge in mammalian synthetic biology and a desirable asset in gene therapy applications. Here, we introduce a methodology to control the copies and residence time of a gene product delivered in host human cells but also selectively disrupt fragments of the delivery vehicle. A crucial element 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 specific parameters of the system can be adjusted to fine-tune the delivery properties. We envision future applications in complex synthetic biology architectures, gene therapy and trace-free delivery.;


Supplementary materials are available at Nucleic Acids Research Online (see DOI).


mKate2 protein, Cas9 protein, Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR), Guide RNA (gRNA), Gene delivery

US National Institutes of Health (GM098984, GM096271, CA17001801); US National Science Foundation (CBNET-1105524)


CC BY-NC 4.0 (Attribution-Non-commercial), ©2014 The Authors