Protocol for high-precision CRISPR-Cas12a-based SNV detection on synthetic DNA, cell line cfDNA models, and liquid
Kavish A V Kohabir1, Jasper Linthorst2, Rob M F Wolthuis3
1Department of Human Genetics, Amsterdam UMC Location Vrije Universiteit Amsterdam, Amsterdam, the Netherlands; Amsterdam Reproduction & Development, Amsterdam, the Netherlands; Imaging and Biomarkers, Cancer Center Amsterdam, Amsterdam, the Netherlands.
Abstract:
CRISPR-based diagnostics (CRISPRdx) offer promising tools for rapid and cost-effective genetic testing, but achieving single-nucleotide fidelity remains a challenge. Here, we present a protocol for high-precision detection of single-nucleotide variants (SNVs) using a Cas12a-based approach. We describe how to apply our publicly available ARTEMIS algorithm to identify targetable SNVs, design optimized CRISPR RNAs (crRNAs), and perform fluorescence-based CRISPRdx assays on synthetic DNA, cell line-derived cell-free DNA (cfDNA), and liquid biopsy samples. For complete details on the use and execution of this protocol, please refer to Kohabir et al.1.
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