CRISPR Detection and Research on Screening Mutant Gene of Moyamoya Disease Family Based on Whole Exome Sequencing.

Moyamoya disease (MMD) has a high incidence in Asian populations and demonstrates some degree of familial clustering. Whole-exome sequencing (WES) is useful in establishing key related genes in familial genetic diseases but is time-consuming and costly. Therefore, exploring a new method will be more effective for the diagnosis of MMD. We identified familial cohorts showing MMD susceptibility and performed WES on 5 affected individuals to identify susceptibility loci, which identified point mutation sites in the titin () gene (rs771533925, rs559712998 and rs72677250). Moreover, mutations were not found in a cohort of 50 sporadic MMD cases. We also analyzed mutation frequencies and used bioinformatic predictions to reveal mutation harmfulness, functions and probabilities of disease correlation, the results showed that rs771533925 and rs72677250 were likely harmful mutations with GO analyses indicating the involvement of in a variety of biological processes related to MMD etiology. CRISPR-Cas12a assays designed to detect mutations provided results consistent with WES analysis, which was further confirmed by Sanger sequencing. This study recognized as a new familial gene marker for moyamoya disease and moreover, demonstrated that CRISPR-Cas12a has the advantages of rapid detection, low cost and simple operation, and has broad prospects in the practical application of rapid detection of MMD mutation sites.