	US 12,110,517 B2
	Optimized protein linkers and methods of use
Sharon Leigh Guffy, Chapel Hill, NC (US); and Joseph Matthew Watts, Cary, NC (US)
Assigned to Pairwise Plants Services, Inc., Durham, NC (US)
Filed by Pairwise Plants Services, Inc., Durham, NC (US)
Filed on Jun. 14, 2023, as Appl. No. 18/334,407.
Application 18/334,407 is a continuation of application No. 17/381,465, filed on Jul. 21, 2021, granted, now 11,718,838.
Claims priority of provisional application 63/054,449, filed on Jul. 21, 2020.
Prior Publication US 2024/0002821 A1, Jan. 4, 2024
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 9/22 (2006.01); C07K 14/00 (2006.01); C12N 9/78 (2006.01); C12N 15/11 (2006.01); C12N 15/90 (2006.01)

CPC C12N 9/22 (2013.01) [C07K 14/001 (2013.01); C12N 9/78 (2013.01); C12N 15/11 (2013.01); C12N 15/907 (2013.01); C12Y 305/04002 (2013.01); C07K 2319/00 (2013.01); C12N 2310/20 (2017.05)]

22 Claims

1. A Type V Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated (Cas) (CRISPR-Cas) system comprising:

(a) a fusion protein comprising a Cas12a domain, a linker comprising an amino acid sequence of any one of SEQ ID NOs: 1-24, and a polypeptide of interest, wherein the Cas12a domain is linked to the polypeptide of interest via any one of the amino acid sequences of SEQ ID NOs: 1-24, or a nucleic acid encoding the fusion protein; and

(b) a guide nucleic acid comprising a spacer sequence and a repeat sequence, wherein the guide nucleic acid is capable of forming a complex with the Cas12a domain of the fusion protein and the spacer sequence is capable of hybridizing to a target nucleic acid, thereby guiding the Cas12a domain and the polypeptide of interest to the target nucleic acid, whereby the system is capable of modifying or modulating the target nucleic acid.