Plants with higher yield, biomass, and stress tolerance.

Description

"PLANTS WITH HIGHER YIELD, BIOMASS" "AND STRESS TOLERANCE" DESCRIPTIVE REPORT WITH REFERENCE TO RELATED REQUESTS

[0001] This Request claims priority over the Provisional Request of United States Patent No. 63 / 494448, filed April 5, 2023, the full contents of which are incorporated herein by reference. TECHNICAL FIELD

[0002] The present invention is directed to plants that exogenously express the polynucleotide(s), nucleic acid construct(s), polypeptide(s) of the invention and methods for producing them. In particular, the present invention is directed to the introduction of polynucleotide(s), nucleic acid construct(s) and / or polypeptide(s) of the invention into plant(s) to increase the biomass and / or yield, and / or drought tolerance of the plant(s). SEQUENCE LISTING REFERENCE EMBEDDING

[0003] This Application is being filed together with a Sequence Listing in electronic format. The Sequence Listing is provided as a file entitled INGRN.005WO_ST26.xml, created on April 2, 2024, with a size of 108,117 bytes. The information in the electronic Sequence Listing is incorporated herein by reference in its entirety. BACKGROUND

[0004] Changes in climate conditions, population growth, and food shortages represent a challenge Petition 870250109327, dated 11 / 28 / 2025, page 5 / 107 2 / 78 formidable for agriculture. Conventional methods for improving crops and horticultural plants have often used selective breeding technologies to identify plants with desired properties. This type of selective breeding technology has several disadvantages, as it requires a lot of labor and results in plants with heterogeneous genetic complements.

[0005] In agriculture, genetic engineering has led to crops with a variety of advantageous improvements, including yield, biomass, pest resistance, herbicide resistance, vigor, and stress tolerance. Transgenic varieties in commercial use have reduced the use of chemical pesticides by 37%, increased crop yield by 22%, and increased farmers' profits by 68%. Most current transgenic technologies, however, increase yield indirectly through their anti-insect and anti-herbicide properties, etc. Due to increasing needs and challenges, there remains a strong need for the development of plants that can withstand their constantly changing environment while simultaneously increasing yield and biomass. SUMMARY OF THE INVENTION

[0006] In some respects, the techniques described herein refer to a plant into which a nucleic acid molecule encoding a factor associated with fat mass and obesity (FTO) is introduced, wherein the FTO has at least the following domains: nuclear localization signal, promoter, coding sequence and terminator sequence.

[0007] In some respects, a plant into which a nucleic acid molecule encoding an FTO is introduced, in which the FTO has at least the following domains: a promoter, a nuclear localization signal, a coding sequence, and a terminator.

[0008] In other respects, the FTO nucleic acid molecule is derived from vertebrates, invertebrates, algae, or oomycetes. Petition 870250109327, dated 11 / 28 / 2025, p. 6 / 107 3 / 78

[0009] In some respects, the plant contains a nucleic acid molecule that encodes an FTO that has at least 80% identity with any of the SEQ IDs 34-38.

[0010] The plant exhibits at least a twofold increase in biomass compared to a plant that does not contain the nucleic acid molecule in some respects. In other respects, the plant exhibits at least a twofold increase in productivity compared to a plant that does not contain the nucleic acid molecule.

[0011] The plant, in some respects, is selected from the group that consists of Arabadopsis, canola, wheat, corn, soybean, millet, sorghum, cotton, tomato, grape, peanut, rice, lettuce, melon, sweet potato, carrot, cucumber, sugarcane, alfalfa, Camelina or PennyCress.

[0012] In some respects, a plant, plant cell, tissue or seed, each having in its genome a nucleic acid molecule comprising a nucleotide sequence of SEQ ID NO: 34-38. In other respects, the plant, plant cell, tissue or seed has a nucleotide sequence with at least 80% homology with any of the SEQ ID NOs 34-38.

[0013] The nucleic acid molecule also has a promoter, a nuclear localization signal, a coding sequence and a termination sequence, and is expressed in the nucleus in some respects.

[0014] The plant that expresses the nucleic acid sequence shows at least a twofold increase in seed yield, biomass or yield in some respects.

[0015] The plant that expresses the nucleic acid sequence is selected from the group consisting of canola, wheat, soybean, millet, sorghum, cotton, tomato, grape, peanut, rice, lettuce, melon, sweet potato, carrot, cucumber, sugarcane, alfalfa, Camelina or PennyCress.

[0016] In other respects, a plant that expresses the amino acid sequence has at least 90% homology with any of the SEQ IDs 6-30 and 39-71. The plant also expresses the amino acid sequence in the nucleus in some respects. Petition 870250109327, dated 11 / 28 / 2025, p. 7 / 107 4 / 78

[0017] The plant that expresses the amino acid sequence exhibits at least a twofold increase in seed yield, biomass, or yield in some respect and may be selected from the group consisting of canola, wheat, soybean, millet, sorghum, cotton, tomato, grape, peanut, rice, lettuce, melon, sweet potato, carrot, cucumber, sugarcane, alfalfa, Camelina, or PennyCress.

