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Updated: May 20, 2025
Sequencing of mRNA from Whole Blood using Nanopore Sequencing
Published on: June 3, 2019
Determination of CD177 (human neutrophil antigen 2) polymorphisms using nanopore sequencing
Kirstine Kløve-Mogensen1, Thure Mors Haunstrup1,2,3, Anne-Louise Fjordside Bilde1
1Department of Clinical Immunology, Aalborg University Hospital, Aalborg, Denmark.
Background And Objectives:
Human neutrophil antigen 2 (HNA-2), encoded by the CD177 gene, is considered one of the most important neutrophil antigens in human medicine, but molecular testing of CD177 is complicated and therefore not a standard procedure for investigating CD177 expression. CD177 expression can vary from 0% to 100%, and to date, the molecular basis for altered or non-expressed genes has not been determined. Reliance on phenotyping and crossmatching to investigate these neutropenic clinical cases is inconvenient for patients and demands substantial resources within the laboratory. The purpose of this study was therefore to test a new molecular testing approach based on long-read nanopore sequencing.
Materials And Methods:
DNA from 44 Danish blood donors with different levels of CD177 expression, 22 of whom were found to be CD177 null, was selected as test samples. All the DNA was sequenced for the first eight exons and the beginning of exon 9 of CD177.
Results:
All incidences of CD177 null cases could be associated with the known variant c.787A>T;p.K263X (rs20182172), and a correlation was observed between c.787A>T heterozygosity and a reduced expression of CD177, which is consistent with previously published findings. The c.1291G>A;p.G431R (rs78718189) variant was found to be linked to the atypical expression of CD177. The nanopore assay revealed a total of 14 variants in 7 exons in the 44 tested samples.
Conclusion:
On the basis of these observations, we conclude that long-read nanopore sequencing can be a reliable tool for the routine laboratory molecular testing of CD177.
