Molecular Characterization and Event-Specific Real-Time PCR Detection of Two Dissimilar Groups of Genetically Modified Petunia (Petunia x hybrida) Sold on the Market.

Marleen M Voorhuijzen,Benjamin Pickel,Ingrid M J Scholtens,Anke Belter,Ottmar Goerlich,Theo W Prins,Andrea Stolz,Jeroen P Van Dijk,Esther J Kok,Joachim Bendiek,Lutz Grohmann,Claudia Brünen-Nieweler

doi:10.3389/fpls.2020.01047

Abstract

Petunia plants with unusual orange flowers were noticed on the European market and confirmed to be genetically modified (GM) by the Finnish authorities in spring 2017. Later in 2017, inspections and controls performed by several official laboratories of national competent authorities in the European Union detected several GM petunia varieties with orange flowers, but also another group of unusually colored flowers. In the latter group, a so far undetected gene coding for a flavonoid 3’5’ hydroxylase (F3’5’H) responsible for the purple color was identified by German and Dutch authorities, suggesting that the petunias found on the markets contain different genetic constructs. Here, a strategy is described for the identification of GM petunia varieties. It is based on an initial GMO screening for known elements using (real-time) PCR and subsequent identification of the insertion sites by a gene walking-like approach called ALF (amplification of linearly-enriched fragments) in combination with Sanger and MinION sequencing. The results indicate that the positively identified GM petunias can be traced back to two dissimilar GM events used for breeding of the different varieties. The test results also confirm that the transgenic petunia event RL01-17 used in the first German field trial in 1991 is not the origin of the GM petunias sold on the market. On basis of the obtained sequence data, event-specific real-time PCR confirmatory methods were developed and validated. These methods are applicable for the rapid detection and identification of GM petunias in routine analysis. In addition, a decision support system was developed for revealing the most likely origin of the GM petunia.

Highlights

In spring 2017 Finnish authorities first reported that ornamental petunias (Petunia x hybrida or P. hybrida) placed on the market were genetically modified (GM)1
The GM petunia sale as gardening plants in Finland was notified to the European Commission, who immediately informed all member states that no application for the import, cultivation, or marketing of GM petunia was ever submitted in the European Union (EU)
After testing a large number of different petunia varieties sold on the market, the official control laboratories revealed a large set of PCR screening results3

Summary

Introduction

In spring 2017 Finnish authorities first reported that ornamental petunias (Petunia x hybrida or P. hybrida) placed on the market were genetically modified (GM)1 These plants have an unusual orange flower color that could not have been generated by conventional breeding. In subsequent inspections and controls, various petunia varieties having orange, and otherwise unusual colored flowers were tested GM-positive by the official laboratories of the national competent authorities in the EU. It became apparent, that in several Member States the domestic breeding companies were affected and had unintentionally traded these GM petunia events in their downstream distribution and marketing chains for many years. The same situation was promptly revealed in the United States after it was officially announced, and in October 2017 it was reported that at least 124 varieties of GM petunias had been unintentionally imported and distributed interstate without proper US authorization

Methods

Results

Conclusion