Fig. 4: Examples of four samples for three loci in the discovery study.

For a–c, the conventional study results are shown on the left and the results of our diagnostic method using GridION are shown on the right. Posi positive control, Nega negative control, NTC no template control, RP-PCR repeat-primed PCR. a Upper left panel shows the flanking PCR result for the CACNA1A locus. Patient 13 and six other patients (1–6) were tested using a 2% agarose gel and only the patient labeled “2” was judged as positive. Upper right panel shows T-LRS results detecting CACNA1A as the rank #1 locus. Lower right panel shows the confirmatory flanking PCR and fragment analysis of the CACNA1A locus. Flanking PCR was evaluated on a 2.5% agarose gel. b Left panel shows flanking PCRs result for the ATXN8OS/ATXN8 locus. Right panel shows the T-LRS result detecting ATXN8OS/ATXN8 as the rank #1 locus but with ambiguous pathogenicity. The rank #2 locus was also rejected for pathogenicity. c Left panel shows an abnormally expanded PCR amplicon in BEAN1, RP-PCR detecting a pathogenic TGGAA repeat, and Sanger sequencing for an SCA31-linked SNP in which one of the patients (Patient 14) did not have this SNP. Right panel shows the T-LRS result detecting the rank #1 locus as BEAN1 and SCA31-linked SNP genotyping shown in the integrative genomics viewer. The result was matched between the two methods, but multiple experiments were needed with the conventional method.