US 12,275,966 B2
Genetically engineered cells
Jennifer A. Doudna, Berkeley, CA (US); Basem Al-Shayeb, Berkeley, CA (US); Jillian F. Banfield, Berkeley, CA (US); and Patrick Pausch, Berkeley, CA (US)
Assigned to The Regents of the University of California, Oakland, CA (US)
Filed by The Regents of the University of California, Oakland, CA (US)
Filed on May 16, 2022, as Appl. No. 17/745,630.
Application 17/745,630 is a continuation of application No. 17/225,878, filed on Apr. 8, 2021, granted, now 11,377,646.
Application 17/225,878 is a continuation in part of application No. PCT/US2020/021213, filed on Mar. 5, 2020.
Claims priority of provisional application 62/948,470, filed on Dec. 16, 2019.
Claims priority of provisional application 62/907,422, filed on Sep. 27, 2019.
Claims priority of provisional application 62/855,739, filed on May 31, 2019.
Claims priority of provisional application 62/815,173, filed on Mar. 7, 2019.
Prior Publication US 2022/0340889 A1, Oct. 27, 2022
Int. Cl. C12N 9/22 (2006.01); A61K 31/7088 (2006.01); A61K 38/46 (2006.01); C12N 15/10 (2006.01); C12N 15/11 (2006.01); C12N 15/90 (2006.01); C12Q 1/6818 (2018.01); A61K 38/00 (2006.01); A61K 48/00 (2006.01)
CPC C12N 9/22 (2013.01) [A61K 31/7088 (2013.01); A61K 38/465 (2013.01); C12N 15/102 (2013.01); C12N 15/11 (2013.01); C12N 15/111 (2013.01); C12N 15/907 (2013.01); C12Q 1/6818 (2013.01); A61K 38/00 (2013.01); A61K 48/00 (2013.01); C07K 2319/02 (2013.01); C07K 2319/09 (2013.01); C12N 2310/20 (2017.05); C12N 2800/80 (2013.01)] 23 Claims
 
1. A eukaryotic cell comprising:
a) a nuclease comprising an amino acid sequence that is at least 99% identical to the amino acid sequence set forth in SEQ ID NO: 115;
b) a target nucleic acid comprising a target strand (TS) and a non-target strand (NTS); and
c) a recombinant guide nucleic acid comprising in a 5′ to 3′ orientation:
i) a first region that binds to the nuclease; and
ii) a second region that is complementary to a portion of the TS, wherein the target nucleic acid comprises a protospacer adjacent motif (PAM), wherein the PAM is on the NTS of the target nucleic acid, and wherein the PAM comprises the sequence 5′-VTTN-3′, wherein T is thymine, N is any nucleotide, and V is selected from adenine, guanine, and cytosine.