A sensitive and specific real-time PCR targeting DNA from wheat, barley and rye to track gluten contamination in marketed foods

García García, AinaMadrid García, RaquelSohrabi, HamidrezahCruz, Silvia de laGarcía Lacarra, TeresaMartín De Santos, María Del RosarioGonzález Alonso, María Isabel2024-01-202024-01-202019García-García, A., Madrid, R., Sohrabi, H., De La Cruz, S., Garcı́a, T., Martı́n, R., & González, I. (2019). A sensitive and specific real-time PCR targeting DNA from wheat, barley and rye to track gluten contamination in marketed foods. LWT, 114, 108378. https://doi.org/10.1016/j.lwt.2019.1083780023-643810.1016/j.lwt.2019.108378https://hdl.handle.net/20.500.14352/94204This work reports a real-time PCR assay to specifically detect the presence of DNA from the main gluten-containing cereals wheat, barley and rye (WBR) in foodstuffs. The WBR real-time PCR uses two specific primers and a Taqman probe for amplification of a 118 bp DNA fragment common to the three target cereals on the ribosomal internal transcribed spacer (ITS) region. In-house validation of the method was performed employing three binary arrays containing different concentrations (100 000–10 mg/kg) of a wheat/barley/rye mixture in maize flour: untreated, soft heat-treated at 160 °C/13 min and intense heat-treated at 200 °C/20 min. Results showed optimal PCR amplification efficiency, reproducibility and sensitivity with an overall limit of detection of 10–50 mg/kg of the target cereals, theoretically equating to approximately 1–5 mg/kg of gluten. Applicability of the assay was further assessed through a market analysis of 220 food products by the real-time PCR assay and the official R5-based ELISA. Comparative results highlighted the ability of the proposed methodology as a highly sensitive and robust procedure to detect the presence of potentially harmful concentrations of undeclared gluten-containing cereals in some of the tested samples, supporting results of the immunological assays currently used for gluten determination in foods.engAttribution 4.0 Internationalhttp://creativecommons.org/licenses/by/4.0/A sensitive and specific real-time PCR targeting DNA from wheat, barley and rye to track gluten contamination in marketed foodsjournal article1096-1127https://doi.org/10.1016/j.lwt.2019.108378restricted access636.09Taqman real-time PCRGluten-containing cerealsGluten-related disordersCommercial food productsVeterinaria31 Ciencias Agrarias