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Use of real-time PCR and fluorimetry for rapid detection of rifampin and isoniazid resistance-associated mutations in Mycobacterium tuberculosis

Opened Access Use of real-time PCR and fluorimetry for rapid detection of rifampin and isoniazid resistance-associated mutations in Mycobacterium tuberculosis
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Autor: Torres Sánchez, María José
Criado, Antonio
Palomares Folía, José Carlos
Aznar Martín, Javier
Departamento: Universidad de Sevilla. Departamento de Microbiología
Fecha: 2000
Publicado en: Journal of Clinical Microbiology, 38 (9), 3194-3199.
Tipo de documento: Artículo
Resumen: Very fast amplification of DNA in small volumes can be continuously monitored with a rapid cycler that incorporates fluorimetric detection. Primers were designed to amplify a 157-bp fragment of the rpoB gene spanning codons 526 and 531 and a 209-bp fragment of the katG gene spanning codon 315 of Mycobacterium tuberculosis. Most mutations associated with resistance to rifampin (RMP) and isoniazid (INH) in clinical isolates occur in these codons. Two pairs of hybridization probes were synthesized; one in each pair was 3' labeled with fluorescein and hybridized upstream of the codon with the mutation; the other two probes were 5' labeled with LightCycler-Red 640. Each pair of probes recognized adjacent sequences in the amplicon. After DNA amplification was finished by using a LightCycler, the temperature at which the Red 640 probe melted from the product was determined in a 3-min melt program. Twenty M. tuberculosis clinical isolates susceptible to streptomycin, INH, RMP, and ethambutol ...
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Cita: Torres Sánchez, M.J., Criado, A., Palomares Folia, J.C. y Aznar Martín, J. (2000). Use of real-time PCR and fluorimetry for rapid detection of rifampin and isoniazid resistance-associated mutations in Mycobacterium tuberculosis. Journal of Clinical Microbiology, 38 (9), 3194-3199.
Tamaño: 99.16Kb
Formato: PDF

URI: http://hdl.handle.net/11441/64866

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