Two new RHD alleles with deletions spanning multiple exons.
Journal
Transfusion
ISSN: 1537-2995
Titre abrégé: Transfusion
Pays: United States
ID NLM: 0417360
Informations de publication
Date de publication:
03 2021
03 2021
Historique:
revised:
29
09
2020
received:
19
08
2020
accepted:
11
10
2020
pubmed:
27
11
2020
medline:
15
7
2021
entrez:
26
11
2020
Statut:
ppublish
Résumé
The most common large-deletion RHD allele (RHD*01N.01) includes the entire coding sequence, intervening regions and untranslated regions. The rest of large-deletion RHD alleles reported to-date consist of single-exon deletions, such as RHD*01N.67 which includes exon 1. Samples from two donors with RhD-negative serology yielded unclear or inconclusive results when subject to confirmatory testing on RHD genotyping arrays. To determine their RHD genotypes, genomic DNA was analyzed with a combination of allele-specific PCR, long-range PCR, Sanger sequencing, and next-generation sequencing assays. Allele-specific PCR failed to detect products for RHD exons 1 to 3 in one sample and RHD exons 1 to 5 in the other. A quantitative next-generation sequencing assay confirmed deletion of exons 1 to 3 and 1 to 5 respectively, and detected the absence of an RHD gene in trans in both samples. Long-range PCR and Sanger sequencing enabled identification of the breakpoints for both alleles. Both deletions start within the 5' Rhesus box (upstream of the identity region for the 1-to-3 deletion, downstream of it for the 1-to-5 deletion), and end within introns. Resolution of unclear or inconclusive results from targeted genotyping arrays often leads to the discovery of new alleles. The 5' Rhesus box may be a hot spot for genetic recombination events, such as the large deletions described in this report.
Sections du résumé
BACKGROUND
The most common large-deletion RHD allele (RHD*01N.01) includes the entire coding sequence, intervening regions and untranslated regions. The rest of large-deletion RHD alleles reported to-date consist of single-exon deletions, such as RHD*01N.67 which includes exon 1.
MATERIALS AND METHODS
Samples from two donors with RhD-negative serology yielded unclear or inconclusive results when subject to confirmatory testing on RHD genotyping arrays. To determine their RHD genotypes, genomic DNA was analyzed with a combination of allele-specific PCR, long-range PCR, Sanger sequencing, and next-generation sequencing assays.
RESULTS
Allele-specific PCR failed to detect products for RHD exons 1 to 3 in one sample and RHD exons 1 to 5 in the other. A quantitative next-generation sequencing assay confirmed deletion of exons 1 to 3 and 1 to 5 respectively, and detected the absence of an RHD gene in trans in both samples. Long-range PCR and Sanger sequencing enabled identification of the breakpoints for both alleles. Both deletions start within the 5' Rhesus box (upstream of the identity region for the 1-to-3 deletion, downstream of it for the 1-to-5 deletion), and end within introns.
CONCLUSIONS
Resolution of unclear or inconclusive results from targeted genotyping arrays often leads to the discovery of new alleles. The 5' Rhesus box may be a hot spot for genetic recombination events, such as the large deletions described in this report.
Substances chimiques
Rh-Hr Blood-Group System
0
Rho(D) antigen
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
682-686Informations de copyright
© 2020 AABB.
Références
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