Computational protocol: Whole exome sequencing reveals mutations in NARS2 and PARS2, encoding the mitochondrial asparaginyl-tRNA synthetase and prolyl-tRNA synthetase, in patients with Alpers syndrome

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Protocol publication

[…] Whole exome sequencing (WES) was carried out by BGI Hong Kong Co., Limited (Hong Kong, China), using Illumina HiSeq 2000 high-throughput sequencing technology. Agilent (Santa Clara, CA) SureSelect Human All Exon 51M kit was used for exome capture.Quality assessment of the sequence reads was performed by generating QC statistics with FastQC (http://www.bioinformatics.bbsrc.ac.uk/projects/fastqc). Read alignment to the reference human genome (hg19, UCSC assembly, February 2009) was done using BWA (Burrows–Wheeler alignment) with default parameters (Li and Durbin ). A summary of the sequencing data is shown in Table. After removal of PCR duplicates using Picard tools (http://picard.sourceforge.net) and file conversion using SAMtools (Li et al. ) quality score recalibration, indel realignment and variant calling were performed with the Genome Analysis Toolkit package (McKenna et al. ). For the identification of potentially pathogenic variants, we used Ingenuity ® Variant Analysis ™ software (http://www.ingenuity.com/variants), from Ingenuity Systems. All mutations identified by WES were verified by Sanger sequencing using primers amplifying the exons harboring the mutations in NARS2 and PARS2. Sequencing analysis was performed using an ABI PRISM ® 3100 Genetic analyzer and the BigDye Terminator v.1.1 Cycle Sequencing Kit (Applied Biosystems Foster City, CA). Primer sequences and PCR conditions are available upon request. […]

Pipeline specifications

Software tools FastQC, BWA, Picard, SAMtools, GATK, Ingenuity Variant Analysis
Application WES analysis
Organisms Homo sapiens
Diseases Diffuse Cerebral Sclerosis of Schilder, Liver Failure, Neurodegenerative Diseases