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High-Resolution HLA-Typing by Next-Generation Sequencing of Randomly Fragmented Target DNA

Methods Mol Biol. 2018; 
Michael Wittig, Simonas Juzenas, Melanie Vollstedt, Andre Franke
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Oligo pool … 2. Custom oligo pool of the bait design (see above) (CustomArray). 3. Phusion High-Fidelity PCR Kit (NEB). 4. Ethanol 80% and 70%. 5. Elution buffer or TE 0.1%. 6 … 3.75. 10 μM reverse primer. 3.75. Custom oligo pool from CustomArray (1:100 dilution). 2.5. DMSO. 1.5 … Get A Quote

摘要

PCR- or probe-based targeted capturing enables the enrichment of specific genomic loci prior to Next-Generation Sequencing (NGS). Here, we describe a probe-based protocol, which allows for high-resolution HLA typing of DNA samples by NGS. We also describe existing software tools that can be used for the subsequent HLA data analysis. Key prerequisites that warrant an accurate HLA calling are specific mappings of the sequencing reads, phasing of the mapped reads, and the possibility to perform a manual inspection/curation of the read mapping.

关键词

HLA analysis, HLA typing, In-solution capture, NGS, Next-Generation Sequencing, RNA baits, Sequencing, Targeted enrichment