Agricultural compositions comprising remodeled nitrogen fixing microbes
Inventors
REISINGER, MARK • SANDERS, ERNEST • BROGLIE, RICHARD • Temme, Karsten
Assignees
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Publication Number
US-11678668-B2
Publication Date
2023-06-20
Expiration Date
Abstract
The present disclosure provides non-intergeneric remodeled microbes that are able to fix atmospheric nitrogen and deliver such to plants in a targeted, efficient, and environmentally sustainable manner. The utilization of the taught microbial products will enable farmers to realize more productive and predictable crop yields without the nutrient degradation, leaching, or toxic runoff associated with traditional synthetically derived nitrogen fertilizer. The remodeled microbes taught herein are able to be combined with leading agricultural chemistry and elite germplasm. Furthermore, the disclosure provides seed treatments comprising remodeled microbes.
Core Innovation
The invention provides a composition comprising a plurality of non-intergeneric remodeled bacteria together with at least one plant hormone selected from auxins, cytokinins, gibberellins, abscisic acid, ethylene, brassinosteroids, jasmonic acid, and strigolactones. The non-intergeneric remodeled bacteria are configured to produce fixed N and to colonize plant roots, enabling targeted nitrogen delivery to non-leguminous crops. The platform is used in agricultural compositions, including seed-treatment formulations, that include the remodeled bacteria together with specified components.
The non-intergeneric remodeled bacteria include at least one genetic variation introduced into genes and associated regulatory elements including nifA, nifL, ntrB, ntrC, polynucleotide encoding glutamine synthetase, glnA, glnB, glnK, drat, amtB, polynucleotide encoding glutaminase, glnD, glnE, nifJ, nifH, nifD, nifK, nifY, nifE, nifN, nifU, nifS, nifV, nifW, nifZ, nifM, nifF, nifB, nifQ, and combinations thereof. The disclosure further includes nitrogen regulation gene elements including the nifLA operon, promoter insertions, and variants affecting GlnE and related nitrogen-regulation activities, including adenylyl-removing activity changes.
The invention further describes guided microbial remodeling and microbial breeding using genetic variation introduced into a gene selected from nitrogen fixation, assimilation, and nitrogenase-associated targets. The disclosed outcomes include fixed N production and root colonization ability, with quantification approaches described including acetylene reduction assay (ARA), ammonia excretion assays, isotopic approaches, and nitrogen delivery calculations based on colonization multiplied by activity.
Claims Coverage
Independent claim clm-00001 covers a composition combining a plurality of non-intergeneric remodeled bacteria with at least one plant hormone, while requiring quantitative thresholds for fixed N production and root colonization, and requiring specific genetic variations in the remodeled bacteria. The independent claim has three main inventive aspects: the composition of remodeled bacteria plus selected plant hormones, performance thresholds for fixed N and root colonization, and genetic variations introduced into selected nitrogen-related genes.
Composition of non-intergeneric remodeled bacteria plus plant hormone
A composition comprising a plurality of non-intergeneric remodeled bacteria and at least one plant hormone selected from auxins, cytokinins, gibberellins, abscisic acid, ethylene, brassinosteroids, jasmonic acid, and strigolactones.
Fixed N production and root colonization thresholds for remodeled bacteria
The non-intergeneric remodeled bacteria produce fixed N of at least about 1×10^-17 mmol N per bacterial cell per hour and have an average colonization ability per unit of plant root tissue of at least about 1.0×10^4 bacterial cells per gram of fresh weight of plant root tissue.
Genetic variations in specified nitrogen fixation and assimilation genes
A member of the plurality of non-intergeneric remodeled bacteria comprises at least one genetic variation introduced into a gene selected from nifA, nifL, ntrB, ntrC, polynucleotide encoding glutamine synthetase, glnA, glnB, glnK, drat, amtB, polynucleotide encoding glutaminase, glnD, glnE, nifJ, nifH, nifD, nifK, nifY, nifE, nifN, nifU, nifS, nifV, nifW, nifZ, nifM, nifF, nifB, nifQ, a gene associated with biosynthesis of a nitrogenase enzyme, and combinations thereof.
The independent claim centers on a composition that combines remodeled bacteria and selected plant hormones, where the remodeled bacteria meet fixed-N and root-colonization thresholds and carry genetic variations in a defined set of nitrogen fixation and nitrogen assimilation genes, including nifA, nifL, GlnE-related targets, amtB, and nitrogenase-associated targets.
Stated Advantages
Enhanced nitrogen fixation and root colonization, as reflected by the stated fixed N production and colonization ability thresholds.
Targeted nitrogen delivery to non-leguminous crops.
Documented Applications
Agricultural compositions, including seed-treatment formulations.
Use in agricultural contexts where the composition is associated with plant roots and with greenhouse/field outcomes, including root colonization readouts.
Targeted nitrogen delivery to non-leguminous crops.
Transgenic plants and agricultural pesticidal protein systems.
Plant-protectant and nutrient-related agricultural compositions including crop protectant additives, fertilizers, nitrogen stabilizers, and urease inhibitors.
Microbiome-associated nitrogen functions analyzed through Guided Microbial Remodeling workflows.
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