Novel regulatory variant in ABO intronic RUNX1 binding site inducing A3 phenotype

Abstract

AbstractBackground and ObjectivesMixed‐field agglutination in ABO phenotyping (A3, B3) has been linked to genetically different blood cell populations such as in chimerism, or to rare variants in either ABO exon 7 or regulatory regions. Clarification of such cases is challenging and would greatly benefit from sequencing technologies that allow resolving full‐gene haplotypes at high resolution.Materials and MethodsWe used long‐read sequencing by Oxford Nanopore Technologies to sequence the entire ABO gene, amplified in two overlapping long‐range PCR fragments, in a blood donor presented with A3B phenotype. Confirmation analyses were carried out by Sanger sequencing and included samples from other family members.ResultsOur data revealed a novel heterozygous g.10924C>A variant on the ABO*A allele located in the transcription factor binding site for RUNX1 in intron 1 (+5.8 kb site). Inheritance was shown by the results of the donor's mother, who shared the novel variant and the anti‐A specific mixed‐field agglutination.ConclusionWe discovered a regulatory variant in the 8‐bp RUNX1 motif of ABO, which extends current knowledge of three other variants affecting the same motif and also leading to A3 or B3 phenotypes. Overall, long‐range PCR combined with nanopore sequencing proved powerful and showed great potential as an emerging strategy for resolving cases with cryptic ABO phenotypes.