[0018] In some embodiments, a plant comprising plant cells containing a nucleic acid molecule encoding a fat mass and obesity-associated factor (FTO) gene is provided. In some embodiments, the expression of said nucleic acid molecule results in one or more of the following increases: yield, biomass, growth rate, drought tolerance, and abiotic stress tolerance compared to a plant of the same variety without the nucleic acid molecule encoding the FTO gene. In some embodiments, the FTO nucleic acid molecule is from vertebrates, invertebrates, algae, or oomycetes. In some embodiments, the nucleic acid molecule further comprises a promoter, a nuclear localization signal, a coding sequence, and a termination sequence.

[0019] In some embodiments, the nucleic acid molecule encoding an FTO has at least 80% identity with any of the SEQ IDs 34-38.

[0020] In some embodiments, the plant exhibits at least a twofold increase in biomass compared to the plant that does not contain the nucleic acid molecule. In some embodiments, the plant exhibits at least a twofold increase in productivity compared to the plant that does not contain the nucleic acid molecule. In some embodiments, the plant exhibits increased drought tolerance compared to the plant that does not contain the nucleic acid molecule.

[0021] In some modalities, the plant is selected from the group consisting of Arabadopsis, canola, wheat, corn, soybean, millet, sorghum, cotton, tomato, grape, peanut, rice, lettuce, melon, sweet potato, Petition 870250109327, dated 11 / 28 / 2025, p. 8 / 107 5 / 78 Carrot, cucumber, sugarcane, alfalfa, Camelina or Penny Cress.

[0022] In some embodiments, the FTO gene is expressed in a nucleus of said plant cells.

[0023] In some embodiments, the FTO gene encodes a polypeptide comprising either of the SEQ IDs 6-30 or 3971 or a polypeptide comprising at least 90% identity with this.

[0024] In some embodiments, a method is provided for generating a modified plant comprising increased yield, growth rate, biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency, drought tolerance and / or abiotic stress tolerance compared with a wild-type plant of the same variety. In some embodiments, the method comprises: (a) genetically modifying a plant cell with an exogenous polynucleotide encoding a polypeptide comprising an amino acid sequence with at least 80% identity to the selected sequence from the group comprising any of the SEQ ID NOs: 6-30 and 39-71, and (b) regenerating the modified plant from said plant cell modified with an exogenous polynucleotide. In some embodiments, the method further comprises producing modified plants from the plant produced in step (b).In some forms, the polypeptide originates from vertebrates, invertebrates, algae, or oomycetes.

[0025] In some embodiments, the modified plant exhibits at least a two-fold increase in biomass compared to a plant that does not contain the nucleic acid molecule. In some embodiments, the modified plant exhibits at least a two-fold increase in productivity compared to a plant that does not contain the nucleic acid molecule. In some embodiments, the modified plant exhibits increased drought tolerance compared to a plant that does not contain the nucleic acid molecule. In some Petition 870250109327, dated 11 / 28 / 2025, p. 9 / 107 6 / 78 varieties, the plant is selected from the group that consists of Arabadopsis, canola, wheat, corn, soybean, millet, sorghum, cotton, tomato, grape, peanut, rice, lettuce, melon, sweet potato, carrot, cucumber, sugarcane, alfalfa, Camelina or PennyCress. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Some embodiments of the invention are described herein by way of example only, with reference to the accompanying drawings. The drawings are for illustrative purposes only and are for discussion purposes only.

[0027] FIG. 1 is a schematic illustration of the pET28a(+) plasmid containing the T7 promoter used to express the polynucleotide sequences of the invention, as in some embodiments of the invention.

[0028] FIG. 2 is a schematic illustration of the pET28a plasmid containing acFTO, SEQ ID No. 9, as in one embodiment of the invention.

[0029] FIG. 3 is a gel image illustrating the expression of FTO homologs in E. coli after induction, as in some embodiments of the invention.

[0030] FIG. 4 is a schematic representation of root length in Seteria plants transformed with hFTO-GFP and mcFTO (SEQ ID No. 7), as in an embodiment of the invention.

[0031] FIG. 5 is a diagram illustrating the seed weights by event and gene of interest (GOI), as in one embodiment of the invention.

[0032] FIG. 6 is a diagram illustrating the expression of the transgene FTO in Seteria expressing the SEQ IDs n°s 1, 2 and 7, as in an embodiment of the invention.

[0033] FIG. 7 is a diagram illustrating the root length in Seteria expressing SEQ ID No. 7, as in a form of Petition 870250109327, dated 11 / 28 / 2025, p. 10 / 107 7 / 78 invention.

[0034] FIG. 8 is a diagram representing the Seteria biomass transformed with SEQ IDs no. 9, 6, 8 and 3, as in an embodiment of the invention.

[0035] FIG. 9 is a diagram representing the production of Seteria seeds transformed with SEQ IDs no. 9, 6, 8 and 3, as in one embodiment.

[0036] FIG. 10 is a diagram representing the expression of the SEQ ID No. 1 in soybeans, as in an embodiment of the invention.

[0037] FIG. 11A illustrates images of drought-treated soybeans with an empty vector control (FIG. 11A) and soybeans transformed with SEQ ID no. 8 (FIG. 11B). FIG. 11B illustrates images of drought-treated soybeans with an empty vector control (FIG. 11A) and soybeans transformed with SEQ ID No. 8 (FIG. 11B).

