G01025: Novel methods for detection of GM materials in foods and food ingredients

Study Duration: February 2003 to February 2005 (extended to April 2005)

Contractor: LGC

Trace nucleic acid detection technologies, such as real-time polymerase chain reaction (PCR) and the invader assay, have the potential to detect single copy nucleic acid molecules within complex food matrices.

This project had attempted to find an alternative to PCR as a method of amplification of genomic DNA by using rolling circle amplification (RCA) and combine this with mass spectrometry (MS) to achieve quantification of genetically modified (GM) material in food.

The use of RCA for amplification of genomic DNA was not successful. Detection of a specific GM DNA fragment was achieved by MS, but only after amplification by PCR.

Real-time PCR is currently the method of choice for detection of genetically modified (GM) DNA in food and feed. The need for sensitive detection methods is in part driven by current trends in regulation. The EU have set a 0.9% (w/w) limit (as a percentage of the total GM material in a food matrix) for the presence of GM ingredients in potentially processed foods and this is a good example of where measurement can struggle to match regulatory demands.

This project sought to examine and develop the application of two of the most promising emerging technologies in the context of the identification and quantitation of GM ingredients; RCA and MS. The overall investigation, aimed to determine whether these technologies offered the potential specificity and sensitivity needed to offer a replacement to current methodology.

The primary advantage of RCA is the ability to make copies of the target DNA under isothermal conditions i.e. at a uniform temperature. PCR, by contrast, requires the use of a dedicated instrument that performs the range of incubation temperatures required for this process. However, in this project, despite the claims in the literature, it has been demonstrated that RCA can be performed on synthetic pieces of DNA or PCR reaction products bearing the appropriate target DNA sequences (from soya and maize), but not on ‘real’ genomic DNA. It would therefore appear that, at present, RCA does not represent an alternative technique to that of PCR for the detection of GM material in food.

The second part of the project sought to use MS for the detection of GM ingredients. This is a technique that enables very accurate separation, identification and quantitation of specific ions in a sample based upon their mass charge ratio. Two of the most popular techniques used are electrospray ionisation (ESI) and matrix-assisted laser desorption ionisation (MALDI) MS. Both ESI and MALDI MS were assessed for the detection of PCR amplified DNA and primer extension products. Using MS, the specific detection and differentiation of GM and non-GM soya after DNA cleanup was possible, demonstrating the potential of this technique for quantitative measurement of DNA. However, further work will be needed to optimise this approach compared to existing techniques for the quantitation of GM ingredients in food products.

Pang S, Qureshi F, Shanahan D, Harris N (2007) Investigation of the use of rolling circle amplification for the detection of GM food. European Food Research and Technology. 225 (1) 59 - 66

Shanahan D, Stokes P, Burns M, Harris N (2007) A novel method for the detection of Round Up Ready TM Soya using liquid chromatography-electrospray ionisation mass spectrometry. European Food research and Technology. 225 (3&4) 483 - 491