IP Library › Granted Patent US 9,885,026
Granted Patent B2
US 9,885,026 · App. 14/997,474 · Granted Feb 6, 2018

Modified cascade ribonucleoproteins and uses thereof

Inventors: Stan Johan Jozef Brouns (Wageningen, NL); John Van Der Oost (Renkum, NL)
Assignee: Caribou Biosciences, Inc.
C12N9/22A61K48/005C07K14/245C07K14/47C12N9/16C12N15/62C12N15/66C12N15/70C12N15/74C12N15/81C12N15/82C12N15/86C12N15/902C12N15/907A61K38/00C07K2319/09C07K2319/22C07K2319/60C07K2319/71C07K2319/80C07K2319/85C12N2310/20C12Y301/21004
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Quick Facts
Patent No.
US 9,885,026
App. No.
14/997,474
Filed
Jan 15, 2016
Granted
Feb 6, 2018
Kind
B2
Art Unit
1636
USPC
435/462
Abstract

A clustered regularly interspaced short palindromic repeat (CRISPR)-associated complex for adaptive antiviral defence (Cascade); the Cascade protein complex comprising at least CRISPR-associated protein subunits Cas7, Cas5 and Cas6 which includes at least one subunit with an additional amino acid sequence possessing nucleic acid or chromatin modifying, visualizing, transcription activating or transcription repressing activity. The Cascade complex with additional activity is combined with an RNA molecule to produce a ribonucleoprotein complex. The RNA molecule is selected to have substantial complementarity to a target sequence. Targeted ribonucleoproteins can be used as genetic engineering tools for precise cutting of nucleic acids in homologous recombination, non-homologous end joining, gene modification, gene integration, mutation repair or for their visualization, transcriptional activation or repression. A pair of ribonucleotides fused to FokI dimers may be used to generate double-strand breakages in the DNA to facilitate these applications in a sequence-specific manner.

Claims (8)

1. A method of modifying a target nucleic acid comprising:

contacting the target nucleic acid with a Type I Cascade ribonucleoprotein complex comprising a designed fusion protein comprising a FokI fused to the N-terminus of a Cse1, wherein the designed fusion protein further comprises a linker polypeptide between the FokI and the Cse1, and a CRISPR RNA (crRNA) molecule; and

wherein the modifying is binding and/or cleaving the target nucleic acid.

2. The method of claim 1 , wherein the target nucleic acid is a double-stranded DNA (dsDNA).

3. The method of claim 1 , wherein the Type I Cascade ribonucleoprotein complex further comprises a Type I CRISPR-associated protein subunit selected from the group consisting of Cas6, Cas5, Cse2, and Cas7.

4. The method of claim 1 , wherein the FokI is a KKR Sharkey.

5. The method of claim 1 , wherein the method of modifying a target nucleic acid is carried out in vitro in a cell free environment.

6. The method of claim 1 , wherein the FokI is an ELD Sharkey.

Assignments (2)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Dec 19, 2016
From: VAN DER OOST, JOHN; BROUNS, STAN JOHAN JOZEF
To: WAGENINGEN UNIVERSITEIT
Reel/Frame 041010/0279 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Dec 19, 2016
From: WAGENINGEN UNIVERSITEIT
To: CARIBOU BIOSCIENCES, INC.
Reel/Frame 041010/0327 →
Priority Claims (1)
GB 1122458.1 · Dec 30, 2011 · national
Continuity (3)
Continuation 14326099 · Jul 8, 2014
Continuation 14240735
Related Publication 20160186214A1 · Jun 30, 2016