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Simultaneous Genotyping of Human Platelet Antigens by Hot Start Sequence‐Specific Polymerase Chain Reaction with DNA Polymerase AmpliTaq Gold
Author(s) -
Chen DongFeng,
Pastucha Ladislav T.,
Chen HaiYan,
Kadar Janos G.,
Stangel Walter
Publication year - 1997
Publication title -
vox sanguinis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.68
H-Index - 83
eISSN - 1423-0410
pISSN - 0042-9007
DOI - 10.1046/j.1423-0410.1997.7230192.x
Subject(s) - genotyping , polymerase chain reaction , typing , biology , microbiology and biotechnology , gold standard (test) , hot start pcr , polymerase , polymerase chain reaction optimization , gene , genetics , genotype , multiplex polymerase chain reaction , medicine
Objectives: Human platelet alloantigen (HPA) typing has potential clinical relevance in a variety of contexts. We can improve methods for HPA genotyping by complementing our knowledge of the DNA sequence polymorphisms of HPA genes and experience with various DNA‐based HPA typing techniques. Methods: A newly available DNA polymerase, AmpliTaq Gold (Perkin Elmer), provided in an inactive state and activated by heat, makes it possible to perform a hot start polymerase chain reaction (PCR) in order to prevent nonspecific amplification during the setup of PCR. To establish a practical procedure for HPA‐1, 2, 3 and 5 genotyping, we applied the AmpliTaq Gold for a hot start PCR and employed 8 pairs of published sequence‐specific primers (SSP). A simple simultaneous genotyping of these 4 HPA systems could be rapidly achieved with high specificity. Results: The HPA gene frequencies observed in 126 randomly selected German blood donors were 0.82 and 0.18 for HPA‐1a and 1b, 0.92 and 0.08 for HPA‐2a and 2b, 0.63 and 0.37 for HPA‐3a and 3b and 0.90 and 0.10 for HPA‐5a and Sb, respectively. Conclusion: Using our hot start PCR‐SSP procedure with AmpliTaq Gold a simple, rapid and reproducible genotyping for HPA‐1, 2, 3 and S systems could be achieved.