title: Nanoparticle-based DNA biosensor for visual detection of genetically modified organisms.
abstract: Although screening of raw ingredients and food products for genetically modified organisms (GMO) may be accomplished by detecting either the exogenous DNA or the novel protein, DNA is the preferred analyte because of its superior stability during food processing. The development of DNA biosensors is of increasing importance due to the growing demand for rapid and reliable methods for GMO detection. We report the first DNA biosensor in a dry-reagent dipstick configuration for visual detection and confirmation of GMO-related sequences by hybridization within minutes. The sensor is disposable and does not require special instrumentation. It detects the 35S promoter and nopaline synthase (NOS) terminator sequences that are present in the majority of transgenic plants. The target sequences are amplified by the polymerase chain reaction (PCR) and hybridized (7min) with probes bearing oligo(dA) tail. The biotinylated product is applied to the sensor followed by immersion in the appropriate buffer. Migration of the buffer rehydrates gold nanoparticles conjugated to oligo(dT), which hybridize with the oligo(dA) tails. The hybrids are captured by immobilized streptavidin at the test zone of the sensor giving a characteristic red line due to the accumulation of the nanoparticles. The excess of nanoparticle conjugates are captured at the control zone by immobilized oligo(dA) strands. Amplified 35S or NOS DNA is detectable at 0.16nM. Soybean powder certified reference material with 0.1% GMO content is clearly detectable after 35 and 40 amplification cycles for 35S and NOS sequence, respectively. The sensor was also applied to real samples from various sources.
authors.affiliation: Department of Chemistry, University of Patras, Patras 26500, Greece.
authors.forename: Despina P
authors.initials: DP
authors.lastname: Kalogianni
authors.forename: Theodora
authors.initials: T
authors.lastname: Koraki
authors.forename: Theodore K
authors.initials: TK
authors.lastname: Christopoulos
authors.forename: Penelope C
authors.initials: PC
authors.lastname: Ioannou
mesh_terms: D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D018744:DNA, Plant / Q000032:analysis* / Q000235:genetics*; D004209:Disposable Equipment; D004867:Equipment Design; D019544:Equipment Failure Analysis; D017403:In Situ Hybridization / Q000295:instrumentation* / Q000379:methods; D049329:Nanostructures / Q000737:chemistry; D036103:Nanotechnology / Q000295:instrumentation / Q000379:methods; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation* / Q000379:methods; D030821:Plants, Genetically Modified / Q000235:genetics*; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D018744:DNA, Plant; D011933:Reagent Kits, Diagnostic