Crop Improvement Through Targeted Genome Optimization Requires More Than Scissors

The crops that feed the world today are the product of hundreds of years of improvement through breeding by combining and selecting desired characteristics such as higher yield, resistances to disease and drought. In the past decades, the combination and selection of native traits through breeding was extended by mutation breeding, and more recently complemented with genetic engineering, enabling the introduction of novel genetic diversity beyond that originally present in the crops. Until recently, the introduction of mutations or new genes was random and the best performing combinations were obtained only after many rounds of laborious and costly analyses and selection steps.

Since early 2000, the crop improvement toolbox has been extended with the possibility to introduce mutations and new genes in a more targeted manner. Several technology concepts for such ‘targeted genome optimization' (TGO) have been developed. The concept based on targeting loci for improvement by local induction of a DNA double-strand break (DSB), and the specific optimization by utilizing the plant's natural DSB repair pathways, is to date the most versatile and efficient approach.

TGO through the directional repair of DSBs relies on the ability to induce DNA breaks very specifically at target positions selected for genetic improvement. The first ‘molecular scissor’ technology platform allowing such precise targeting were the zinc finger nucleases, followed by meganucleases, and more recently by TALENs and CRISPR-type nucleases. The availability of these molecular scissors, and certainly the simplicity offered by the latter two, has led to a rapid adoption of what most people refer as to 'genome editing' in crop genetics R&D, both in academia and industry.

Our view on the current status of plant TGO is that 3 types of optimization should be distinguished because they are not all equally well-developed for in all crops. While gene knock out through non-homologous end joining repair has been demonstrated with success in many crops, homologous repair that enables precise gene editing and allele replacement, as well as targeted gene insertion, is still inefficient in many crops. Thus, although we celebrate the ability to easily design and produce molecular scissors that, when having access to transformation technology and tissue culture expertise, enable the generation of plants with new properties due to gene KOs, we call for an increased effort in tackling the efficiency and accuracy of homology-directed repair to achieve a quantum leap towards precision plant breeding.

Academic and industry partners need to join forces to enable the technological advancements that will ensure the targeted and precise addition of genes, and the precise editing of native gene sequences, the latter of which is currently most challenging. Both aspects of targeted genome optimization have been shown to be feasible, but are yet still too low and very unpredictable in frequency and precision.

We will introduce the different TGO technologies and highlight the challenges we see towards developing those beyond the creation of gene knock outs, and capturing the full potential of TGO for near future crop improvement.