title: Targeting Conserved Genes in Alternaria Species.
abstract: Real-time polymerase chain reaction (PCR) is a molecular biology technique based on the detection of the fluorescence produced by a reporter molecule, which increases as the reaction proceeds proportionally to the accumulation of the PCR product within each amplification cycle. The fluorescent reporter molecules include dyes that bind to the double-stranded DNA (i.e., SYBR®Green) or sequence-specific probes (i.e., Molecular Beacons or TaqMan®Probes). Real-time PCR provides a tool for accurate and sensitive quantification of target fungal DNA. Here, we describe a TaqMan real-time PCR method for specific detection and quantification of Alternaria spp. The method uses Alternaria-specific primers and probe, targeting the internal transcribed spacer regions ITS1 and ITS2 of the rRNA gene, and a positive amplification control based on 18S rRNA gene.
authors.affiliation: Facultad de Veterinaria, Departamento de Nutrición, Bromatología y Tecnología de los Alimentos, Universidad Complutense de Madrid, Avda. Puerta de Hierro s/n, Madrid, 28040, Spain.
authors.forename: Miguel Ángel
authors.initials: MÁ
authors.lastname: Pavón
authors.affiliation: Facultad de Veterinaria, Departamento de Nutrición, Bromatología y Tecnología de los Alimentos, Universidad Complutense de Madrid, Avda. Puerta de Hierro s/n, Madrid, 28040, Spain.
authors.forename: Inés María
authors.initials: IM
authors.lastname: López-Calleja
authors.affiliation: Facultad de Veterinaria, Departamento de Nutrición, Bromatología y Tecnología de los Alimentos, Universidad Complutense de Madrid, Avda. Puerta de Hierro s/n, Madrid, 28040, Spain.
authors.forename: Isabel
authors.initials: I
authors.lastname: González
authors.affiliation: Facultad de Veterinaria, Departamento de Nutrición, Bromatología y Tecnología de los Alimentos, Universidad Complutense de Madrid, Avda. Puerta de Hierro s/n, Madrid, 28040, Spain.
authors.forename: Rosario
authors.initials: R
authors.lastname: Martín
authors.affiliation: Facultad de Veterinaria, Departamento de Nutrición, Bromatología y Tecnología de los Alimentos, Universidad Complutense de Madrid, Avda. Puerta de Hierro s/n, Madrid, 28040, Spain. tgarcia@vet.ucm.es.
authors.forename: Teresa
authors.initials: T
authors.lastname: García
mesh_terms: D000528:Alternaria / Q000145:classification* / Q000235:genetics*; D017124:Conserved Sequence; D021901:DNA, Intergenic; D019143:Evolution, Molecular; D005800:Genes, Fungal; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D021901:DNA, Intergenic
keywords: Alternaria spp.; Internal transcribed spacer; TaqMan real-time PCR