RFC1repeat expansion analysis from whole genome sequencing data simplifies screening and increases diagnostic rates

Biallelic expansions and a motif change in RFC1 are a common cause of cerebellar ataxia, neuropathy, and vestibular areflexia syndrome. Molecular diagnosis relies on a complicated combination of repeat primed PCR and Southern blotting. We developed a whole genome sequencing based method for RFC1 repeat detection. The combination of sequence motifs and allele length analysis in 29,478 individuals showed that 92.5% of samples have no expanded allele or have expansions of known benign motifs on one allele. In total, 103 samples were classified as biallelic carriers of pathogenic expansions and the frequency of the most common pathogenic allele AAGGG was estimated as 3.9%. Our RFC1 classification method was validated by molecular diagnostic methods achieving sensitivity and specificity of 100% and 97.4%, respectively. Additionally, we catalogued 8 rare repeat motifs, further elucidating the high sequence complexity within the RFC1 locus. In this study we report a new method to identify patients carrying RFC1 pathogenic repeat expansions from WGS data, reducing the need for the inefficient molecular workflow currently used for clinical diagnoses. This method correctly identifies recessive pathogenic repeat expansions in RFC1 and allows screening of existing large scale WGS datasets of unsolved cases.