Suppressing transgene silencing to enhance Engineering Biology – investigating the role of RDR6 in transgenic camelina via gene editing and metabolic engineering

Engineering Biology is predicated on genetic manipulation and being able to alter or rewrite a host genome. In plants, the ability to introduce foreign DNA and genes has been well established for over 40 years and has underpinned the GM revolution in agriculture. However, a well-recognized issue which affects transgenesis in plants is transgene silencing. This can take the form of transcriptional silencing or post-transcriptional gene silencing (PTGS). This is due in part to plants having a highly efficient innate surveillance system that detects non-native. One consequence of these powerful mechanisms to detect and inactive non-native sequences is that transgenic plants can lose their capacity to generate the trait they have been engineered with. Thus, there is significant interest in being able to mitigate transgene silencing not just for academic interest but also for translation and impact.  

Research at Rothamsted has developed transgenic camelina lines which have been engineered to accumulate the omega-3 long chain polyunsaturated fatty acids via seed-specific expression of up to eight foreign genes, and multiple events have been taken forward into field testing. However, we have observed several examples of lines showing transgene silencing (Han et al., 2020). We have also developed lines which are less susceptible to PTGS (based on the removal of provocative non-native sequences).  These plants and constructs will form part of the foundation on which this PhD project will be built.  In addition, we have recently used gene editing to generate rdr6 mutants in Camelina –In Arabidopsis, RDR6 (RNA-dependent RNA Polymerase 6) is a crucial enzyme in RNA silencing pathways, acting to copy RNA templates to produce double-stranded RNA, which are then further processed by enzymes such as Dicer (DCL4) and creating programming nucleotide sequences for various innate silencing systems. We hypothesize that expressing our complex transgenic cassettes for PUFAs in the rdr6 background will be significantly less susceptible to transgene silencing. This will be investigated both in the glasshouse and in the field via GM field trials. 

This project is expected to take place mainly at Rothamsted Research (Harpenden) with close collaboration with Royal Holloway University of London, and additional training and networking opportunities with the Engbio4GBP students at all partner institutions. In addition to training in specific research skills associated with the project, you will have a few days in-residence workshops for basic and more advanced training in AI/machine learning/computer-based data processing/programming/ research communication, commercialisation and scalability.