Multiplex Genotyping Developed for Rare Blood Types

All existing sequence-specific primers polymerase chain reaction (PCR-SSP) assays for blood typing, genotype one or several single-nucleotide polymorphisms (SNPs) individually, but DNA pooling is a way that reduces the amount of genotyping required.

Scientists at the Shanghai Blood Center (China) established a sensitive multiplex PCR-SSP assay testing pooled DNA to detect the rare Fyb and S alleles. It was applied to screen a total of 4,490 donor samples via testing of 898 DNA pools. The samples in the positive pools were further tested individually. Genomic DNA was extracted from ethylenediaminetetraacetate (EDTA) anticoagulated blood samples using the TIANamp Genomic DNA Kit (Tiangen Biotech; Beijing, China). Every five DNA samples of the 4,490 donors screened were pooled before the multiplex PCR.

Two hundred and fifty-four donors were tested positive for the Duffy Fyb allele, whereas 101 donors were positive for the S allele. Among the 254 Fy(b+) donors, five were Fy(a−b+) and 249 were Fy(a+b+). Among the 101 S+ donors, three were S+s− and 98 were S+s+. The rare Fyb and S alleles comprised 2.28% and 1.16%, respectively. The PCR-SSP assays were confirmed by sequencing analysis and serological testing.

The authors concluded that this genetic high-throughput screening provides a means of detecting relatively large numbers of donors while providing, to a certain extent, antigen-negative blood for “difficult” patients with rare antibodies. The multiplex PCR assay was combined with DNA pooling to reduce the number of tests required, making large-scale screening feasible, and a cost-effective strategy. The study was published on January 22, 2013, in the journal Transfusion Medicine.

Related Links:
Shanghai Blood Center
Tiangen Biotech