Methods and compositions for improving plant traits
Inventors
Temme, Karsten • Tamsir, Alvin • BLOCH, Sarah • CLARK, Rosemary • TUNG, Emily • Hammill, Kevin • Higgins, Douglas • Davis-Richardson, Austin
Assignees
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Publication Number
US-11565979-B2
Publication Date
2023-01-31
Expiration Date
Abstract
Methods and systems are provided for generating and utilizing a bacterial composition that comprises at least one genetically engineered bacterial strain that fixes atmospheric nitrogen in an agricultural system that has been fertilized with more than 20 lbs of Nitrogen per acre.
Core Innovation
The invention relates to a method of decreasing an amount of nitrogen-containing fertilizer required for producing a yield of a crop by inoculating soil of a field with a plurality of genetically engineered bacteria that fix atmospheric nitrogen. The genetically engineered bacteria comprise at least one genetic variation introduced into at least one gene, or non-coding polynucleotide, of a nitrogen fixation or assimilation genetic regulatory network, and are capable of fixing atmospheric nitrogen in the presence of exogenous nitrogen.
The plurality of genetically engineered bacteria 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 and produce fixed N at a rate of at least about 1×10^-17 mmol N per bacterial cell per hour. After inoculation, a crop is planted in the inoculated soil and a nitrogen-containing fertilizer is applied at no more than 90% of a dose, while the yield of the crop is the same as if 100% of the dose is applied under similar conditions in the absence of the plurality of genetically engineered bacteria.
The document also describes microbial breeding and evolution workflows, including host-microbe evolution and trait screening and selection, and describes microbial identification using 16S and diagnostic non-native junction qPCR. It further describes nitrogen fixation or nitrogen delivered measurement frameworks, including quantifying colonization and nitrogen fixation using assays such as isotope tracing and acetylene reduction assay, and presents supporting examples including strain colonization, gene expression, ammonia excretion, field yield comparisons, and deposited strains under the Budapest Treaty with accession and strain lineage information.
Claims Coverage
The claim coverage centers on a fertilizer-reduction method using genetically engineered atmospheric nitrogen-fixing bacteria with defined performance thresholds and yield equivalence. Three inventive features are consistently presented: engineered bacteria with variation in a nitrogen fixation or assimilation regulatory network, colonization and fixed-N production thresholds, and reduced nitrogen-containing fertilizer application to no more than 90% of a dose while preserving yield.
Inoculation with atmospheric nitrogen-fixing genetically engineered bacteria
Inoculating soil of a field with a plurality of genetically engineered bacteria that fix atmospheric nitrogen, wherein the genetically engineered bacteria comprise at least one genetic variation introduced into at least one gene, or non-coding polynucleotide, of a nitrogen fixation or assimilation genetic regulatory network, such that the genetically engineered bacteria are capable of fixing atmospheric nitrogen in the presence of exogenous nitrogen.
Bacterial colonization and fixed-N production thresholds
The plurality of genetically engineered bacteria 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 and produce fixed N at a rate of at least about 1×10^-17 mmol N per bacterial cell per hour.
Planting and reduced fertilizer dose with yield equivalence
Planting a crop in the inoculated soil and applying a nitrogen-containing fertilizer corresponding to no more than 90% of a dose, where the yield of the crop is the same as if 100% of the dose of nitrogen-containing fertilizer is applied under similar conditions between planting and harvesting the crop in the absence of the plurality of genetically engineered bacteria.
Overall, the claim coverage requires genetically engineered atmospheric-nitrogen-fixing bacteria with genetic variation in nitrogen fixation or assimilation genetic regulatory networks, defined colonization and fixed-N production thresholds, and reduced nitrogen fertilizer application while maintaining crop yield equivalence relative to a full-dose control without the engineered bacteria.
Stated Advantages
Decreases the amount of nitrogen-containing fertilizer required for producing a yield of a crop while maintaining the same yield as if 100% of the nitrogen-containing fertilizer dose were applied in the absence of the plurality of genetically engineered bacteria.
Avoids intergeneric engineered microbes to reduce regulatory and adoption barriers.
Documented Applications
An agricultural system use case in which a crop is planted in field soil inoculated with genetically engineered atmospheric-nitrogen-fixing bacteria, enabling nitrogen-containing fertilizer dose reduction while producing the same yield as a full-dose fertilizer comparison without the bacteria.
Reducing the amount of nitrogen-containing fertilizer needed to produce a crop yield while maintaining the same yield under similar conditions.
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