Analysis of the Unintended and Unexpected Effects of Transgene Insertion on the Endogenous Host Genome

Agriculture is highly susceptible to climate change effects and in Sub-Saharan Africa these rapid climate changes are predicted to introduce additional pressures to food security. Genetically modified (GM) crops with diverse traits are currently developed and produced to generate social, economic and environmental benefits focussed on resource-poor farmers. However, current safety assessments of these GM crops are thought to be biased as they focus on potential hazards posed by the inserted transgene and its products. As a result, a project was initiated to study the unintended effects of the integration of the transgene within the host genome. The Malus domestica polygalacturonase inhibiting protein 1 (Mdpgip1) gene was overexpressed in Solanum tuberosum cv BP1 for enhanced resistance against Verticillium wilt. Crude MdPGIP1 extracts prepared from the Mdpgip1 transgenic potato plant successfully inhibited the activities of Verticillium dahliae PGs (VdPGs) in in vitro studies showing that the transgene encodes an active protein. Genome walking studies revealed that the T-DNA containing the Mdpgip1 transgene was successfully inserted into the potato genome. Furthermore, genome walking revealed that no non-T-DNA sequences from the pCAMBIA2300 binary vector were inserted into the host potato genome during transformation. Genome walking studies further indicated the insertion site of the Mdpgip1 transgene as being adjacent to the photosystem Q_B gene on chromosome 1 of the potato genome. cDNA-RDA studies revealed that  the  insertion of the Mdpgip1 resulted in unintended effects in the nuclear genome that could affect the functions of the chloroplast (PsaC),  mitochondria (Afg3)  and ribosomes (25S rRNA).