A mutation creating an alternative splice site in case 6. (A) Normal splicing of HLA-B*40:02. (B) The C>G mutation in intron 3 created an alternative 5′ splicing site with a strong consensus sequence, GTGAGT, which can change the splicing of HLA-B*40:02, leading to a stop codon formation. (C) mRNA from 3 different leukocyte subsets was reverse-transcribed and amplified with alternatively spliced mRNA-specific primers. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA was amplified as an internal control. (D) The expression level of HLA-B4002 by B4002⁻A⁺ granulocytes of case 6 (i) was higher than that by B4002⁻A⁺ granulocytes of case 12 (ii), which lost HLA-B4002 because of nonsense and frameshift mutations. gDNA, genomic DNA.