Genomic DNA was extracted from peripheral blood leukocytes using the QIAamp DNA blood kit according to the manufacturer’s protocol. A total of 10 ng DNA per sample was used for sequencing using the CH capture panel, which was designed based on the Illumina Truseq Custom Amplicon v1.5 kit. Thirteen pathogenic genes (DUOX2, TG, TPO, TSHR, TTF1, TTF2, PAX8, NKX2-5, GNAS, THRA, TSHB, IYD, and SLC5A5) were screened in all patients, including the entire coding regions and exon-intron boundaries. The genetic fragments were between 250 and 280 bp and were prepared using the Covaris and Agencourt AMPure XP kits, which include purified and captured gene fragments. Adaptor-ligated amplicons were prepared using the Illumina Paired-End Sample Preparation kit. Illumina multi-PE-adaptors were bound to terminal genes and target enrichment was performed by multiplex PCR. After 12 PCR cycles, amplicons were purified using Agencourt AMpurc SPRI XP beads and captured on the Illumina MiSeq 2000 instrument.
Ampurc spri xp beads
Manufactured by Illumina
AMpure SPRI XP beads are a paramagnetic bead-based reagent used for the purification and size-selection of DNA and RNA samples. The beads bind to nucleic acids, allowing for efficient removal of unwanted contaminants and size-selection through controlled binding and elution conditions.
Automatically generated - may contain errors
2 protocols using ampurc spri xp beads
1
Targeted Sequencing of Thyroid-related Genes
2
Targeted Illumina Sequencing of Pathogenic Genes
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Genomic DNA was extracted from peripheral blood leukocytes using the QIAamp DNA blood kit according to the manufacturer's protocol. A total of 10 ng DNA per sample was used for sequencing using the CH capture panel, which was designed base on the Illumina Truseq Custom Amplicon v1.5 kit. Thirteen pathogenic genes were screened in all patients, including the entire coding regions and exon-intron boundaries. The genetic fragments were between 250-280 bp and were prepared using the Covaris and Agencourt AMPure XP kits, which include purified and captured gene fragments. Adaptor-ligated amplicons were prepared using the Illumina Paired-End Sample Preparation kit. Illumina multi-PE-adaptors were bound to terminal genes and target enrichment was performed by multiplex PCR. After 12 PCR cycles, amplicons were purified using Agencourt AMpurc SPRI XP beads and captured on an Illumina MiSeq 2000 instrument.
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