Type III CRISPR-Cas System for Robust RNA Knockdown and Imaging in Eukaryotes
Tech ID: 32820 / UC Case 2022-128-0
Patent Status
| Country | Type | Number | Dated | Case |
| United States Of America | Published Application | 20240026323 | 01/25/2024 | 2022-128 |
Brief Description
Type III CRISPR-Cas systems recognize and degrade RNA molecules using an RNA-guided mechanism that occurs widely in microbes for adaptive immunity against viruses. The inventors have demonstrated that this multi-protein system can be leveraged for programmable RNA knockdown of both nuclear and cytoplasmic transcripts in mammalian cells.
Using single-vector delivery of the S. thermophilus Csm complex, RNA knockdown was achieved with high efficiency (90-99%) and minimal off-targets, outperforming existing technologies of shRNA- and Cas13-mediated knockdown. Furthermore, unlike Cas13, Csm is devoid of trans-cleavage activity and thus does not induce non-specific transcriptome-wide degradation and cytotoxicity. Catalytically inactivated Csm can also be used for programmable RNA-binding, which the inventors exploit for live-cell RNA imaging. This work demonstrates the feasibility and efficacy of multi-subunit CRISPR-Cas effector complexes as RNA-targeting tools in eukaryotes.
Suggested uses
This invention has the potential to completely supplant RNAi and Cas13-based RNA knockdown technologies.
Suggested uses include:
- research purposes requiring gene perturbation at the transcript (RNA) but not genome (DNA) level.
- efficiently knocking down both cytoplasmic and nuclear RNAs.
- RNA imaging in live cells.
- RNA detection for diagnostic purposes.
- utilizing the enzyme's ssDNase and/or cyclic oligoadenylate synthesis activities.
Advantages
The advantages include:
- yields robust, highly efficient RNA knockdown with minimal off-targets and no cell toxicity
- cheap, easy to use, and easy to deliver as an all-in-one vector
- allows for gene perturbation at the transcript (RNA) level
- efficiently knocks down both cytoplasmic and nuclear RNAs
Related Materials
Contact
- Terri Sale
- terri.sale@berkeley.edu
- tel: View Phone Number.
Inventors
- Doudna, Jennifer A.
Other Information
Keywords
siRNA, shRNA, knockdown, ncRNA, RNA, Type III, CRISPR, Cas, Csm, Cas13, RNAi
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- Genome Editing via LNP-Based Delivery of Efficient and Stable CRISPR-Cas Editors
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- CRISPR CASY COMPOSITIONS AND METHODS OF USE
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- Improved Cas12a Proteins for Accurate and Efficient Genome Editing
- CRISPR-CAS EFFECTOR POLYPEPTIDES AND METHODS OF USE THEREOF
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- Methods Of Use Of Cas12L/CasLambda In Plants
- Type V CRISPR/CAS Effector Proteins for Cleaving ssDNA and Detecting Target DNA
- THERMOSTABLE RNA-GUIDED ENDONUCLEASES AND METHODS OF USE THEREOF (GeoCas9)
- Structure-Guided Methods Of Cas9-Mediated Genome Engineering
- Efficient Site-Specific Integration Of New Genetic Information Into Human Cells
- CRISPR-Cas Effector Polypeptides and Methods of Use Thereof
- Virus-encoded DNA-binding Proteins
- Class 2 CRISPR/Cas COMPOSITIONS AND METHODS OF USE
- Compositions and Methods of Use for Variant Csy4 Endoribonucleases
- Methods and Compositions for Controlling Gene Expression by RNA Processing