[0038] FIG. 12A illustrates images of drought-treated soybeans with an empty vector control (FIG. 12A), soybeans transformed with SEQ ID No. 7 (FIG. 12B), and soybeans transformed with SEQ ID No. 7 (FIG. 12C). FIG. 12B illustrates images of drought-treated soybeans with an empty vector control (FIG. 12A), soybeans transformed with SEQ ID #7 (FIG. 12B), and soybeans transformed with SEQ ID #7 (FIG. 12C). FIG. 12C illustrates images of drought-treated soybeans with an empty vector control (FIG. 12A), soybeans transformed with SEQ ID No. 7 (FIG. 12B), and soybeans transformed with SEQ ID No. 7 (FIG. 12C).

[0039] FIG. 13 illustrates the expression of the hFTO acFTO, niFTO protein detected by Western Blot in soybean plants.

[0040] FIG. 14 illustrates the detection of hFTO-GFP expression in soybean plant nuclei. Petition 870250109327, dated 11 / 28 / 2025, page 11 / 107 8 / 78

[0041] FIG. 15A illustrates the position of the plants used for the drought trial on transformed soybean plants with SEQ ID NO: 9. FIG. 15A shows a map used to position the correct plant in the correct location for the experiment and FIG. 15B shows the well-irrigated and dried plants surrounded by the soybean border to avoid edge effects. FIG. 15B illustrates the position of the plants used for the drought trial on transformed soybean plants with SEQ ID NO: 9. FIG. 15A shows a map used to position the correct plant in the correct location for the experiment, and FIG. 15B shows the well-irrigated and dry plants surrounded by the soybean border to avoid edge effects.

[0042] FIG. 16A illustrates a method of image acquisition for determining wilting score and FIG. 16B illustrates the root length of well-irrigated and dry soybean plants transformed with SEQ ID NO: 9.

[0043] FIG. 17A illustrates the drought test on soybean plants transformed with SEQ ID NO: 9. FIG. 17A shows a map used to position the correct plant in the correct position for the experiment, FIG. 17B shows the wilting score of the leaves of well-irrigated and dry soybean plants, and FIG. 17C shows the root length of well-irrigated and dry soybean plants, transformed with SEQ ID NO: 9. FIG. 17B illustrates the drought test in soybean plants transformed with SEQ ID NO: 9. FIG. 17A shows a map used to position the correct plant in the correct location for the experiment, FIG. 17B shows the wilting score of the leaves of well-irrigated and dry soybean plants, and FIG. 17C shows the root length of well-irrigated and dry soybean plants transformed with SEQ ID NO: 9. FIG. 17C illustrates the drought test on soybean plants. Petition 870250109327, dated 11 / 28 / 2025, page 12 / 107 9 / 78 transformed with SEQ ID NO: 9. FIG. 17A shows a map used to position the correct plant in the correct location for the experiment, FIG. 17B shows the wilting score of the leaves of well-irrigated and dry soybean plants, and FIG. 17C shows the root length of well-irrigated and dry soybean plants, transformed with SEQ ID NO: 9.

[0044] FIG. 18A illustrates the drought test in soybean plants transformed with SEQ ID NO: 7. FIG. 18A shows an aerial image of the plants, FIG. 18B shows a side view of the plants, FIG. 18C shows the root morphology of well-irrigated and dry soybean plants transformed with SEQ ID NO: 7, and FIG. 18C shows the root morphology of well-irrigated and dry soybean plants transformed with SEQ ID NO: 7. FIG. 18D shows the root length of well-irrigated and dry soybean plants transformed with SEQ. ID NO. 7 and FIG. 18E shows the wilting score of the leaves of well-irrigated and dry soybean plants transformed with SEQ. ID NO. 7. FIG. 18B illustrates the drought test in soybean plants transformed with SEQ ID NO: 7. FIG. 18A shows an aerial image of the plants, FIG. 18B shows a side view of the plants, FIG. 18C shows the root morphology of well-irrigated and dry soybean plants transformed with SEQ ID NO: 7, and FIG. 18C shows the root morphology of well-irrigated and dry soybean plants transformed with SEQ ID NO: 7. FIG. 18D shows the root length of well-irrigated and dry soybean plants transformed with SEQ. ID NO. 7, and FIG. 18E shows the wilting score of the leaves of well-irrigated and dry soybean plants transformed with SEQ. ID NO. 7. Figure 18C illustrates the drought test in soybean plants transformed with SEQ ID NO: 7. Figure 18A shows an aerial image of the plants, Figure 18B shows a side view of the plants, Figure 18C shows the root morphology of well-irrigated and dry soybean plants transformed with SEQ ID NO: 7, and Figure 18C shows the morphology of the Petition 870250109327, dated 11 / 28 / 2025, page 13 / 107 Figure 18D shows the root length of well-irrigated and dry soybean plants transformed with SEQ ID NO. 7, and Figure 18E shows the wilting score of well-irrigated and dry soybean plant leaves transformed with SEQ ID NO. 7. FIG. 18D illustrates the drought test in soybean plants transformed with SEQ ID NO: 7. FIG. 18A shows an aerial image of the plants, FIG. 18B shows a side view of the plants, FIG. 18C shows the root morphology of well-irrigated and dry soybean plants transformed with SEQ ID NO: 7, and FIG. 18C shows the root morphology of well-irrigated and dry soybean plants transformed with SEQ ID NO: 7. FIG. 18D shows the root length of well-irrigated and dry soybean plants transformed with SEQ. ID NO. 7, and FIG. 18E shows the wilting score of the leaves of well-irrigated and dry soybean plants transformed with SEQ. ID NO. 7. FIG. 18E illustrates the drought test in soybean plants transformed with SEQ ID NO: 7. FIG. 18A shows an aerial image of the plants, FIG. 18B shows a side view of the plants, FIG. 18C shows the root morphology of well-irrigated and dry soybean plants transformed with SEQ ID NO: 7, and FIG. 18D shows the root length of well-irrigated and dry soybean plants transformed with SEQ. ID NO. 7, and FIG. 18E shows the wilting score of the leaves of well-irrigated and dry soybean plants transformed with SEQ. ID NO. 7. DETAILED DESCRIPTION

