Abstract Background KEL1 antigen expression is routinely tested in blood donors in Switzerland. A donor sample with an apparent rare KEL:1,‐2 phenotype was genotyped KEL*01.01/KEL*02. A series of detailed molecular analyses were performed to solve this discrepancy, and a novel variant KEL*02 allele was identified. Materials and Methods Standard serological column agglutination and in‐house adsorption‐elution testing were used for the detection of KEL antigens. Genomic DNA was isolated and analyzed by commercial sequence‐specific primer (SSP)‐PCR, in‐house multiplex SSP‐PCR, and KEL exon sequencing. Total RNA was isolated from blood samples; polyadenylated RNA was reverse‐transcribed, and cDNA was amplified with allele‐specific primers and sequenced. SpliceAI and PolyPhen‐2 were used to evaluate the impact of nucleotide variations and amino acid changes on splice effects and protein function, respectively. Results The donor sample was initially typed KEL:1,‐2. However, SSP‐PCR revealed the genotype KEL*01.01/KEL*02 and subsequent adsorption‐elution testing indicated very weak KEL2 expression. Exon sequencing showed the heterozygous missense mutation c.139C>T leading to the amino acid substitution p.(Arg47Trp). PolyPhen‐2 predicted this change to be benign, whereas SpliceAI analysis indicated a putative change in splice sites. Allele‐specific amplification and sequencing revealed that the KEL*02 derived transcript lacks a significant portion of exon 3, causing a frameshift. Conclusion We identified a novel missense mutation c.139C>T in KEL*02 . Although the variant nucleotide locates in the center of exon 3, far away from the exon/intron boundary, it leads to variant splicing of the transcript, resulting in very weak expression of a truncated protein only detectable by adsorption‐elution testing.
Schimanski et al. (Tue,) studied this question.