EPO transgene detection in dried blood spots for antidoping application.
DBS
DNA analysis
EPO
gene doping
real-time PCR
Journal
Drug testing and analysis
ISSN: 1942-7611
Titre abrégé: Drug Test Anal
Pays: England
ID NLM: 101483449
Informations de publication
Date de publication:
Nov 2021
Nov 2021
Historique:
revised:
20
04
2021
received:
09
02
2021
accepted:
03
05
2021
pubmed:
8
5
2021
medline:
17
3
2022
entrez:
7
5
2021
Statut:
ppublish
Résumé
The modification of gene expression to treat diseases is a field of research with exponential growth. As doping in sport closely follows emerging therapies, a surveillance of the modification of gene expression to enhance performance is needed. The gene coding for erythropoietin (EPO) is one target of interest. Since 2010, several protocols have been proposed to identify EPO gene doping by focusing on the presence in blood of a transgene that differ in size from the endogenous gene sequence, normally found in the human DNA. In this work, our aim was to validate an easily applicable method for EPO gene doping detection in dried blood spots (DBS). We evaluated the detection of EPO transgene in 20-μl DBS after the spike of a plasmid carrying the EPO transgene in whole blood. Three different DBS were compared: Nucleic-Card™, Whatman® 903, and the volumetric 20-μl VAMS™. Detection was performed with real-time polymerase chain reaction (PCR) and validated with two Taqman assays (one commercial and one custom) specific for the EPO transgene. The initial testing procedure could be done using one assay (custom) and the confirmation using the second one (commercial Taqman) with a final check of the size of the PCR product. Starting from 20-μl dried blood, 1000 copies of EPO transgene could efficiently be detected with the three types of DBS, VAMS showing a slightly better sensitivity. No loss of sensitivity was observed after 1-month storage of DBS at room temperature. This method could be applied to DBS collected during doping controls and allows reanalysis.
Substances chimiques
EPO protein, human
0
Erythropoietin
11096-26-7
Types de publication
Comparative Study
Journal Article
Validation Study
Langues
eng
Sous-ensembles de citation
IM
Pagination
1888-1896Subventions
Organisme : Agence Française de Lutte contre le Dopage
Informations de copyright
© 2021 John Wiley & Sons, Ltd.
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