case_images: PMC10726134_01
case_images: Peripheral blood smear and family tree of the patient. (A) Spherical-shaped erythrocytes were found in the peripheral blood film. (B), Family tree of a Chinese family with the patient exhibiting hereditary spherocytosis. Square and circle denote male and female, respectively.
case_images: fgene-14-1309040-g001.jpg
case_images: PMC10726134_01_fgene-14-1309040-g001.jpg
case_images: F1
case_images: RBCs showed a disparity in size, and spherical-shaped erythrocytes (7%) were found in the peripheral blood film (Figure 1A). The PCR product only presented in the proband, not in his parents (Figure 1B; Figure 3B). This confirmed that the intronic inversion was a de novo mutation.
case_images: Next-generation sequencing. The intronic inversion mutation located between intron 19 and 20 of the SPTB gene was visualized by the integrative genomic viewer. The discordant reads are labelled green and blue.
case_images: fgene-14-1309040-g002.jpg
case_images: PMC10726134_01_fgene-14-1309040-g002.jpg
case_images: F2
case_images: The inversion variant resided between the 5'-gene breakpoint of intron 19 and 3'-gene breakpoint of intron 20 in the SPTB gene (Figure 2; Figure 3A) that could disrupt the canonical splice site.
case_images: Sanger sequencing validation and a schematic diagram of the intronic inversion mutation in SPTB. (A) Schematic diagram of the SPTB gene. F1, forward primer 1; R1, reverse primer 1; F2, forward primer 2; R2, reverse primer 2. (B) The intronic inversion variant carried by the proband was confirmed using Sanger sequencing. The black and red arrow indicates the 5'-gene breakpoint of intron 19 and 3'-gene breakpoint of intron 20 in the SPTB gene, respectively.
case_images: fgene-14-1309040-g003.jpg
case_images: PMC10726134_01_fgene-14-1309040-g003.jpg
case_images: F3
case_images: The inversion variant resided between the 5'-gene breakpoint of intron 19 and 3'-gene breakpoint of intron 20 in the SPTB gene (Figure 2; Figure 3A) that could disrupt the canonical splice site. The inversion was represented with a schematic diagram (Figure 3A). TGATGCAGAAACAGAAAACC-5'; 5'-gene breakpoint of intron 19 in SPTB: forward primer 2, 3'-GGAGCAAGTTATTTGGAACCTCAG-5' and reverse primer 2, 3'- ACAAAGGAGAAGACCCAGCACC-5' (Figure 3A). The PCR product only presented in the proband, not in his parents (Figure 1B; Figure 3B). This confirmed that the intronic inversion was a de novo mutation.
case_images: RNA sequencing analysis. (A) RNA sequencing showed skipped exon 20 in the SPTB mRNA of the patient, but not the control. (B) Gene-level significance (-log10(P), y-axis) versus Z-score, with SPTB labelled as the expression outlier (red dot) of sample from the proband.
case_images: fgene-14-1309040-g004.jpg
case_images: PMC10726134_01_fgene-14-1309040-g004.jpg
case_images: F4
case_images: RNA sequencing analysis found that the intronic inversion variant was associated with exon 20 skipping in the SPTB mRNA (Figure 4A), and revealed one underexpression outlier, which was the SPTB gene (Figure 4B).