[0045] Previous epigenetic research has generally focused on reversible DNA and histone modifications. Although hundreds of RNA modifications have been discovered, N6-methyladenoesin Petition 870250109327, dated 11 / 28 / 2025, page 14 / 107 11 / 78 (m6A) was considered the most abundant RNA modification in mRNA across all eukaryotes.

[0046] The mRNAs of all plants contain the N6-methyladenoesin (m6A) modification. m6A regulates the metabolic processing of mRNAs, including splicing, nuclear export, stability, and protein translation through m6A-binding proteins. The modifications are produced by m6A-modifying enzymes (the currently reported modifying enzyme subunits are classified as MTA (METTL3 homolog gene) and FIP37 (WTAP homolog gene)) and have an important regulatory effect on plant growth and development. Although the modification has been known for over forty years, its function was not known until the demethylase of m6A, FTO protein, was characterized (Jia et al., N6-methyladenosine in nuclear RNA is an important substrate of FTO associated with obesity. Nat Chem Biol, 2011, 7 (12): 885-887), reporting the reversibility of the RNA modification for the first time and initiating “RNA epigenetics (or epitranscriptome)” research in 2011.

[0047] During the demethylation process of m6A in RNA, FTO produces two relatively stable new modifications, hm6A (N6-hydroxymethyladenosine) and f6A (N6-formyladenosine), which have potential regulatory effects on RNA processing (Fu et al., FTO-mediated formation of N6-hydroxymethyladenosine and N6-formyladenosine in mammalian RNA. Nat Commun, 2013, 4:1798). Given the importance of this modification, there are advantages in discovering FTO homologs that produce higher yield and biomass in other plants. However, such efforts are difficult and time-consuming, since cellular localization, transcription efficiency, species specificity, and phenotypic effects cannot be predicted due to variations in FTO sequences between species.

[0048] In some embodiments, FTO homologs are provided that increase biomass, yield and stress tolerance in crops disclosed herein. Polypeptides are also provided herein. Petition 870250109327, dated 11 / 28 / 2025, page 15 / 107 12 / 78 FTO homologs, their compositions and methods for developing plants with enhanced beneficial characteristics, including yield, growth rate, biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity and / or tolerance to abiotic stress. Definitions

[0049] The illustrative embodiments described in the detailed description, drawings and claims are not intended to be limiting. Other embodiments may be used and other alterations may be made, without departing from the scope of the subject matter presented here. It is easy to understand that the aspects of this disclosure, as described here in general terms and illustrated in the figures, may be arranged, substituted, combined, separated and projected in a wide variety of different configurations, all explicitly contemplated herein.

[0050] Unless otherwise defined, all technical and scientific terms used herein have the same meaning commonly understood by someone with common knowledge of the art to which the invention relates.

[0051] The terms “compreende”, “compreendendo”, “inclui”, “incluindo”, “tendo” and their conjugates mean “including, but without limitation”.

[0052] The term “consisting of” means “including and limited to”.

[0053] The term “essentially consisting of” means that the composition, method or structure may include additional ingredients, steps and / or parts, but only if the additional ingredients, steps and / or parts do not materially alter the basic and innovative characteristics of the claimed composition, method or structure. Petition 870250109327, dated 11 / 28 / 2025, page 16 / 107 13 / 78

[0054] As used herein, the singular forms “a”, “an” and “the” include plural references, unless the context clearly indicates otherwise. For example, the term “a compound” or “at least one compound” may include a plurality of compounds, including mixtures thereof.

[0055] Throughout this application, various embodiments of this invention may be presented in range format. It should be understood that the range description is merely for convenience and brevity and should not be interpreted as an inflexible limitation to the scope of the invention. Consequently, the description of a range should be considered as having specifically disclosed all possible sub-ranges as well as individual numerical values ​​within that range. For example, the description of a range such as 1 to 6 should be considered as having specifically disclosed sub-ranges such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the range width.

