title: Detection of food-borne pathogens with DNA arrays on disk.
abstract: A DNA oligonucleotide array for duplex pathogen detection on a DVD platform is developed. The assay involves hybridization of PCR products and optical detection using compact disc technology. Different DNA array constructions for attachment of synthetic oligonucleotides on to DVD surface are evaluated, finding that streptavidin-biotin coupling method yielded the highest sensitivity in combination with enzymatic signal amplification. Issues of importance for the DNA array construction such immobilized probes design, PCR product labeling strategy and composition of the hybridization buffer were addressed. The methodology was proved scoring single nucleotide polymorphisms with high selectivity. The assay capability was also demonstrated by the identification of two pathogenic microorganisms in powder milk samples. In fifty minutes, the DVD-array system identifies Salmonella spp. and Cronobacter spp. (previously named Enterobacter sakazakii) precise and simultaneously with a sensitivity of 10(0) and 10(2) cfu/mL, respectively, in infant milk. Results were in good agreement with those obtained by quantitative real-time PCR.
authors.affiliation: Centro de Reconocimiento Molecular y Desarrollo Tecnológico, Departamento de Química, Universitat Politécnica de Valéncia, Camino de Vera s/n, 46071 Valencia, Spain.
authors.forename: T
authors.initials: T
authors.lastname: Arnandis-Chover
authors.forename: S
authors.initials: S
authors.lastname: Morais
authors.forename: L A
authors.initials: LA
authors.lastname: Tortajada-Genaro
authors.forename: R
authors.initials: R
authors.lastname: Puchades
authors.forename: Á
authors.initials: Á
authors.lastname: Maquieira
authors.forename: J
authors.initials: J
authors.lastname: Berganza
authors.forename: G
authors.initials: G
authors.lastname: Olabarria
mesh_terms: D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D005516:Food Microbiology; D008892:Milk / Q000382:microbiology; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide
chemical_list: D017931:DNA Primers