Simultaneous genotyping of human platelet antigens by hot start sequence-specific polymerase chain reaction with DNA polymerase AmpliTaq Gold.

Autor: Chen DF; Blutbank-Immunhämatologie-Transfusionsmedizin, Medizinische Hochschule Hannover, Deutschland., Pastucha LT, Chen HY, Kadar JG, Stangel W
Jazyk: angličtina
Zdroj: Vox sanguinis [Vox Sang] 1997; Vol. 72 (3), pp. 192-6.
DOI: 10.1046/j.1423-0410.1997.7230192.x
Abstrakt: 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.
Databáze: MEDLINE
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