Detection of Gluten-Rich Cereals in Processed Foods with Enhanced Sensitivity by Targeting Mitochondrial DNA Using PCR
- 55 Downloads
Since the correct labeling of foods is the only effective way of protecting celiac disease patients and people allergic to gluten, it is essential to have reliable methods to detect the presence of gluten-containing cereals in foods. DNA-based methods have their merits as complementary approaches to immunochemical assays to detect the possible gluten contamination in processed foodstuff. Nevertheless, insufficient sensitivity is a major drawback for DNA-based analytical methods. Accordingly, DNA markers within mitochondrial DNA were identified in this study as the targets of PCR for the detection of gluten-containing cereals. Primer pairs Mt1F/Mt1R and Mt2F/Mt2R target a unique DNA segment conserved among wheat, barley, rye, and oat, whereas Mt3F/Mt3R targets the other segment that is present only in wheat, barley, and rye. In regard to detection limits, all the three designed primers could detect the presence of 0.2 pg wheat, barley, and rye DNA, 10−5 (10 ppm) of wheat, barley, and rye DNA diluted with soya DNA, or 10−5 (10 ppm) of wheat flour mixed within corn flour. The extremely high copy numbers of mitochondrial genome per cell may explain and justify the astonishing sensitivity of the methods described in this study. The detection of wheat DNA in thermally processed foods by this method is evidence of the suitability and applicability of the method to examine foods with gluten-free labels.
KeywordsCeliac disease Gluten-free foods Wheat allergy Gliadin PCR Molecular diagnosis
We would like to thank Professor Weiming Leu at the Graduate Institute of Biotechnology, National Chung Hsing University, Taiwan, for her assistance with bioinformatics analysis. This study was financially supported by the Minister of Science and Technology, R.O.C. (Taiwan), under the grant MOST 106-2313-B-005-036-MY3.
Compliance with Ethical Standards
Conflict of Interest
Nahed Ahmed declares that he has no conflict of interest. Menghsiao Meng declares that he has no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
- Anonymous (2005) International Standard EN ISO 21571Google Scholar
- Catassi C, Fabiani E, Iacono G, D'Agate C, Francavilla R, Biagi F, Volta U, Accomando S, Picarelli A, de Vitis I, Pianelli G, Gesuita R, Carle F, Mandolesi A, Bearzi I, Fasano A (2007) A prospective, double-blind, placebo-controlled trial to establish a safe gluten threshold for patients with celiac disease. Am J Clin Nutr 85:160–166CrossRefGoogle Scholar
- Haraszi R, Chassaigne H, Maquet A, Ulberth F (2011) Analytical methods for detection of gluten in food-method developments in support of food labeling legislation. J AOAC Int 94:1006–1025Google Scholar
- Mazzara M, Savini C, Delobel C, Broll H, Damant A, Paoletti C, van den Eede G (2008) Definition of minimum performance requirements for analytical methods of GMO Testing European Network of GMO Laboratories (ENGL). Joint Research Centre and Office for Official Publications of the European Communities. 8 p. OPOCE, BrusselsGoogle Scholar
- Ogihara Y, Yamazaki Y, Murai K, Kanno A, Terachi T, Shiina T, Miyashita N, Nasuda S, Nakamura C, Mori N, Takumi S, Murata M, Futo S, Tsunewaki K (2005) Structural dynamics of cereal mitochondrial genomes as revealed by complete nucleotide sequencing of the wheat mitochondrial genome. Nucleic Acids Res 33:6235–6250CrossRefGoogle Scholar
- Piknová Ľ, Brežná B, Kuchta T (2008) Detection of gluten-containing cereals in food by 5′-nuclease real-time polymerase chain reaction. J Food Nutr Res 47:114–119Google Scholar