The authors of this study succeeded in designing a specific PCR method to detect a Cibus rapeseed variety that carries a herbicide tolerance in its genome. An important prerequisite for the application of the method must be fulfilled: the changes in the respective gene location of the examined varieties must be known beforehand.
First, the area that contains the point mutation in the genome is amplified and then sequenced, i.e. the DNA sequence is decoded. This makes it possible to detect small variants that can occur naturally in different varieties, but also variants that originated from the use of genome editing or mutagenesis. The authors examined two rapeseed varieties, one containing a single change in the AHAS1C gene (Cibus rapeseed), and another variety with a single change in the AHAS3A gene produced by chemical mutagenesis (BASF rapeseed). The varieties both convey herbicide tolerance.
Based on the DNA sequences of the respective varieties, the authors of the study are developing a qPCR (quantitative PCR) method which makes it possible to specifically detect the respective change in the AHAS1C gene of the Cibus rapeseed or in the AHAS3A gene of the BASF rapeseed and to determine the respective allele quantitatively. The procedure was tested and verified by another, independent laboratory.
The study provides a method to detect known small changes that have, for example, been made in the genome by genome editing (or other processes) and that can be used in the approval processes of such varieties. However, if two varieties show the same alteration in the same position of the genome, which have arisen through different processes (e.g. through CRISPR/Cas, chemical mutagenesis or naturally occurring mutations), the method described here does not allow to determine the process that was used. The authors discuss that identification of the method is not necessary to detect unauthorized GMOs.