Optimised padlock probe ligation and microarray detection of multiple (non-authorised) GMOs in a single reaction

Abstract

Background: To maintain EU GMO regulations, producers of new GM crop varieties need to supply an event-specific method for the new variety. As a result methods are nowadays available for EU-authorised genetically modified organisms (GMOs), but only to a limited extent for EU-nonauthorised GMOs (NAGs). In the last decade the diversity of genetically modified (GM) ingredients in food and feed has increased significantly. As a result of this increase GMO laboratories currently need to apply many different methods to establish to potential presence of NAGs in raw materials and complex derived products.

Four new SYBR®Green qPCR screening methods for the detection of Roundup Ready®, LibertyLink®, and CryIAb traits in genetically modified products

“…Raymond et al [14] and Nadal et al [15] combined PCR with capillary electrophoresis in a very effective way, but requiring costly investment in additional equipment. Finally, an effective novel approach based on so-called padlock-probes [18] was developed, which could in the future represent an elegant solution to cover the broad diversity of GMO [16]. However, this technology is less sensitive and per se requires time-consuming optimization [19].…”

“…Several multiplex qPCR formats are available reducing the number of analyses and facilitating high throughput but requiring multi-channel detection devices and often including costly detection probes for at least some of the targets [8][9][10]. In other cases, the applied chemistries are quite complex or are to be combined with other technologies that are less appropriate for routine applications [11][12][13][14][15][16]. Several of these approaches involve the use of PCR with multiple targets and consecutive detection and identification of the amplification products using microarrays [11][12][13].…”

Development of a Molecular Platform for GMO Detection in Food and Feed on the Basis of “Combinatory qPCR” Technology

“…the cryIAb/Ac) are being used and detection methods have been developed (Bravo et al, 2007). It should however be noted that the above-mentioned methods are mostly either end-point detection on agarose gel or real-time qPCR using TaqMan® chemistry (Hamels et al, 2009;Raymond et al, 2010;Nadal et al, 2009;Prins et al, 2008). Development of screening methods using the SYBR ® Green qPCR technology only started recently Barbau-Piednoir et al, 2011;Mbongolo Mbella et al, 2011) although this approach offers a number of advantages over the TaqMan chemistry.…”

Safety of genetically modified (GM) crop ingredients in animal feed

“…Examples of these are multi-methods, such as microarray analyses, which are currently being developed at RIKILT -Institute of Food Safety (e.g. Prins et al, 2008).…”