title: DNA-Seq of homo sapiens: ovarian carcinoma tissue
design: Oncomine BRCA Research Assay (Thermo Fisher Scientific, Inc) is used to investigate BRCA1 and BRCA2 exons, with an average of 64 bases of intronic flanking sequences upstream and downstream of each exon, according to manufacturer's protocol. Briefly, five m sections from FFPE samples were manually microdissected to isolate the highest percentage of neoplastic cells. Genomic DNA was extracted with protease K (incubation ON at 55C) and quantified with Qubit dsDNA BR kit. The libraries were prepared with the IonAmpliSeq Library kit 2.0 and quantified with Qubit dsDNA HS kit following the manufacturers instructions. The libraries are diluted to 25 pm, pooled and loaded on the Ion Chef to perform emulsion PCR and chip loading. Data were processed using Torrent Suite v5.12. Variant calling from sequencing data was generated by the Variant Caller plugin. To eliminate erroneous base calling, variant coverage has been set >40, variant frequency on each sample >5 and quality value >30. The resulting variants were annotated using both the Ensemble Variant Effect Predictor pipeline and the Ion Reporter analysis software. Each variant is displayed on IGV2.11.1.