序列特异引物聚合酶链反应在HLA-B27检测中的应用

摘要

目的 探讨序列特异引物聚合酶链反应(PCR-SSP)方法在人类白细胞抗原B27检测中的应用.方法 对经流式细胞术分析定型的279例疑似强直性脊柱炎患者样本采用PCR-SSP法检测HLA-B27的表达.以流式细胞术检测结果为判断标准,对结果不一致样本,进一步应用PCR-直接测序法进行B27测序分析.结果 与流式细胞术结果相比,只有1例结果不一致.经测序分析,其碱基序列与PCR-SSP法特异性引物3 '末端错配;且发现该样本序列存在多个杂合碱基,经与HLA-B27等位基因数据库比对,该杂合序列与最相近B* 2733亚型存在2个碱基差异.结论 PCR-SSP法可适用于HLA-B27的临床检测.%Objective To investigate the effect of polymerase chain reaction using sequence-specific primers (PCR-SSP) in detecting Human leukocyte antigen B27 (HLA-B27).Methods Peripheral blood samples from 279 cases of suspected patients with ankylosing spondylitis (AS) were collected,which had been identified by flow cytometry (FCM).Genomic DNA was analyzed by PCR-SSP.Comparing with the results of flow cytometry,the discrepancies were then analyzed by the sequence-based typing method.Results Except for one sample,the results of PCR-SSP were in agreement with flow cytometry.The sequencing results of the discrepancy showed its nucleotide sequence mismatch with the 3'end of the specific primer and several heterozygous nucleotides in exon 2 and 3.In alignment with HLA Alleles database,there are 2 nucleotide changes from B * 2733 in heterozygous sequences.Conclusion The PCR-SSP technique is applicable for the clinical detection of HLA-B27.