基于Ion Torrent S5平台的二代测序技术对HLA-DRB1基因分型模棱两可进行分析

目的 基于Ion Torrent S5平台NGS法进行HLA-DRB1基因分型检测的模棱两可结果的种类、比例进行分析.方法 对471例脐带血造血干细胞库标本采用基于Ion Torrent S5平台的二代测序法检测HL4-DRB1位点,并同步对其中的94例标本采用PCR-SBT测定HL4-DRBI位点,余下377例标本采...

Analyze the ambiguous results of HLA-DRB1 genotyping using NGS method based on the Ion Torrent S5 platform

Objective To analyze the kinds and ratio of ambiguous results of the HLA-DRB1 genotyping using next generation sequencing(NGS) method based on Ion Torrent S5 platform. Methods A total of 471 samples from Blood Center of Zhejiang Province were genotyped for HLA-DRB1 loci by the NGS method,and meanwhile 94 samples among them were detected for HLA-DRB1 loci by PCR-SBT method,and the other 377 samples were detected for HLA-DRB1 loci by PCR-SSO flow fluorescent bead method. The genotyping results of HLA-DRB1 loci by the NGS method were assigned using auto-analytical software. The number of the third field of HLA-DRB1 * was used as high resolution level. The ambiguous combination ratio of the results was calculated by the directly count method. Results The results of NGS detection of HLA-DRB1 gene in 470 samples were consistent with PCR-SBT or PCR-SSO. One allele was missed by NGS method,and the coincidence rate was 99. 8%.Among the 471 samples detected by NGS method,there were 160 samples with ambiguous HLA-DRB1 allelic genotypes,with a proportion of 33. 97%(160/471). The most common combination of ambiguous results is DRB1 * 09 ∶ 01 ∶ 02/09 ∶ 21.Conclusion The NGS technology based on Ion Torrent S5 platform can significantly reduce the ambiguity of HLA-DRB1 genotyping result,while there were still a few HLA-DRB1 genotyping ambiguous results. The risk for allele-dropout should be pay attention in the NGS method.