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Pitfalls When Determining HNA-1 Genotypes and Finding Novel Alleles.
- Source :
-
International Journal of Molecular Sciences . Aug2024, Vol. 25 Issue 16, p9127. 18p. - Publication Year :
- 2024
-
Abstract
- Genetic variation in the FCGR3B gene is responsible for different variants of human neutrophil antigen 1 (HNA-1). Laboratory techniques currently utilized for routine HNA-1 genotyping, predominantly PCR-sequence-specific primer (PCR-SSP) and PCR-sequence-based typing (PCR-SBT), lack specificity for FCGR3B. This study compares the capabilities and limitations of existing technologies including an in-house TaqMan PCR, a commercial PCR-SSP test, PCR-SBT and multiplex ligation-dependent probe amplification (MLPA) with those of a long-read nanopore sequencing assay. Testing was performed with both related and unrelated Danish samples with different copy numbers and/or rare alleles. Long-read nanopore sequencing was validated by blind testing of ten English samples. The results showed that FCGR3B copy numbers correlate with a dose-dependent distribution of alleles that complicates genotyping by TaqMan PCR, PCR-SSP and PCR-SBT, due to co-amplification of the homologous FCGR3A gene. MLPA can correctly quantify the dose-dependent distribution but not detect novel variants. Long-read nanopore sequencing showed high specificity for FCGR3B and was able to detect dosage-dependent distribution, and rare and novel variants that were previously not described. Current HNA-1 genotyping methods cannot produce unambiguous allele-level results, whereas long-read nanopore sequencing has shown the potential to resolve observed ambiguities, identify new HNA-1 variants and allow definitive allele assignment. [ABSTRACT FROM AUTHOR]
- Subjects :
- *GENETIC variation
*ALLELES
*GENOTYPES
*LABORATORY techniques
*NEUTROPHILS
Subjects
Details
- Language :
- English
- ISSN :
- 16616596
- Volume :
- 25
- Issue :
- 16
- Database :
- Academic Search Index
- Journal :
- International Journal of Molecular Sciences
- Publication Type :
- Academic Journal
- Accession number :
- 179349306
- Full Text :
- https://doi.org/10.3390/ijms25169127