A next-generation sequencing method for gene doping detection that distinguishes low levels of plasmid DNA against a background of genomic DNA.
Gene Ther
; 26(7-8): 338-346, 2019 08.
Article
em En
| MEDLINE
| ID: mdl-31296934
ABSTRACT
Gene doping confers health risks for athletes and is a threat to fair competition in sports. Therefore the anti-doping community has given attention on its detection. Previously published polymerase chain reaction-based methodologies for gene doping detection are targeting exon-exon junctions in the intron-less transgene. However, because these junctions are known, it would be relatively easy to evade detection by tampering with the copyDNA sequences. We have developed a targeted next-generation sequencing based assay for the detection of all exon-exon junctions of the potential doping genes, EPO, IGF1, IGF2, GH1, and GH2, which is resistant to tampering. Using this assay, all exon-exon junctions of copyDNA of doping genes could be detected with a sensitivity of 1296 copyDNA copies in 1000 ng of genomic DNA. In addition, promotor regions and plasmid-derived sequences are readily detectable in our sequence data. While we show the reliability of our method for a selection of genes, expanding the panel to detect other genes would be straightforward. As we were able to detect plasmid-derived sequences, we expect that genes with manipulated junctions, promotor regions, and plasmid or virus-derived sequences will also be readily detected.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Plasmídeos
/
Testes Genéticos
/
Análise de Sequência de DNA
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Transgenes
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Dopagem Esportivo
/
Sequenciamento de Nucleotídeos em Larga Escala
Tipo de estudo:
Diagnostic_studies
/
Prognostic_studies
Limite:
Humans
Idioma:
En
Ano de publicação:
2019
Tipo de documento:
Article