Developing a toolbox to engineer quantitative trait variation in legume species using CRISPR/Cas technologies.
Abstract
The impact of intensive agriculture on the environment is immense. This is especially dire with regard to the natural nitrogen (N) cycle, where the human driven interference, primarily associated with industrial fertilization, has reached unsustainable levels. Unlike cereals, legumes, such as soybean, alfalfa and common bean, have the ability to use atmospheric nitrogen, which limits the need for industrial fertilization. A more wide-spread use of legumes could alleviate some of the negative impacts on the biogeochemical cycle while also providing a useful alternative to meat consumption, an important factor in sustainability. To reach this goal, further improvements of legume crops with regard to their nitrogen economy and yield are essential. In the model legume, Lotus japonicus, the HAR1 receptor kinase plays a central role in the plant nitrogen and phosphate nutrition by regulating beneficial symbioses and root system architecture. In this thesis study, I used the HAR1 locus as a paradigm for the development of a CRISPR/Cas-based toolkit, with the ultimate goal of generating a range of synthetic variation at agriculturally important traits. While genome modifications at the HAR1 locus are yet to be demonstrated, the toolbox required to perform these experiments was developed. It should facilitate rapid expansion on the repertoire of alleles available for accelerated breeding of new, high yielding legume varieties that are better attuned with the natural environment.