Supplementary Material for: Unveiling novel and rare variants in the α-1,4-galactosyltransferase gene leading to rare p phenotype in Indian patients

Introduction:
The rare p phenotype, characterized by the absence of P, P1, and Pk antigens, produces anti-PP1Pk antibody that can cause severe hemolytic reactions and recurrent miscarriages. This phenotype is rare globally but shows notable prevalence in specific populations. This study focuses on the molecular characterization of six Indian patients to provide further insight into the genetic basis of the p phenotype.
Methods:
A targeted Next-Generation Sequencing (tNGS) assay covering 51 genes associated with 41 blood group systems was utilized to investigate the molecular basis of the A4GALT gene in six patients with a serologically confirmed p phenotype. The results were analysed and annotated using bioinformatics tools, including the Integrative Genomics Viewer (IGV), with novel variants confirmed by Sanger sequencing. Family members of each patient were also screened to identify potential rare donors.
Results:
Genomic analysis revealed novel and rare variants in the A4GALT gene, all confirming the p phenotype. Five frameshift mutations (c.72dupC, c.218delG, c.592delC, c.972_997del and c.547_548del) and one nonsense mutation (c.C392G) were detected, resulting in truncated, non-functional proteins. The p phenotype was confirmed in a subset of family members, identifying three new donors for rare blood transfusion requirements.
conclusion:
This study presents the molecular characterization of the p phenotype in Indian patients, identifying novel variants not yet registered in the ISBT database. These findings enhance understanding of the p phenotype and highlight the significance of family screening for identifying rare blood donors. The study underscores the critical need for rare donor registries, especially for patients at high risk of severe hemolytic reactions during transfusion.