[0056] Whenever a numerical range is indicated here, it must include any quoted numeral (fractional or integer) within the indicated range. The expressions “ranging between” a first indicative number and a second indicative number and “ranging from” a first indicative number “to” a second indicative number are used here interchangeably and are intended to include the first and second numbers indicated and all fractional and integer numerals between them.

[0057] As used here, the term “method” refers to ways, means, techniques, and procedures for performing a particular task, including, but not limited to, those ways, means, techniques, and procedures known to, or readily developed from, ways, means, techniques, and procedures known to practitioners of the chemical, pharmacological, biological, biochemical, and medical arts.

[0058] When referring to sequence listings Petition 870250109327, dated 11 / 28 / 2025, page 17 / 107 14 / 78 specific, such reference should be understood as also encompassing sequences that substantially correspond to their complementary sequence, including small sequence variations resulting from, for example, sequencing errors, cloning errors, or other alterations that result in base substitution, base deletion, or base addition, provided that the frequency of such variations is less than 1 in 50 nucleotides, alternatively, less than 1 in 100 nucleotides, alternatively, less than 1 in 200 nucleotides, alternatively, less than 1 in 500 nucleotides, alternatively, less than 1 in 1000 nucleotides, alternatively, less than 1 in 5,000 nucleotides, alternatively, less than 1 in 10,000 nucleotides.

[0059] As used herein, the term “plant yield” refers to the quantity (e.g., determined by weight, volume, processed product, or size) or quantity (number) of tissues or organs produced by a plant or by a growing season. Therefore, increased yield can affect the economic benefit that can be obtained from the plant in a given growing area and / or growing period.

[0060] The term “about,” as used herein, means greater than or less than the stated value or range of values ​​by 10 percent, but is not intended to designate any value or range of values ​​solely for this broader definition. Each value or range of values ​​preceded by the term “about” is also intended to encompass the concretization of the stated absolute value or range of values.

[0061] As used herein, the term “polynucleotide” refers to a single- or double-stranded nucleic acid sequence that is isolated and provided in the form of an RNA sequence, a complementary polynucleotide sequence (cDNA), a genomic polynucleotide sequence, and / or a composite polynucleotide sequence (e.g., a combination of the foregoing).

[0062] The term “isolated” refers to at least partially separated from the natural environment, for example, from a plant cell. Petition 870250109327, dated 11 / 28 / 2025, p. 18 / 107 15 / 78

[0063] As used herein, the term “complementary polynucleotide sequence” refers to a sequence resulting from the reverse transcription of messenger RNA using a reverse transcriptase or any other RNA-dependent DNA polymerase. Such a sequence can be further amplified in vivo or in vitro using a DNA-dependent DNA polymerase.

[0064] As used herein, the term “genomic polynucleotide sequence” refers to a derived (isolated) sequence from a chromosome and therefore represents a contiguous part of a chromosome.

[0065] As used herein, the term “compound polynucleotide sequence” refers to a sequence that is at least partially complementary and at least partially genomic. A compound sequence may include some exonal sequences necessary to encode the polypeptide of the present invention, as well as some intronic sequences interposed between them. The intronic sequences may be from any source, including other genes, and will typically include conserved splicing signal sequences. Such intronic sequences may also include cis-acting expression regulatory elements. The nucleic acid sequences encoding the polypeptides of the present invention may be optimized for expression.Examples of such sequence modifications include, among others, an altered G / C content to more closely approximate that typically found in the plant species of interest and the removal of codons atypically found in the plant species, commonly referred to as codon optimization.

[0066] The term “codon optimization” refers to the selection of appropriate DNA nucleotides for use in a structural gene or fragment thereof that approximates the codon usage in the plant of interest. Therefore, an optimized gene or nucleic acid sequence refers to a gene in which the nucleotide sequence of a native or naturally occurring gene has been modified to utilize codons. Petition 870250109327, dated 11 / 28 / 2025, page 19 / 107 16 / 78 statistically preferred or statistically favored in the plant. The naturally occurring coding nucleotide sequence may already, before any modification, contain a number of codons that correspond to a statistically favored codon in a specific plant species. Therefore, codon optimization of the native nucleotide sequence may involve determining which codons within the native nucleotide sequence are not statistically favored with respect to a specific plant, and modifying those codons according to a codon usage table of the specific plant to produce a codon-optimized derivative. A modified nucleotide sequence may be wholly or partially optimized for codon usage in the plant, provided that the protein encoded by the modified nucleotide sequence is produced at a higher level than the protein encoded by the corresponding native or naturally occurring gene.The construction of synthetic genes through the alteration of codon usage is described, for example, in Patent Application PCT 93 / 07278, which is incorporated herein by reference in its entirety. Thus, the invention encompasses the nucleic acid sequences described above; fragments thereof, hybridizable sequences with them, sequences homologous to them, sequences encoding similar polypeptides using different codons, altered sequences characterized by mutations, such as deletion, insertion or substitution of one or more nucleotides, occurring naturally or induced by humans, in a random or targeted manner.

