A novel homozygous splice‐site mutation of JK gene leads to Jk(a‐b‐) phenotype

Abstract Objectives This study aimed to investigate the molecular mechanism of the Jk(a‐b‐) phenotype in a Chinese transfusion patient. Background Many different mutation types relating to Jk(a‐b‐) phenotype have been reported. However, the splice‐site mutation is relatively rare and the related functional verification is lacking. Materials and Methods In this study, the blood sample was collected from a transfusion patient with the Jk(a‐b‐) phenotype. Serotyping was performed using routine serological methods. The exons sequences and coding regions of the JK gene were amplified using polymerase chain reaction and directly sequenced. To perform a minigene splicing assay, the intronic mutation sequences were cloned into a pSPL3 splice reporting vector. The splicing reporter minigene assay was performed in HEK 293T cells. Results The Jk(a‐b‐) phenotype of the blood sample was identified through serological testing. Sequencing results revealed that the sample had a novel homozygous splice‐site mutation JK*02N (NM_015865.7: c.663+3A>C). Further analysis, including cDNA sequencing and minigene splicing assay, confirmed that the novel splice‐site mutation resulted in exon skipping. Interestingly, different numbers of exons being skipped were obtained by the two methods. Conclusion This study revealed a novel homozygous splicing‐site mutation associated with the Jk(a‐b‐) phenotype in Chinese population. Our results emphasise the importance of the in vitro functional method minigene splicing assay, while also acknowledging its potential limitations when compared to cDNA sequencing.