Multiplex detection of antibiotic resistance genes using padlock probes.
Diagn Microbiol Infect Dis
; 77(2): 118-25, 2013 Oct.
Article
em En
| MEDLINE
| ID: mdl-23948548
ABSTRACT
The elucidation of resistance mechanisms is of central importance to providing and maintaining efficient medical treatment. However, molecular detection methods covering the complete set of resistance genes with a single test are still missing. Here, we present a novel 100-plex assay based on padlock probes in combination with a microarray that allows the simultaneous large-scale identification of highly diverse ß-lactamases. The specificity of the assay was performed using 70 clinical bacterial isolates, recovering 98% of the ß-lactamase nucleotide sequences present. Additionally, the sensitivity was evaluated with PCR products and genomic bacterial DNA, revealing a detection limit of 10(4) DNA copies per reaction when using PCR products as the template. Pre-amplification of genomic DNA in a 25-multiplex PCR further facilitated the detection of ß-lactamase genes in dilutions of 10(7) cells/mL. In summary, we present an efficient, highly specific, and highly sensitive multiplex detection method for any gene.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas de Bactérias
/
Beta-Lactamases
/
Sondas de DNA
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Análise de Sequência com Séries de Oligonucleotídeos
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Farmacorresistência Bacteriana
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Reação em Cadeia da Polimerase Multiplex
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
En
Revista:
Diagn Microbiol Infect Dis
Ano de publicação:
2013
Tipo de documento:
Article