[0067] As used in this document, the term “plant” includes an entire plant and any offspring, cell, tissue, or part of a plant. The term “plant parts” includes any part(s) of a plant, including, for example and without limitation: seed (including mature seed and immature seed); a plant cutting; a plant cell; a plant cell culture; a plant organ (e.g., pollen, embryos, flowers, fruits, buds, leaves, roots, stems, and explants). A plant tissue or plant organ may be a seed, callus, or Petition 870250109327, dated 11 / 28 / 2025, p. 20 / 107 17 / 78 any other group of plant cells organized into a structural or functional unit. A plant cell or tissue culture may be able to regenerate a plant with the physiological and morphological characteristics of the plant from which the cell or tissue was obtained, and to regenerate a plant with substantially the same genotype as the plant. In contrast, some plant cells are not capable of being regenerated to produce plants. Regenerable cells in a plant cell or tissue culture may be embryos, protoplasts, meristematic cells, calluses, pollen, leaves, anthers, roots, root tips, silk, flowers, grains, ears, cobs, husks, or stems.

[0068] Plant parts include harvestable parts and parts useful for propagation of offspring plants. Plant parts useful for propagation include, for example and without limitation: seed; fruit; cutting; seedling; tuber; and rootstock. A harvestable part of a plant may be any useful part of a plant, including, for example and without limitation: flower; pollen; seedling; tuber; leaf; stem; fruit; seed; and root.

[0069] A plant cell is the structural and physiological unit of the plant. Plant cells, as used herein, include protoplasts and cell-walled protoplasts. A plant cell may be in the form of a single isolated cell or an aggregate of cells (e.g., a friable callus and a cultured cell) and may be part of a higher organized unit (e.g., a plant tissue, a plant organ, and a plant). Thus, a plant cell may be a protoplast, a gamete-producing cell, or a cell or set of cells that can regenerate into an entire plant. Thus, a seed, which comprises multiple plant cells and is capable of regenerating into an entire plant, is considered a “plant part” in the embodiments presented herein.

[0070] The plant can be in any form, including suspension cultures, embryos, meristematic regions, callus tissue, leaves, gametophytes, sporophytes, pollen, and microspores. Petition 870250109327, dated 11 / 28 / 2025, p. 21 / 107 18 / 78

[0071] It should be noted that the yield of a plant can be affected by several parameters, including, but not limited to, plant biomass; plant vigor, stress tolerance, growth rate; seed yield; quantity of seeds or grains; seed or grain quality; oil yield; oil, starch and / or protein content in harvested organs (e.g., seeds or vegetative parts of the plant); number of flowers (florets) per panicle (expressed as a ratio between the number of filled seeds and the number of primary panicles); harvest index; number of plants grown per area; number and size of harvested organs per plant and per area; number of plants per growing area (density); number of organs harvested in the field; total leaf area; carbon assimilation and partitioning (the distribution / allocation of carbon within the plant); shade resistance; number of harvestable organs (e.g., seeds), seeds per pod, weight per seed;and modified architecture [such as increased stem diameter, thickness, or improved physical properties (e.g., elasticity)].

[0072] As used herein, the term “seed yield” refers to the number or weight of seeds per plant, seeds per pod or per growing area, or the weight of a single seed, or the oil extracted per seed. Therefore, seed yield can be affected by seed dimensions (e.g., length, width, perimeter, area and / or volume), number of (filled) seeds and seed filling rate, and seed oil content. Therefore, increasing seed yield per plant can affect the economic benefit that can be obtained from the plant in a given growing area and / or growing season; and increasing seed yield per growing area can be achieved by increasing seed yield per plant and / or increasing the number of plants grown in the same area.

[0073] The term “seed” (also called “grain” or “corn”), as used herein, refers to a small embryonic plant. Petition 870250109327, dated 11 / 28 / 2025, page 22 / 107 19 / 78 enclosed in a covering called the integument (usually containing some stored food), the product of the matured ovule of gymnosperm and angiosperm plants, which occurs after fertilization and some growth within the mother plant.

[0074] The term “oil content,” as used herein, refers to the amount of lipids in a given plant organ, whether in the seeds (seed oil content) or in the vegetative part of the plant (vegetative oil content), and is typically expressed as a percentage of dry weight (10% seed moisture) or wet weight (for the vegetative part). It should be noted that oil content is affected by the intrinsic oil production of a tissue (e.g., seed, vegetative part), as well as by the mass or size of the oil-producing tissue per plant or per growing period. In one embodiment, increasing the plant oil content can be achieved by increasing the size / mass of the oil-containing plant tissue(s) per growing period. Thus, increasing the oil content of a plant can be achieved by increasing yield, growth rate, biomass, and plant vigor.

[0075] As used herein, the term “plant biomass” refers to the amount (e.g., measured in grams of air-dried tissue) of a plant’s tissue produced in a growing season, which may also determine or affect plant yield or yield per area of ​​cultivation. An increase in plant biomass may occur in the whole plant or in parts thereof, such as aerial (harvestable) parts, vegetative biomass, roots, and seeds.

[0076] As used herein, the term “root biomass” refers to the total weight of the plant’s root(s). Root biomass can be determined directly by weighing the total root material (fresh and / or dry weight) of a plant. Additionally or alternatively, root biomass can be determined indirectly by measuring the root cover, root density, and / or root length of a plant. It should be noted that plants with greater root cover Petition 870250109327, dated 11 / 28 / 2025, page 23 / 107 Plants with 20 / 78 exhibit greater efficiency in fertilizer use (e.g., nitrogen) and / or greater efficiency in water use compared to plants with less root cover.

