Computational protocol: Identification of a de novo DYNC1H1 mutation via WES according to published guidelines

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Protocol publication

[…] We conducted WES of a proband-parent trio to identify the causal gene. The SureSelect Human All ExonV5 Kit (Agilent) was used for exome capture. The IlluminaHiseq 2500 platform (San Diego, CA) was utilized for genomic DNA sequencing of the proband and her parents in Novogene (Beijing, China).Raw image analyses and base calling were performed using Illumina’s Pipeline (version 1.3.4) with default parameters. Sequence data were aligned to the reference human genome (hg19) using the Burrows-Wheeler Aligner (BWA), and duplicate reads were removed using Picard tools. We used the Genome Analysis ToolKit (GATK) to perform the re-alignment and variation (SNP and Indel) detection. Annovar was utilized to catalogue the detected variations. Then, we filtered variations with a homopolymer length >6 (and synonymous substitutions) or that were common (>0.5%) in dbSNP (, HapMap, and the 1000 Genomes Project ( http://www.1000 Variants that were not present in any of the above databases were considered novel.Given to the characteristics of the pedigree, homozygous, compound heterozygous or de novo variations were considered to be candidate causal variations. [...] We confirmed the candidate causal variations identified via WES and conducted co-segregation analyses among the family. The primers of the candidate variations were designed using Primer 3 ( The genomic DNA was PCR-amplified using Roche Fast Start PCR Master Mix polymerase (Roche Diagnostics Corp, USA). PCR products were sequenced with Applied BiosystemsBigDye terminator sequencing chemistry and then run on an ABI3730xl genetic analyzer according to the manufacturer’s instructions (Applied Biosystems,CA, USA). Sequence analysis was performed with Lasergene software (DNASTAR, Madison, WI, USA). […]

Pipeline specifications

Software tools BWA, Picard, GATK, ANNOVAR, Primer3
Databases dbSNP
Applications WES analysis, qPCR
Organisms Homo sapiens
Diseases Movement Disorders, Nervous System Diseases, Genetic Diseases, Inborn