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PCR with sequence‐specific primer‐based simultaneous genotyping of human platelet antigen‐1 to ‐13w
Author(s) -
Lyou JauYi,
Chen YingJu,
Hu HuiYu,
Lin JeongShi,
Tzeng ChengHwai
Publication year - 2002
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1046/j.1537-2995.2002.00172.x
Subject(s) - genotyping , typing , loss of heterozygosity , primer (cosmetics) , biology , antigen , genotype , polymerase chain reaction , genetics , gene , allele , chemistry , organic chemistry
BACKGROUND : Accurate human platelet antigen (HPA) typing is important for patients with diagnosis of alloimmune thrombocytopenic syndromes and provision of HPA‐matched blood components for these patients. STUDY DESIGN AND METHODS : Thirteen sequence‐specific primers (SSPs) designed on the basis of known published polymorphisms for HPA‐1 to HPA‐13w, respectively, were employed for simultaneous HPA genotyping. All PCR amplifications were carried out with identical cycling conditions in 96‐well plates containing primer mixtures. A total of 300 blood samples from unrelated volunteer donors in Taiwan were included in the study. RESULTS: All primers had specific amplification products. The typing results were available within 4 hours each time for up to four blood samples tested. Among the 13 HPAs, HPA‐3 had the greatest heterozygosity with a gene frequency of 0.3267, 0.4967, and 0.1767 forHPA‐3a/HPA‐3a, HPA‐3a/HPA‐3b,andHPA‐3b/HPA3‐b,respectively. For the remaining 12 HPAs, the predominance ofa/ahomozygosity was noted for HPA‐1, ‐2, ‐4, ‐5, and ‐6, with a frequency ranging from 0.9200 to 0.9967. The frequency ofa/ahomozygosity was 1.0000 for HPA‐7w to ‐13w, except for HPA‐10w, for which one case was observed to beHPA‐10aw/HPA‐10bwheterozygous. Excluding HPA‐3,b/bhomozygosity was noted in only one case (HPA‐6b/HPA‐6b ). The prevalence rates of HPA‐1 to ‐13w in this study were consistent with previous reports using different methods. CONCLUSION : An extended, streamlined PCR‐SSP protocol for simultaneous genotyping of HPA‐1 to HPA‐13w was established. This allows fast and reliable diagnosis of alloimmune thrombocytopenia, and is readily applicable to large‐scale genetic population studies.