Using Next-Generation Sequencing Technology to Confirm the HLA Rare Alleles Detected by PCR-SSOP.
10.19746/j.cnki.issn.1009-2137.2023.01.032
- Author:
Xian-Xin ZHONG
1
;
Wang-Da WU
1
;
Zhan-Rou QUAN
2
;
Su-Qing GAO
3
Author Information
1. Department of Clinical Laboratory Examination, Shenzhen Traditional Chinese Medicine Hospital, Shenzhen 518033, Guangdong Province, China.
2. Institute of Transfusion Medicine of Shenzhen Blood Center, Shenzhen 518020, Guangdong Province, China.
3. Institute of Transfusion Medicine of Shenzhen Blood Center, Shenzhen 518020, Guangdong Province, China.E-mail: gaosuqing1@163.com.
- Publication Type:Journal Article
- Keywords:
HLA;
PCR sequence-based typing;
PCR sequence-specific oligonudeotide probe;
allele;
next generation sequencing
- MeSH:
Humans;
Alleles;
Polymerase Chain Reaction;
Genotype;
High-Throughput Nucleotide Sequencing;
Histocompatibility Testing/methods*;
Technology
- From:
Journal of Experimental Hematology
2023;31(1):203-208
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To confirm the HLA genotypes of the samples including 4 cases of magnetic bead probe HLA genotyping result pattern abnormality and 3 cases of ambiguous result detected by PCR sequence-specific oligonudeotide probe (SSOP) method.
METHODS:All samples derived from HLA high-resolution typing laboratory were detected by PCR-SSOP. A total of 4 samples of magnetic bead probe HLA genotyping result pattern abnormality and 3 samples of ambiguous result were further confirmed by PCR sequence-based typing (SBT) technology and next-generation sequencing (NGS) technology.
RESULTS:A total of 4 samples of magnetic bead probe HLA genotyping result pattern abnormality were detected by PCR-SSOP method. The results of SBT and NGS showed that the HLA-A genotype of sample 1 did not match any known genotypes. NGS analysis revealed that the novel allele was different from the closest matching allele A*31:01:02:01at position 154 with G>A in exon 2, which resulting in one amino acid substitution at codon 28 from Valine to Methionine (p.Val28Met). The HLA-C genotype of sample 2 was C*03:119, 06:02, sample 3 was C*03:03, 07:137, and sample 4 was B*55:02, 55:12. A total of 3 samples with ambiguous result were initially detected by PCR-SSOP method. The re-examination results of SBT and NGS showed that the HLA-B genotype of sample 5 was B*15:58, 38:02, sample 6 was DRB1*04:05, 14:101, and sample 7 was DQB1*03:34, 05:02. Among them, alleles C*03:119, C*07:137 and DRB1*14:101 were not included in the Common and Well-documented Alleles (CWD) v2.4 of the Chinese Hematopoietic Stem Cell Donor Database.
CONCLUSION:The abnormal pattern of HLA genotyping results of magnetic probe by PCR-SSOP method suggests that it may be a rare allele or a novel allele, which needs to be verified by sequencing.
