Abstract: Objective Design of a real-time PCR method for rapid screening of rare Di(b-) blood types inspired by 3 clinical cases. Methods Blood type and unexpected antibodies were identified by serological methods. The coding region of SLC4A1 gene was amplified by PCR and sequenced to analyze the DNA sequence. Real-time TaqMan-MGB PCR was used for rapid identification of DI*A/DI*B from clinical cases and control samples. Results The unexpected antibody identification of 3 clinical cases was uniformly positive with RBCs, suggesting the presence of unexpected antibodies against high-frequency antigens. DNA sequencing confirmed that the 3 cases were TT at 2561bp in exon 19 of the SLC4A1 gene, which is rare Di(b-) blood type. Real-time fluorescent PCR primers and probes designed for rapid screening of matching blood can accurately identify the SLC4A1 genotype of samples. Conclusion The real-time fluorescent TaqMan-MGB PCR method we designed can quickly, accurately and efficiently identify DI*A and DI*B genes, and can be used to rapidly screen Di(b-) blood to meet the clinical blood needs.

Key words: Rare Di(b-) blood type, SLC4A1, Real-time PCR