To guarantee the traceability on the market and the freedom of choice for
consumers, genetically modified organisms (GMO) legislations have been
established in many countries, including in Europe (EU). However, the
implementation of these legislations by the enforcement laboratories is becoming
complex due mainly to the increasing number and diversity of GMO. To cope
with the problematic of EU unauthorized GMO, this PhD aims to improve and
strengthen the existing GMO detection system using high-tech approaches.
First, as a study case, an overview of genetically modified (GM) rice, developed
around the world was carried out to collect information related inter alia
on elements found in their transgenic cassette. Second, according to this
information, key targets, frequently found in GMO (p35S and tNOS) or
exclusively observed in EU unauthorized GMO (t35S pCAMBIA), were selected
to develop a strategy allowing to detect and characterize a broad range of
GMO. This strategy, fully integrated in the GMO routine analysis, consists
to characterize sequences surrounding detected key transgenic elements using
a DNA walking approach. By this way, the acquisition of sequences from
the junction between the transgenic cassette and the plant genome as well as
the associations of elements typically found in transgenic constructs allow to
confirm the presence of GMO in food/feed matrices. Due to its good performance
thoroughly assessed via several unprocessed and processed food/feed matrices,
this strategy represents a key tool, easily implementable by the enforcement
laboratories. With the aim to even more simplify the workflow and increase the
throughput of this strategy, the sequencing step was performed using the Next
Generation Sequencing (NGS) technology instead of the Sanger technology.
In parallel, the detection of GMO in alimentary matrices using exclusively the
NGS technology, through a whole genome sequencing (WGS) approach, was
also investigated. As this last approach does not theoretically require any prior
information about the targeted sequences, GMO composed only of unknown
transgenic elements could be detected.
This work has thus allowed to provide additional strategies to the current GMO
detection system in order to characterize a larger spectrum of GMO, both
authorized or not.