[0077] As used herein, the term “root cover” refers to the total area or volume of soil or any growing medium covered by the roots of a plant. According to some embodiments of the invention, root cover is the minimum convex volume covered by the plant roots.It should be noted that, since each plant has a characteristic root system—for example, some plants have a shallow root system (e.g., only a few centimeters below the soil level), while others have a deep root system in the soil (e.g., several tens of centimeters or several meters deep in the soil below the soil level)—the measurement of a plant's root cover can be performed at any depth in the soil or growing medium of the plant, and the comparison of root cover between plants of the same species (e.g., a transgenic plant exogenously expressing the polynucleotide of some embodiments of the invention and a control plant) should be performed by measuring the root cover at the same depth. According to some embodiments of the invention, root cover is the minimum convex area covered by the roots of a plant at a specific depth.A non-limiting example of measuring root cover is shown in FIGS. 4 and 7. As used herein, the term “root density” refers to the density of roots in a given area (e.g., soil area or any plant growing medium). Root density can be determined by counting the number of roots per predetermined area at a predetermined depth (in units of number of roots per area, e.g., mm2, cm2, or m2).

[0078] As used herein, the expression “root length” refers to the total length of the longest root of a single plant.

[0079] As used herein, the expression “growth rate” Petition 870250109327, dated 11 / 28 / 2025, page 24 / 107 "21 / 78 of root length" refers to the variation in total root length per plant per unit of time (e.g., per day).

[0080] As used herein, the expression “growth rate” refers to the increase in the size of the plant organ / tissue over time (it can be measured in cm2 per day or cm / day).

[0081] As used herein, the expression “photosynthetic capacity” (also known as “Amax”) is a measure of the maximum rate at which leaves are able to fix carbon during photosynthesis. It is typically measured as the amount of carbon dioxide fixed per square meter per second, for example, as pmol m-2 s-1. Plants are able to increase their photosynthetic capacity by several modes of action, such as increasing the total leaf area (e.g., increasing leaf area, increasing the number of leaves, and increasing plant vigor, e.g., the plant's ability to develop new leaves over time), as well as increasing the plant's ability to efficiently perform carbon fixation in the leaves. Therefore, the increase in total leaf area can be used as a reliable measurement parameter for the increase in photosynthetic capacity.

[0082] As used herein, the term “plant vigor” refers to the amount (measured by weight) of tissue produced by the plant in a given period. Therefore, increased vigor can determine or affect plant yield or yield per growing period or growing area. Furthermore, early vigor (seed and / or seedling) results in better field stand. Improving early vigor is an important goal of modern rice breeding programs, both in temperate and tropical rice cultivars. Long roots are important for proper soil anchorage in water-sown rice. Where rice is sown directly in flooded fields and where plants need to emerge quickly through the water, longer shoots are associated with vigor. Where direct seeding is practiced, mesocotyls Petition 870250109327, dated 11 / 28 / 2025, page 25 / 107 22 / 78 and longer coleoptiles are important for good seedling emergence. The ability to modify initial vigor in plants would be of great importance in agriculture. For example, low initial vigor has been a limitation to the introduction of maize (Zea mays L.) hybrids based on germplasm from the Corn Belt in the European Atlantic.

[0083] In some embodiments, a plant characteristic, such as yield, growth rate, biomass, vigor, oil content, fiber yield, fiber quality, fiber length, photosynthetic capacity and fertilizer use efficiency (e.g., nitrogen use efficiency), can be determined under stress (e.g., abiotic stress, nitrogen-limiting conditions) and / or stress-free (normal) conditions.

[0084] As used herein, the expression “no-stress conditions” refers to growing conditions (e.g., water, temperature, light-dark cycles, humidity, salt concentration, fertilizer concentration in the soil, supply of nutrients such as nitrogen, phosphorus, and / or potassium) that do not significantly exceed the everyday climatic and other abiotic conditions that plants may encounter, and that allow for the optimal growth, metabolism, reproduction, and / or viability of a plant at any stage of its life cycle (e.g., in a cultivated plant, from seed to mature plant and back to seed). People skilled in the art are aware of the normal soil and climatic conditions for a given plant in a given geographical location.It should be noted that although stress-free conditions may include some slight variations from ideal conditions (which vary from one plant type / species to another), such variations do not cause the plant to stop growing without the ability to resume growth.

[0085] The term “fiber” generally includes thick-walled conducting cells such as vessels and tracheids, and fibrillar aggregates of many individual fibrous cells. Therefore, the term “fiber” includes lignin and also refers to (a) conducting and non-conducting cells of. Petition 870250109327, dated 11 / 28 / 2025, page 26 / 107 23 / 78 thick xylem walls; (b) fibers of extra-xylar origin, including those of the phloem, bark, soil tissue, and epidermis; and (c) fibers from stems, leaves, roots, seeds, and flowers or inflorescences (such as those of Sorghum vulgare, used in the manufacture of brushes and brooms). Examples of fiber-producing plants include, among others, agricultural crops such as cotton, silk cotton (Ceiba pentandra), desert willow, creosote bush, fern, balsa, kenaf, roselle, jute, sisal, abaca, flax, corn, sugarcane, hemp, ramie, kapok, coconut fiber, bamboo, Spanish moss, and Agave spp. (e.g., sisal).

