Access to fixed or available forms of nitrogen limits the productivity of crop plants and thus food production. Nitrogenous fertilizer production currently represents a significant expense for the efficient growth of various crops in the developed world. There are significant potential gains to be had from reducing dependence on nitrogenous fertilizers in agriculture in the developed world and in developing countries, and there is significant interest in research on biological nitrogen fixation and prospects for increasing its importance in an agricultural setting. Biological nitrogen fixation is the conversion of atmospheric N2 to NH3, a form that can be used by plants. However, the process is restricted to bacteria and archaea and does not occur in eukaryotes. Symbiotic nitrogen fixation is part of a mutualistic relationship in which plants provide a niche and fixed carbon to bacteria in exchange for fixed nitrogen. This process is restricted mainly to legumes in agricultural systems, and there is considerable interest in exploring whether similar symbioses can be developed in nonlegumes, which produce the bulk of human food. We are at a juncture at which the fundamental understanding of biological nitrogen fixation has matured to a level that we can think about engineering symbiotic relationships using synthetic biology approaches. This minireview highlights the fundamental advances in our understanding of biological nitrogen fixation in the context of a blueprint for expanding symbiotic nitrogen fixation to a greater diversity of crop plants through synthetic biology.
Engineering cereal crops that are self-supported by nitrogen fixation has been a dream since the 1970s when nitrogenase was transferred from Klebsiella pneumoniae to Escherichia coli. A renewed interest in this area has generated several new approaches with the common aim of transferring nitrogen fixation to cereal crops. Advances in synthetic biology have afforded the tools to rationally engineer microorganisms with traits of interest. Nitrogenase biosynthesis has been a recent target for the application of new synthetic engineering tools. Early successes in this area suggest that the transfer of nitrogenase and other supporting traits to microorganisms that already closely associate with cereal crops is a logical approach to deliver nitrogen to cereal crops.
The availability of fixed nitrogen is critical for life in many ecosystems, from extreme environments to agriculture. Due to the energy demands of biological nitrogen fixation, organisms must tailor their metabolism during diazotrophic growth to deliver the energy requirements to nitrogenase in the form of ATP and low potential electrons.
The extreme academic and social disruption caused by COVID-19 in the spring and summer of 2020 led to the loss of many student internships. We report here our creation of a novel internship for students majoring in the biological sciences.
A five-week research project was designed as part of a summer internship for high school students, and could also be used with educators or in introductory undergraduate research courses. This is a guided-inquiry-based project, framed within the significant issue of supplementing fertilizer use in agriculture with nitrogen-fixing microorganisms. This experience exposes students to how scientists are studying real-world problems; it teaches them basic research techniques, and promotes inquiry-based learning in a real research environment. It also fills a current gap in K-12 education that lacks enough microbiology emphasis. Research interns collect soil samples from various fields and use culture-dependent and culture-independent techniques to test whether there are nitrogen-fixing microorganisms that can be isolated and identified in each soil sample. Students work in a research laboratory making nitrogen-free media; culturing, isolating, and identifying microorganisms; extracting soil DNA; and amplifying the 16S rRNA and nifH genes. We administer a pre-test and a post-test, and students present their research both in a short talk and with a poster. By hosting high school students in a research laboratory and immersing them in laboratory science, we hope to inspire them to pursue a STEM-related career.
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