PMC7673426_01
Genetic analysis of a large homozygous DOCK8 gene deletion. (A) Multiplex PCR on genomics DNA was performed by co-amplification of DOCK8 exon 1 to exon 5 with a reference gene (CFTR exon 4). Electrophoresis of PCR products in 3% agarose gel indicated the absence of DOCK8 exons 2-4 in the patient. The arrow indicates the PCR product of the reference gene. (B) After several rounds of PCR, primers located in intron 1 (DOCK8-Intron1FB, 5'-GCAGCATTGGCAAAGTGTTCTG-3') and intron 4 (DOCK8-Intron4RC, 5'-CACACTTACAGATGTGTGCTTG-3') successfully amplified the breakpoint region. Sequencing of this PCR product revealed a deletion of 80,133 bp with an insertion of "T." The indel mutation started from chr9:218156 to 298288 (GRCh38.p12), DOCK8 intron 1 position +3,126 to intron 4 position -6,292 (IVS1+3126_IVS4-6292delinsT).
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F1
The patient was born to a 34-year-old primigravida, and the mother and the father are first-degree cousins, hence she was born from a consanguineous union (Supplementary Figure 1). Whole-exome sequencing identified a homozygous large deletion of exons 2-4 in the DOCK8 gene, which was confirmed by multiplex polymerase chain reaction (PCR) on genomic DNA for DOCK8 coding regions (Figure 1A). Sanger sequencing of this product revealed a deletion of 80,133 bp with an insertion of "T" (Figure 1B).
Deletion breakpoint of the patient. (A) Sequence of the 5' breakpoint region, the 3' breakpoint region and the breakpoint fusion sequence in the patient. (B) Analysis of repeat elements in the 2,000 bp flanking each of the breakpoint regions showed short interspersed elements (Alu and SINE2). The 3' breakpoint is located in an Alu sequence, and the 5' breakpoint is not located, but closely adjacent to a SINE2 element. SINE, short interspersed nuclear element.
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F2
Administration of antibiotics improved the symptoms; however, she developed multiple skin-colored papulonodular skin lesions with central umbilication on the entire skin (Supplementary Figure 2A). Within 1 year of follow-up, the patient showed persistent atopic dermatitis, and developed opacification of the left eye, accompanied by dimming of her vision (Supplementary Figure 2B). The 3' breakpoint is located in an Alu transposable element, but the 5' breakpoint is not located in any transposable elements (Figure 2).