Background:Spinocerebellar ataxia type 3/Machado-Joseph disease (SCA3/MJD) is a progressive autosomal dominant neurodegenerative disease caused by abnormal CAG repeats in exon 10 of ATXN3. The accumulation of the mutant ataxin3 proteins carrying polyglutamine (polyQ) lead to selective degeneration of neurons. Therapeutic strategies were used to inhibit mutant ATXN3 expression, including antisense oligonucleotides, RNA interference and more recently CRISPR/Cas9 genome-editing based approaches. Since the pathogenesis of SCA3 has not been fully elucidated, and no effective therapies can be used, it is crucial to investigate the pathogenesis and seek new therapeutic strategies of SCA3/MJD. Methods: Here we used the paired sgRNA/Cas9 nickases and Cre-loxP mediated homologous recombination (HR) strategy to precisely modify the abnormal CAG expansions in the ATXN3 of SCA3/MJD patient derived induced pluripotent stem cells (SCA3/MJD-iPSCs). Meanwhile, we investigated the disease related phenotypes in differentiated neurons, including electrophysiological characteristics, IC2-positive aggregations, mitochondrial membrane potentials (MMPs), glutathione (GSH) expressions, intracellular reactive oxygen species (ROS), Ca2+ concentrations and malondialdehyde (MDA) levels. Results: SCA3/MJD-iPSCs can be corrected by the replacement of the abnormal CAG expansions with normal repeats using HR. Besides, corrected SCA3/MJD-iPSCs retained pluripotent and normal karyotype, which could be differentiated into neuron cells (NCs) and maintained electrophysiological characteristics. The expression of differentiation markers and electrophysiological characteristics were similar among the control individuals, SCA3/MJD patients and isogenic control SCA3/MJD groups. Furthermore, this study proved that the phenotypic abnormalities in SCA3/MJD-iPSCs derived NCs, including aggregated polyQ toxic protein, decreased MMPs and GSH expressions, increased ROS, Ca2+ concentrations and MDA levels, all were rescued in the corrected SCA3/MJD-NCs. Conclusion: The present study firstly suggested that the genetically corrected SCA3/MJD-iPSCs and associated phenotypic abnormalities, which will provide an ideal models for molecular mechanism research and autologous stem cell therapy.