Whole Exome Sequencing to Identify Genetic Variants

Whole exome sequencing was performed on eight parent-patient trios (families 1–6). Genomic DNA was captured using the SureSelect XT Human All Exon V5 capture library (Agilent Technologies, Santa Clara, CA, USA), and sequenced using the Illumina HiSeq 2000 (Illumina, San Diego, CA, USA) with 100 bp paired-end reads. Exome data processing, variant calling and variant annotation were performed as described previously^{35 (link), 36 (link)}. Multiplex targeted sequencing analysis was performed on one additional patient (patient 7.1). Amplicon libraries corresponding to the 44 exons of the EPG5 gene were prepared with an Ion AmpliSeq Custom Panel (Thermo Fisher Scientific, Waltham, MA, USA), and sequenced with an Ion Torrent Personal Genome Machine (PGM) system^{37 (link)}. Sequence data was analyzed using a CLC Genomics Workbench 7.0 (CLC bio, Aarhus, Denmark) and Ion reporter (Thermo Fisher Scientific). Identified mutations were validated in all nine patients by Sanger sequencing. The breakpoints in patient 7.1 were PCR amplified and sequenced. The ClinVar accession numbers for the DNA variants reported in this paper are SCV000328413, SCV000328414, SCV000328415, SCV000328416, SCV000328417, SCV000328418, and SCV000328419 (ClinVar, https://www.ncbi.nlm.nih.gov/clinvar/).

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Hori I., Otomo T., Nakashima M., Miya F., Negishi Y., Shiraishi H., Nonoda Y., Magara S., Tohyama J., Okamoto N., Kumagai T., Shimoda K., Yukitake Y., Kajikawa D., Morio T., Hattori A., Nakagawa M., Ando N., Nishino I., Kato M., Tsunoda T., Saitsu H., Kanemura Y., Yamasaki M., Kosaki K., Matsumoto N., Yoshimori T, & Saitoh S. (2017). Defects in autophagosome-lysosome fusion underlie Vici syndrome, a neurodevelopmental disorder with multisystem involvement. Scientific Reports, 7, 3552.

Publication 2017

SureSelect XT Human All Exon V5 capture library HiSeq 2000 Ion AmpliSeq Custom Panel CLC Genomics Workbench 7.0 Ion reporter

Clc 7 Exome Exons Gene Genome Human Library Mutations Parent Patient Trios

Corresponding Organization :

Other organizations : Nagoya City University, Osaka University, Yokohama City University, Tokyo Medical and Dental University, Hokkaido University, Kitasato University, Nishi Niigata Chuo National Hospital, Osaka Medical Center for Cancer and Cardiovascular Diseases, Wakayama Medical University, The University of Tokyo, Ibaraki Children's Hospital, University of Tsukuba, National Center of Neurology and Psychiatry, Showa University, RIKEN Center for Integrative Medical Sciences, Hamamatsu University School of Medicine, Osaka National Hospital, Takatsuki General Hospital, Keio University

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Variable analysis

independent variables

Whole exome sequencing was performed on eight parent-patient trios (families 1–6)
Multiplex targeted sequencing analysis was performed on one additional patient (patient 7.1)

dependent variables

Exome data processing, variant calling and variant annotation

control variables

Genomic DNA was captured using the SureSelect XT Human All Exon V5 capture library (Agilent Technologies, Santa Clara, CA, USA)
Sequenced using the Illumina HiSeq 2000 (Illumina, San Diego, CA, USA) with 100 bp paired-end reads
Amplicon libraries corresponding to the 44 exons of the EPG5 gene were prepared with an Ion AmpliSeq Custom Panel (Thermo Fisher Scientific, Waltham, MA, USA)
Sequence data was analyzed using a CLC Genomics Workbench 7.0 (CLC bio, Aarhus, Denmark) and Ion reporter (Thermo Fisher Scientific)

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