Genome editing systems comprising repair-modulating enzyme molecules and methods of their use

A method of altering a nucleic acid at a target position in a cell, or a population of cells, the method comprising contacting the cell, or the population of cells, with

• (a) a gRNA molecule;
• (b) a RNA-guided nuclease molecule;
• and
• (c) a heterologous Repair-Modulating Enzyme Molecule (RMEM);
• wherein the gRNA molecule and the RNA-guided nuclease molecule interact with the nucleic acid, resulting in a cleavage event, and wherein the cleavage event is repaired by at least one DNA repair pathway that is modulated by the RMEM, wherein
• i) the RMEM is Rad52 or TdT, and the RMEM suppresses formation of a deletion in the nucleic acid at the target position in the cell, or in the population of cells;
• ii) the RMEM is Artemis, and the RMEM enhances formation of a deletion in the nucleic acid at the target position in the cell, or in the population of cells;
• iii) the RMEM is Rad52, TdT, Rad51, RPA, or ERCC1, and the RMEM suppresses gene conversion of the nucleic acid at the target position in the cell, or in the population of cells;
• iv) the RMEM is TdT, Rad51, or T5 exonuclease, and the RMEM suppresses gene correction of the nucleic acid at the target position in the cell, or in the population of cells;
• v) the RMEM is Rad52 or 53BP1 dominant negative, and the RMEM enhances gene correction of the nucleic acid at the target position in the cell, or in the population of cells;
• vi) the RMEM is 53BP1 dominant negative or T5 exonuclease, and the RMEM suppresses formation of an insertion in the nucleic acid at the target position in the cell, or in the population of cells;
• or
• vii) the RMEM is TdT, and the RMEM enhances formation of an insertion in the nucleic acid at the target position in the cell, or in the population of cells,
• and wherein the formation of a deletion in the nucleic acid after repair is suppressed by at least 0.1-fold, the formation of a deletion the nucleic acid after repair is enhanced by at least 0.1-fold, the gene conversion is suppressed by at least 0.1-fold, the gene conversion is enhanced by at least 0.1-fold, the gene correction is suppressed by at least 0.1-fold, the gene correction is enhanced by at least 5-fold, the formation of an insertion in the nucleic acid after repair is suppressed by at least 0.1-fold, and/or the formation of an insertion in the nucleic acid after repair is enhanced by at least 0.1-fold;
• in the cell or in the population of cells, relative to a cell or a population of cells not contacted with the RMEM,
• thereby altering the nucleic acid at the target position in the cell, or in the population of cells.