单分子实时测序在HLA基因分型中的应用初探*

doi:10.3969/j.issn.1671-2587.2024.03.010

临床输血与检验 ›› 2024, Vol. 26 ›› Issue (3): 359-365.DOI: 10.3969/j.issn.1671-2587.2024.03.010

单分子实时测序在HLA基因分型中的应用初探^*

储玉霜¹, 肖彦琳¹, 望喆¹, 冯青青¹, 李亭亭², 卞茂红¹

¹安徽医科大学第一附属医院输血科,合肥 230022;
²江苏省泰州市泰兴人民医院检验科,泰州 225499

收稿日期:2024-03-25 出版日期:2024-06-20 发布日期:2024-06-24
通讯作者: 卞茂红,主要从事输血的基础和临床应用研究,（E-mail）mhbian@126.com。
作者简介:储玉霜,主要从事临床检验诊断学方面研究,（E-mail）2466269653@qq.com。
基金资助:
*皖江新兴产业技术发展中心产业化项目（No.WJ21CYHXM11）资助

Application of Single Molecule Real-time Sequencing in HLA Genotyping

CHU Yushuang¹, XIAO Yanlin¹, WANG Zhe¹, FENG Qinging¹, LI Tingting², BIAN Maohong¹

¹Department of Blood Transfusion, the First Affiliated Hospital of Anhui Medical University,Anhui Hefei 230022;
²Department of Laboratory Medicine, Taixing People's Hospital, Jiangsu Taizhou, 225499

Received:2024-03-25 Online:2024-06-20 Published:2024-06-24

摘要/Abstract

摘要： 目的分析单分子实时（SMRT）测序技术判断HLA基因型、单体型的准确性,以探讨该方法应用于临床的可行性。方法留取19例需行肾移植的患者血液标本,提取DNA后利用SMRT测序技术检测样本基因序列,并利用PCR-SBT方法验证其准确性。结果 SMRT技术可鉴定出19例样本的HLA基因型和基因单体型,且结果与PCR-SBT方法相符。1和14号样本PCR-SBT方法的C位点结果存在模棱两可现象,结果为HLA-C*03:02/04;03:03/132,SMRT技术可直接区分,结果为HLA-C*03:02:02,03:03:01。另外,SMRT方法在1号样本的B位点发现新的HLA等位基因。结论 SMRT测序技术应用于HLA高分辨率分型具有高度准确性,在HLA单体型分析具有精准定位多个长间距突变位置的独特优势。

关键词: 单分子实时测序, 长读长测序, HLA分型, 单体型分析

Abstract: Objective To explore the accuracy and the feasibility of this method in clinical application, single molecule real-time (SMRT) sequencing technology was employed to determine HLA genotyping, haploid type. Methods Blood samples were collected from 19 patients who needed kidney transplantation, genomic DNA was extracted respectively, and the sequences of all samples were obtained by SMRT sequencing technology, and then detected by PCR-SBT to verify the accuracy of the result. Results The genotypes and haplotypes of 19 samples could be assigned by SMRT technique, which were consistent with those of the PCR-SBT method. The results of HLA-C locus showed ambiguous combination on samples 1 and 14 using SBT method, with HLA-C*03:02,03:03 and -C*03:04,03:132; while the unique result of them is HLA-C*03:02:02, 03:03:01 using SMRT technology. In addition, a new HLA-B allele of sample 1 was identified using SMRT method. Conclusion SMRT sequencing technology has high accuracy when applied to HLA high-resolution typing, and has unique advantages of accurately locating multiple long-distance mutation in HLA haploid typing.

Key words: Single molecule real-time sequencing, Long read sequencing, HLA typing, Haploid typing

中图分类号:

储玉霜, 肖彦琳, 望喆, 冯青青, 李亭亭, 卞茂红. 单分子实时测序在HLA基因分型中的应用初探^*[J]. 临床输血与检验, 2024, 26(3): 359-365.

CHU Yushuang, XIAO Yanlin, WANG Zhe, FENG Qinging, LI Tingting, BIAN Maohong. Application of Single Molecule Real-time Sequencing in HLA Genotyping[J]. JOURNAL OF CLINICAL TRANSFUSION AND LABORATORY MEDICINE, 2024, 26(3): 359-365.

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单分子实时测序在HLA基因分型中的应用初探^*

Application of Single Molecule Real-time Sequencing in HLA Genotyping

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