Barbara Jerič (Author), Maja Štalekar (Author), Sebastian Vencken (Author), David Dobnik (Author), Polona Kogovšek (Author), Matjaž Stanonik (Author), Miha Arnol (Author), Maja Ravnikar (Author)

Abstract

Increasing research demonstrates the potential of donor-derived cell-free DNA (dd-cfDNA) as a biomarker for monitoring the health of various solid organ transplants. Several methods have been proposed for cfDNA analysis, including real-time PCR, digital PCR, and next generation sequencing-based approaches. We sought to revise the droplet digital PCR (ddPCR)-based approach to quantify relative dd-cfDNA in plasma from kidney transplant (KTx) patients using a novel pilot set of assays targeting single nucleotide polymorphisms that have a very high potential to distinguish cfDNA from two individuals. The assays are capable of accurate quantification of down to 0.1% minor allele content when analyzing 165 ng of human DNA. We found no significant differences in the yield of extracted cfDNA using the three different commercial kits tested. More cfDNA was extracted from the plasma of KTx patients than from healthy volunteers, especially early after transplantation. The median level of donor-derived minor alleles in KTx samples was 0.35%. We found that ddPCR using the evaluated assays within specific range is suitable for analysis of KTx patientsʼ plasma but recommend prior genotyping of donor DNA and performing reliable preamplification of cfDNA.

Keywords

kidney transplantation;droplet digital PCR;plasma cell-free DNA;minor allele quantification;assay evaluation;graft health monitoring;

Data

Language: English
Year of publishing:
Typology: 1.01 - Original Scientific Article
Organization: NIB - National Institute of Biology
UDC: 616.6:615
COBISS: 79997955 Link will open in a new window
ISSN: 2296-858X
Views: 15
Downloads: 8
Average score: 0 (0 votes)
Metadata: JSON JSON-RDF JSON-LD TURTLE N-TRIPLES XML RDFA MICRODATA DC-XML DC-RDF RDF

Other data

Source comment: Soavtorji: Maja Štalekar, Sebastian Vencken, David Dobnik, Polona Kogovšek, Matjaž Stanonik, Miha Arnol and Maja Ravnikar; Nasl. z nasl. zaslona; Opis vira z dne 11. 10. 2021; Št. članka: 748668;
Pages: str. 1-9
Issue: ǂVol. ǂ8
Chronology: 8 October 2021
DOI: 10.3389/fmed.2021.748668
ID: 24555977