WES sequencing libraries were prepare following the previous report with slight modifications72 (link).
In brief, genomic DNA was extracted from snap freezing GCA tumor or matched normal tissue using DNeasy Blood & Tissue Kit (69504, QIAGEN) according to the manufacturer’s instruction. DNA degradation and contamination were monitored on 1% agarose gels. DNA concentration was measured by Qubit DNA Assay Kit in Qubit 2.0 Flurometer (Invitrogen). A total amount of 0.6 μg genomic DNA per sample was fragmented to an average size of 180–280 bp and subjected to DNA library preparation using Illumina TruSeq DNA sample preparation kit. The Agilent SureSelect Human All ExonV5 Kit (5190-6209, Agilent Technologies) was used for exome capture according to the manufacturer’s instruction. In brief, DNA libraries were hybridized with liquid phase with biotin labeled probes from the Agilent SureSelect Human All ExonV5 Kit, then magnetic streptavidin beads were used to capture the exons of genes. Captured DNA fragments were enriched in a PCR reaction with index barcodes for sequencing. Final libraries were purified using AMPure XP beads (A63880, Beckman Coulter) and quantified using the Agilent high sensitivity DNA kit (5067-4626, Agilent Technologies). WES libraries were sequenced on Illumina Novaseq 6000 (Illumina) with 150 bp paired end mode according to the manufacturer instruction.
In brief, genomic DNA was extracted from snap freezing GCA tumor or matched normal tissue using DNeasy Blood & Tissue Kit (69504, QIAGEN) according to the manufacturer’s instruction. DNA degradation and contamination were monitored on 1% agarose gels. DNA concentration was measured by Qubit DNA Assay Kit in Qubit 2.0 Flurometer (Invitrogen). A total amount of 0.6 μg genomic DNA per sample was fragmented to an average size of 180–280 bp and subjected to DNA library preparation using Illumina TruSeq DNA sample preparation kit. The Agilent SureSelect Human All ExonV5 Kit (5190-6209, Agilent Technologies) was used for exome capture according to the manufacturer’s instruction. In brief, DNA libraries were hybridized with liquid phase with biotin labeled probes from the Agilent SureSelect Human All ExonV5 Kit, then magnetic streptavidin beads were used to capture the exons of genes. Captured DNA fragments were enriched in a PCR reaction with index barcodes for sequencing. Final libraries were purified using AMPure XP beads (A63880, Beckman Coulter) and quantified using the Agilent high sensitivity DNA kit (5067-4626, Agilent Technologies). WES libraries were sequenced on Illumina Novaseq 6000 (Illumina) with 150 bp paired end mode according to the manufacturer instruction.
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