Application
Flexible system using Cas9 to modulate gene expression.
Key Benefits
- Edits genes by cleaving DNA as well as inhibiting transcription.
- No need to transfect cells with additional Cas9.
Market Summary
The CRISPR-Cas9 system is a bacterial adaptive immune system that has been appropriated for genetic engineering. The CRISPR-Cas9 system consists of an effector protein, Cas9, and two small RNAs. The two RNAs form a hairpin that complexes with Cas9 and guide the Cas9 to a specific DNA sequence determined by the spacer sequence, and Cas9 can then cleave the DNA in that region. However, the CRISPR-Cas9 system is limited in its ability to transfect or deliver the Cas9 protein to the cell. The protein is large, making delivery complicated, and expression of the protein at high levels may cause cell toxicity.
Technical Summary
Emory researchers have discovered a secondary function for the Cas9 protein in Francisella novicida (FnoCas9). This Cas9 protein blocks transcription but does not cleave the DNA. Importantly, the FnoCas9 remains enzymatically active and can be made to cleave the DNA if given a sgRNA with a longer spacer sequence with more complementary bases to the target sequence. Emory researchers propose to use this novel FnoCasa9-scaRNA system as a method to either suppress transcription of a gene using the shorter scaRNA, or cleave the gene using the longer crRNA. This multi-functional Cas9 can perform both functions without the need to transfect the cell with an additional copy of a defective Cas9. This technology could potentially be used to develop gene therapy. Data also suggests this can be used in a Cas12a (Cpf1)-mediated system.
Developmental Stage
Researchers have shown this technology can be used to sensitize bacteria to antimicrobials.
Publication: Ratner, H. et al. (2019). Molecular Cell, 75(3), 498-510.
