Agilent sureselect human all exome v6 kit

DNA was extracted from peripheral blood samples of the proband and his parents using a QIAamp DNA Blood Mini Kit (Qiagen, Hilden, Germany) in accordance with the manufacturer's protocol. Trio‐whole‐exome sequencing was performed using the Agilent SureSelect Human All Exome V6 kit (Agilent Technologies Inc. Mississauga, ON, Canada) on an Illumina Hiseq X Analyzer (Illumina, San Diego, CA) with 150‐bp paired‐end runs.

Whole‐exome sequencing was performed on the proband. Genomic DNA was extracted from peripheral blood leukocytes (3–5 ml) using an Agilent SureSelect Human All Exome V6 Kit (Agilent Technologies, Santa Clara, CA, USA), according to the manufacturer's instructions. After DNA extraction, the products were sequenced on a NovaSeq 6000 sequencer (Illumina, San Diego, CA, USA), with a reading length of 150 bp. The NovaSeq 6000 platform was used to examine >99% of the target regions. Sequence alignment was performed according to GRCh37/UCSC hg19. The variants were classified into five categories according to the American College of Medical Genetics and Genomics guidelines for the interpretation of genetic variants:¹⁰ pathogenic, likely pathogenic, uncertain significance, likely benign, and benign. The genetic variants were filtered using population databases such as the 1000 Genomes Project (1000G) (http://www.internationalgenome.org/), ESP6500, dbSNP, ClinVar, Exome Aggregation Consortium, and Human Gene Mutation Database. Furthermore, we studied evolutionary conservation. Sanger sequencing was used to validate suspected variants in the parents.