To establish a Drosophila model of MSUD to study disease pathogenesis and develop potential treatments, we first sought to generate BCKDH knockout flies using CRISPR/Cas9-mediated genome editing utilizing homology-dependent repair (HDR), consisting of two guide RNAs and a double-stranded DNA plasmid donor to delete the dDBT gene. Here, PPM1K is linked to maple syrup urine disease.