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

Date
2014-12-18
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Journal Title
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Volume Title
Publisher
Oxford University Press
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Abstract

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.;

Description
Supplementary materials are available at Nucleic Acids Research Online (see DOI).
Keywords
mKate2 protein, Cas9 protein, Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR), Guide RNA (gRNA), Gene delivery
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US National Institutes of Health (GM098984, GM096271, CA17001801); US National Science Foundation (CBNET-1105524)
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CC BY-NC 4.0 (Attribution-Non-commercial), ©2014 The Authors
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