Whole Exome sequencing
Genomic DNA was extracted from whole blood for all patients at the time
of evaluation and treatment at the fertility centre. Parental DNA was
obtained from saliva using the Oragene OG-500 kit (DNA Genotek, Ottawa,
Canada). Exomes of patient-parent trios were prepared for sequencing
using 1 µg of high-quality genomic DNA, which was quantified with the
Qubit dsDNA HS kit (Thermo Fisher Scientific, Waltham, MA, USA).
Whole exome enrichment was
performed using Illumina’s Nextera DNA Exome Capture kit (Illumina, San
Diego, CA, USA) or Twist Bioscience’s Twist Human Core Exome Kit, as per
manufacturer protocols. Individual sample libraries were then indexed
with Illumina Nextera DNA UD indexes (Illumina, San Diego, CA, USA)
prior to pooling libraries together. The final pooled library was
sequenced on the NovaSeq 6000 platform (Illumina, San Diego, CA, USA) to
an average coverage depth of 72X (Illumina Nextera kit) or 99X (Twist
Bioscience’s kit).