Molecular characterization and pathology of the patient’s novel RPS19 mutation. (A) The patient’s 2012 and 2018 chromosome 19 single nucleotide polymorphism genotype: the upper pattern is the B allele frequency and the lower pattern is the log R ratio. Red arrows indicate the B allele frequency splitting widening over time; blue arrows indicate no change in intensity of the total alleles. (B) Identification and visualization of RPS19 transcripts using the Integrative Genome Viewer. De novo transcript assembly identified 2 transcripts that mapped to the RPS19 gene locus (MSTRG.7258.1 and MSTRG.7258.2) and are shown below the reference RPS19 locus. Of note, although MSTRG.7258.1 resembles the reference transcripts, MSTRG.7258.2 lacks exon 4 (red box). (C) The normalized FPKM values of the indicated RPS19 transcripts are shown for all samples. MSTRG.7258.1, the WT transcript, was expressed in all samples, but MSTRG.7258.2 was expressed in the patient sample. (D) Patient and control complementary DNA transcripts were amplified using primers in exon 3 and exon 5 of the RPS19 gene. The red arrow indicates the WT transcript; the blue arrow the truncated transcript skipping exon 4 found in the patient but not control. Results were validated by Sanger sequencing. (E) 40s and 60s ribosomal subunit abundance determined by the areas under the peaks to calculate 40s/60s abundance ratios. Bars indicate standard deviation (n = 3). *P < .05. FPKM, fragments per kilobase per million.