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Methods and compositions for enhancing nuclease-mediated gene disruption

  • US 10,227,610 B2
  • Filed: 02/24/2014
  • Issued: 03/12/2019
  • Est. Priority Date: 02/25/2013
  • Status: Active Grant
First Claim
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1. A method for targeted genomic disruption via microhomology-mediated end joining (MMEJ) in a cell, the method comprising:

  • administering at least one nuclease to the cell, wherein the nuclease cleaves endogenous genomic sequences in the cell, wherein the nuclease is a zinc finger nuclease, a TALE effector domain nuclease (TALEN) and/or a CRISPR/Cas nuclease system; and

    growing the cell in a medium comprising at least one small molecule inhibitor of a DNA-dependent-protein kinase catalytic subunit (DNA-PKcs) protein and a small molecule inhibitor of a Poly-(ADP-ribose) polymerase 1/2 (PARP1/2) protein at 0.5 to 25 wherein the small molecule inhibitor is a nicotinamide;

    a isoquinolinone and a dihydroisoquinolinones;

    a benzimidazole;

    an indole;

    phthalazin-1(2H)-one;

    a quinazolinone;

    an isoindolinone and analogues and derivatives thereof;

    a phenanthridine;

    a phenanthridinone;

    a benzopyrone and analogues and derivatives thereof;

    an unsaturated hydroximic acid derivative and analogues and derivatives thereof;

    a pyridazine;

    caffeine, theophylline;

    thymidine and/or NU7026 and/or NU7441, wherein the endogenous genomic sequences in the cell are disrupted via MMEJ after cleavage by the at least one nuclease.

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