US 12,281,303 B2
Methods and compositions for prime editing nucleotide sequences
David R. Liu, Cambridge, MA (US); Andrew Vito Anzalone, Cambridge, MA (US); and James William Nelson, Cambridge, MA (US)
Assigned to The Broad Institute, Inc., Cambridge, MA (US); and President and Fellows of Harvard College, Cambridge, MA (US)
Filed by The Broad Institute, Inc., Cambridge, MA (US); and President and Fellows of Harvard College, Cambridge, MA (US)
Filed on May 31, 2023, as Appl. No. 18/326,588.
Application 18/326,588 is a continuation of application No. 18/064,738, filed on Dec. 12, 2022.
Application 18/064,738 is a continuation of application No. 17/219,635, filed on Mar. 31, 2021, granted, now 11,643,652.
Application 17/219,635 is a continuation of application No. PCT/US2020/023730, filed on Mar. 19, 2020.
Claims priority of provisional application 63/100,548, filed on Mar. 17, 2020.
Claims priority of provisional application 62/991,069, filed on Mar. 17, 2020.
Claims priority of provisional application 62/944,231, filed on Dec. 5, 2019.
Claims priority of provisional application 62/974,537, filed on Dec. 5, 2019.
Claims priority of provisional application 62/931,195, filed on Nov. 5, 2019.
Claims priority of provisional application 62/913,553, filed on Oct. 10, 2019.
Claims priority of provisional application 62/973,558, filed on Oct. 10, 2019.
Claims priority of provisional application 62/889,996, filed on Aug. 21, 2019.
Claims priority of provisional application 62/922,654, filed on Aug. 21, 2019.
Claims priority of provisional application 62/858,958, filed on Jun. 7, 2019.
Claims priority of provisional application 62/820,813, filed on Mar. 19, 2019.
Prior Publication US 2023/0340465 A1, Oct. 26, 2023
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 15/11 (2006.01); C07K 14/00 (2006.01); C12N 9/12 (2006.01); C12N 9/22 (2006.01); C12N 15/62 (2006.01); C12N 15/90 (2006.01)
CPC C12N 15/11 (2013.01) [C07K 14/001 (2013.01); C12N 9/1276 (2013.01); C12N 9/22 (2013.01); C12N 15/111 (2013.01); C12N 15/62 (2013.01); C12N 15/902 (2013.01); C12N 15/907 (2013.01); C12Y 207/07049 (2013.01); C07K 2319/00 (2013.01); C12N 2310/20 (2017.05); C12N 2310/3517 (2013.01); C12N 2310/3519 (2013.01); C12N 2800/80 (2013.01)] 30 Claims
 
1. A prime editing guide RNA (PEgRNA) comprising:
a) a spacer sequence;
b) a gRNA core that is capable of complexing with a nucleic acid programmable DNA binding protein (napDNAbp) configured to generate a cut site in a first strand of a double-stranded DNA sequence; and
c) an RNA extension arm comprising i) a DNA synthesis template comprising an edit template that encodes one or more edits compared to a region downstream of the cut site in the first strand of the double-stranded DNA sequence, and ii) a primer binding site that is complementary to a region upstream of the cut site in the first strand of the double-stranded DNA sequence.