Defects in the respiratory chain result in human mitochondrial disease and affect ~1/5,000 live births. Onset can be at any age, but severe childhood disease is common and symptoms regularly involve neurological and muscular disease. Defects in respiratory chain Complex I is the most common mitochondrial disease, and results in multi-system disorders. Complex I contains 44 different protein subunits, with 7 subunits encoded by mitochondrial genes and the rest by nuclear genes. Complex I failure leads to defects in aerobic respiration with elevated lactic acid and ketone bodies. Complex I is also known to generate reactive oxygen species, an important contributor to many mitochondrial pathologies. In collaboration with the Murdoch Childrens Research Institute at the Royal Children's Hospital, we analyse cells from patients for defects in assembly of the respiratory chain, in particular Complex I. Using new gene-editing techniques (TALENs and CRISPRs), we are complementing our patient studies with targeted knockouts of specific Complex I subunits and other proteins so we can understand how they help build this important molecular machine.