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目的 采用DNA芯片技术对HLA DR1,DR5 1组基因分型。 方法 根据编码HLA DR1,DR5 1组抗原等位基因序列设计特异分型探针 ,制作DNA分型芯片。通过组间特异引物扩增基因组相应区段DNA ,扩增中用Cy5 dCTP荧光标记 ,扩增标记后的产物与芯片探针杂交 ,通过杂交产生的荧光信号及分布格局确定样品基因亚型。分型结果用标准DNA和PCR SSO验证。 结果 130份样本共检出HLA DR1,DR5 1组位点 34个 ,包括 18个DR15 ,8个DR16 ,6个DR10 ,2个DR1,无假阳性或假阴性 ,准确率和重复率均达 10 0 %。检测总耗时约 3.5h。 结论 DNA芯片用于HLA DR1、DR5 1组基因分型具有高分辨度、高特异性和简单快速的优点 ,适于器官移植临床应用。
Objective To genotype HLA DR1 and DR5 1 using DNA chip technology. Methods Based on the HLA DR1 and DR5 1 antigen alleles, a specific typing probe was designed to make DNA typing chips. Genomic DNA was amplified by using specific primers between two groups. Cy5 dCTP was used to amplify the amplified DNA. Then the amplified products were hybridized with the chip probe. The fluorescence signal and the distribution pattern of the hybridized DNA were used to determine the genotype of the sample. Typing results were validated using standard DNA and PCR SSO. Results A total of 130 HLA DR1 and DR5 loci were detected in 130 samples, including 18 DR15, 8 DR16, 6 DR10 and 2 DR1 without any false positive or false negative, with the accuracy and repetition rate reaching 10 0%. The total test time is about 3.5h. Conclusion DNA chip for HLA DR1, DR5 1 genotyping has the advantages of high resolution, high specificity and simple and rapid, suitable for the clinical application of organ transplantation.