NAIMA: target amplification strategy allowing quantitative on-chip detection of GMOs

We have developed a novel multiplex quantitative DNA-based target amplification method suitable for sensitive, specific and quantitative detection on microarray. This new method named NASBA Implemented Microarray Analysis (NAIMA) was applied to GMO detection in food and feed, but its application can be extended to all fields of biology requiring simultaneous detection of low copy number DNA targets. In a first step, the use of tailed primers allows the multiplex synthesis of template DNAs in a primer extension reaction. A second step of the procedure consists of transcription-based amplification using universal primers. The cRNA product is further on directly ligated to fluorescent dyes labelled 3DNA dendrimers allowing signal amplification and hybridized without further purification on an oligonucleotide probe-based microarray for multiplex detection. Two triplex systems have been applied to test maize samples containing several transgenic lines, and NAIMA has shown to be sensitive down to two target copies and to provide quantitative data on the transgenic contents in a range of 0.1–25%. Performances of NAIMA are comparable to singleplex quantitative real-time PCR. In addition, NAIMA amplification is faster since 20 min are sufficient to achieve full amplification

A PCR-microarray method for the screening of genetically modified organisms

A new method to screen and to identify genetically modified organisms (GMO) is presented in this paper. It is based on the detection of multiple genetic elements common to GMO by their amplification via PCR followed by direct hybridisation of the amplicons on microarray. The pattern of the elements is then compared to a database of the composition of EU-approved GMO and an identification of the GMO is then proposed. The limit of detection of the method was ≤0.1% GMO content (w/w) expressed as the amount of target DNA present in the template for single unprocessed material. The DNA targets were detected both in reference materials and in mixtures with the same detection limit. The specificity for the detection of the different elements was found to be very good with no cross-reaction even in samples with two GMO present at different concentrations. The paper presents examples of GMO identification and discusses the potential and limitation of such approaches and how they can facilitate the work of private and enforcement detection laboratories.This study was supported by the European Commission through the FP5 program “GMOchips” (contract G6RD-CT2000-00419 2000-2003), the Belgian SSCT program and the Integrated Project “Co-Extra”, contract no. 007158 2005-2009, under the 6th Framework Programme, priority 5, food quality and safety.Peer reviewe