MUTE-Seq: An Ultrasensitive Method for Detecting Low-Frequency Mutations in cfDNA With Engineered Advanced-Fidelity FnCas9.

ABSTRACT: In this study, we present the development of the Mutation tagging by CRISPR-based Ultra-precise Targeted Elimination in Sequencing (MUTE-Seq) method. We engineered a highly precise advanced-fidelity FnCas9 variant, named FnCas9-AF2, to effectively discriminate single-base mismatches at all positions of the single guide RNA (sgRNA) target sequences. FnCas9-AF2 exhibited significantly lower off-target effects compared to existing high-fidelity CRISPR-Cas9 variants. MUTE-Seq leverages FnCas9-AF2 for the enrichment of mutant DNA through the exclusive cleavage of perfectly matched wild-type DNA, allowing for sensitive detection of low-frequency cancer-associated mutant alleles. MUTE-Seq enabled sensitive monitoring of minimal residual disease (MRD) from the bone marrow of patients with Acute My

SUBMITTER: Ye S 

PROVIDER: S-EPMC12651102 | biostudies-literature | 2025 Nov

REPOSITORIES: biostudies-literature