HLA-DRB1基因芯片与PCR-SSP分型的比较

目的：比较基因芯片和PCR-SSP用于汉族南方人群HLA-DRB1基因分型的结果,评价分型芯片的准确性。方法：77份待分型样本,分别用等位基因特异的PCR-SSP和基因分型芯片对DRB1分型。芯片分型通过组间特异引物扩增基因组DNA,扩增用荧光标记。扩增标记后的产物与分型芯片的探针杂交,通过杂交产生的荧光信号确定样品的HLA-DRB1基因亚型,比较两种方法分型所获得的结果,不吻合的样本全部经第三方验证或测序。结果：77份样本,两种方法分型全部成功。分型结果的吻合率为93％。不吻合样本6份,其中SSP定型纯合子4份,芯片分型全部为杂合子。经第三方验证,证实芯片对纯合子和杂合子的分型全部正确。另外2份不吻合的样本经测序,证实SSP分型错误1份,芯片分型错误1份。芯片的重复率为96％。结论：基因芯片是一种理想的分型方法,具有特异性高、重复性好、操作简便、所需样本量少、一次可作多份样本的优点。

Comparative Study of HLA-DRB1 Genotyping by Oligoneucleotide Arrays and PCR-SSP

Purpose To evaluate the accuracy and reliability of oligoneucleotide array in comparison with polymerase chain reaction with sequence-specific primers(PCR?SSP) in identification of human leukocyte antigen-DRB1(HLA?DRB1) alleles in Southern Han's population. Methods A total of 77 samples were enrolled in the study. HLA?DRB1 genotyping were performed by PCR?SSP and oligoneucleotide array. A pair of group-specific primers were designed according to the sequence of HLA?DRB1 exon2, then the primers and the Cy5-dCTP were used in the PCR, thus the PCR products were labeled with Cy5. The labeled PCR products were hybridized with the specific allele typing probes which were immobilized on a glass support, and the signals were scanned by scanner and then analyzed. The samples which the HLA?DRB1 typing results by arrays and PCR?SSP were discordant were verified by PCR?SSOP or sequencing. Results All the samples have been genotyped by HLA array and PCR?SSP successfully. 4 homozygous samples typed by PCR?SSP were actually heterozygous by array. The other 2 unidentified samples were typed by sequencing, the results showed that PCR?SSP made a mistake for 1 samples and array for 1. Conclusions The oligoneucleotide array technique is a precise, rapid molecular method for HLA?DRB1 genotyping, and has the advantage of high specificity and integration.