title: A multiplex, bead-based array for profiling plant-derived components in complex food matrixes.
abstract: Authentication of processed food ingredients is becoming an important issue for customers, and some DNA-based analytical methods have been developed, especially for animal products. As food products typically contain several different ingredients, a current challenge is to increase the multiplexing capacity of DNA-based methods, to develop "all-in-one" assays. Oligonucleotide-coupled, bead-based suspension arrays are sensitive and reproducible multiplex analytical tools. We applied the Multi-Analyte Profile (xMAP™) technology to develop an assay able to concurrently detect five different plant components in mixed flours and in processed feed and food. Capture probes were targeted to species-specific DNA polymorphisms present within the first intron of plant β-tubulin genes, which can be amplified by the tubulin-based polymorphism-amplification method (TBP-PCR). The workflow is very simple and straightforward, consisting of a PCR amplification step with universal primers, followed by the direct hybridization assay. Results are highly reproducible. For each single plant species, the absolute detection limit was as low as one target DNA copy. In complex mixtures of flours derived from seeds or from commercial dry "pasta," relative limits of detection ranged, in weight, from 2% for soybean to less than 0.5% for wheat. The specificity of the capture probes and the high sensitivity of the method allowed the successful determination of the analytical composition of three feeds as well as eleven food samples, such as snacks, biscuits, and pasta. The multiplexing ability of the assay (up to 100 different analytes) provides scalability and flexibility, in response to specific needs.
authors.affiliation: Istituto Biologia e Biotecnologia Agraria-Consiglio Nazionale delle Ricerche, Via Bassini 15, 20133, Milano, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Ponzoni
authors.forename: Diego
authors.initials: D
authors.lastname: Breviario
authors.forename: Alessandro
authors.initials: A
authors.lastname: Mautino
authors.forename: Silvia
authors.initials: S
authors.lastname: Gianì
authors.forename: Laura
authors.initials: L
authors.lastname: Morello
mesh_terms: D000821:Animal Feed / Q000032:analysis*; D000818:Animals; D018744:DNA, Plant / Q000032:analysis*; D005433:Flour / Q000032:analysis*; D005524:Food Technology; D017343:Genes, Plant; D005819:Genetic Markers; D020411:Oligonucleotide Array Sequence Analysis; D010944:Plants / Q000145:classification / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D013045:Species Specificity; D014404:Tubulin / Q000235:genetics
chemical_list: D018744:DNA, Plant; D005819:Genetic Markers; D014404:Tubulin