Computational protocol: New perspective in diagnostics of mitochondrial disorders: two years’ experience with whole-exome sequencing at a national paediatric centre

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Protocol publication

[…] WES was performed using TruSeqExome Enrichment Kits according to the manufacturer’s instructions (Illumina). The samples were run on 1/4 of a lane on HiSeq 1500 using 2 × 100 bp paired-end reads. Bioinformatics analysis was performed as previously described []. Briefly, after initial processing with CASAVA, the sequencing reads were aligned to the hg19 reference genome with the Burrows-Wheeler Alignment Tool and further processed by Genome Analysis Toolkit []. Base quality score recalibration, indel realignment, duplicate removal, and SNP/INDEL calling were done as described []. The detected variants were annotated using Annovar and converted to MS Access format for final manual analyses. Alignments were viewed with Integrative Genomics Viewer [, ]. The complete results of WES, including VCF and/or FASTQ files, are available on demand to qualified researchers. All samples were sequenced so that min. 80 % of target was covered 20× or more.The presence of the variants identified by WES was confirmed by Sanger sequencing. […]

Pipeline specifications

Software tools BaseSpace, GATK, ANNOVAR, IGV
Application WES analysis
Organisms Homo sapiens