US 12,467,057 B2
Methods for modifying plastid genomes
Xudong Ye, Chesterfield, MO (US)
Assigned to Monsanto Technology LLC, St. Louis, MO (US)
Filed by Monsanto Technology LLC, St. Louis, MO (US)
Filed on Aug. 16, 2022, as Appl. No. 17/820,172.
Claims priority of provisional application 63/233,921, filed on Aug. 17, 2021.
Prior Publication US 2023/0159944 A1, May 25, 2023
Int. Cl. C12N 15/82 (2006.01); C12N 9/22 (2006.01)
CPC C12N 15/8214 (2013.01) [C12N 9/22 (2013.01)] 22 Claims
 
1. A method of modifying a target plastid sequence comprising at least a portion of a plastid gene, or a sequence complementary to at least a portion of a plastid gene in a plastid genome comprising:
a) introducing a recombinant nucleic acid molecule into a plant cell, wherein the recombinant nucleic acid molecule comprises, from 5′ to 3′:
i) a first homology sequence that is homologous to a first endogenous plastid genome sequence;
ii) a modified target plastid sequence that comprises at least one silent mutation relative to the target plastid sequence, such that the modified target plastid sequence has a sequence identity of less than 70% with the target plastid sequence, while encoding a same or similar amino acid sequence encoded by the plastid gene and such that the modified target plastid sequence comprises at least one silent mutation every 15 nucleotides relative to the target plastid sequence; and
iii) a second homology sequence that is homologous to a second endogenous plastid genome sequence,
wherein the target plastid sequence is located in a plastid genome of the plant cell between the first endogenous plastid genome sequence and the second endogenous plastid genome sequence; and
b) allowing homologous recombination to occur such that the modified target plastid sequence replaces the target plastid sequence in the plastid genome of the plant cell.