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Targeted genome fragmentation with CRISPR/Cas9 improves hybridization capture, reduces PCR bias, and enables efficient high-accuracy sequencing of small targets

By Daniela Nachmanson, Shenyi Lian, Elizabeth K. Schmidt, Michael J. Hipp, Kathryn T. Baker, Yuezheng Zhang, Maria Tretiakova, Kaitlyn Loubet-Senear, Brendan F. Kohrn, Jesse J. Salk, Scott R Kennedy, Rosa Ana Risques

Posted 21 Oct 2017
bioRxiv DOI: 10.1101/207027 (published DOI: 10.1101/gr.235291.118)

Current next-generation sequencing techniques suffer from inefficient target enrichment and frequent errors. To address these issues, we have developed a targeted genome fragmentation approach based on CRISPR/Cas9 digestion. By designing all fragments to similar lengths, regions of interest can be size-selected prior to library preparation, increasing hybridization capture efficiency. Additionally, homogenous length fragments reduce PCR bias and maximize read usability. We combine this novel target enrichment approach with ultra-accurate Duplex Sequencing. The result, termed CRISPR-DS, is a robust targeted sequencing technique that overcomes the inherent challenges of small target enrichment and enables the detection of ultra-low frequency mutations with small DNA inputs.

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