[0086] As used herein, the term “fiber quality” refers to at least one fiber parameter that is agriculturally desired or required in the fiber industry (described further below).Examples of such parameters include, among others, fiber length, fiber strength, fiber suitability, fiber weight per unit length, maturity index, and uniformity (described further below). In some applications, cotton fiber quality is typically measured according to fiber length, strength, and fineness. Consequently, fiber quality is considered higher when the fiber is longer, stronger, and finer.

[0087] As used herein, the term “fiber yield” refers to the amount of fiber produced from the fiber-producing plant.

[0088] As disclosed herein, the transgenic plants of the present invention can be used to improve a myriad of commercially desirable traits, all interrelated, as discussed below.

[0089] As used herein, the term “characteristic” refers to a feature or quality of a plant that can generally (directly or indirectly) enhance the commercial value of the plant.

[0090] As used herein, the term “increase” refers to at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 10%, at least about Petition 870250109327, dated 11 / 28 / 2025, p. 27 / 107 24 / 78 15%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, increase in the characteristic [e.g., yield, seed yield, biomass, growth rate, vigor, oil content, fiber yield, fiber quality, fiber length, photosynthetic capacity, tolerance to abiotic stress and / or nitrogen use efficiency] of a plant compared with a native plant or a wild plant [i.e., a plant not modified with the biomolecules (polynucleotide or polypeptides) of the invention, for example, an untransformed plant of the same species that is grown under the same (e.g., identical growing conditions)] or any intermediate value.

[0091] The phrase “expressing an exogenous polynucleotide in the plant”, as used herein, refers to the upregulation of the expression level of an exogenous polynucleotide in the plant by introducing the exogenous polynucleotide into a plant cell or plant and expressing it by recombinant means, as described below.

[0092] As used herein, “express” refers to expression at the mRNA level and, optionally, at the polypeptide level.

[0093] As used herein, the phrase “exogenous polynucleotide” refers to a heterologous nucleic acid sequence that may not be naturally expressed in the plant (e.g., a nucleic acid sequence from a different species) or whose overexpression in the plant is desired. The exogenous polynucleotide may be introduced into the plant stably or transiently so as to produce a ribonucleic acid (RNA) molecule and / or a polypeptide molecule. It should be noted that the exogenous polynucleotide may comprise a nucleic acid sequence that is identical or partially homologous to an endogenous nucleic acid sequence of the plant.

[0094] As used in this document, the term “endogenous” refers to any polynucleotide or polypeptide present and / or Petition 870250109327, dated 11 / 28 / 2025, p. 28 / 107 25 / 78 naturally expressed in a plant or cell thereof.

[0095] As used in this document, homologous sequences include orthologous and paralogous sequences. The term “paralog” refers to gene duplications within the genome of a species, resulting in paralogous genes. The term “orthologous” refers to homologous genes in different organisms due to ancestral relationship. Thus, orthologs are evolutionary counterparts derived from a single ancestral gene in the last common ancestor of two given species (Koonin EV and Galperin MY (Sequence-Evolution-Function: Computational Approaches in Comparative Genomics. Boston: Kluwer Academic; 2003. Chapter 2, Evolutionary Concept in Genetics and Genomics. Available at: ncbi(dot)nlm(dot)nih(dot)gov / books / NBK20255) and therefore have a high probability of having the same function. Homology (e.g., percent homology, sequence identity + sequence similarity) can be determined using any homology comparison software that calculates a pairwise sequence alignment.

[0096] As used herein, “sequence identity” or “identity” in the context of two nucleic acid or polypeptide sequences includes reference to residues in the two sequences that are the same when aligned. When percentage sequence identity is used in reference to proteins, it is recognized that residue positions that are not identical often differ by conservative amino acid substitutions, where amino acid residues are replaced by other amino acid residues with similar chemical properties (e.g., charge or hydrophobicity) and therefore do not alter the functional properties of the molecule. When sequences differ by conservative substitutions, the percentage sequence identity may be adjusted upward to correct for the conservative nature of the substitution. Sequences that differ by such conservative substitutions are considered to have “sequence similarity” or “similarity”.The means to do it. Petition 870250109327, dated 11 / 28 / 2025, p. 29 / 107 26 / 78 This adjustment is well known to those knowledgeable in the art. Typically, this involves classifying a conservative substitution as a partial mismatch rather than a total mismatch, thus increasing the percentage of sequence identity. Thus, for example, when an identical amino acid receives a score of 1 and a non-conservative substitution receives a score of zero, a conservative substitution receives a score between zero and 1. The score for conservative substitutions is calculated, for example, according to the algorithm of Henikoff S and Henikoff J G. [Amino acid substitution matrices of protein blocks. Proc. Natl. Acad. Sci. USA 1992, 89(22): 10915-9].Identity (e.g., percentage of homology) can be determined using any homology comparison software, including, for example, the BlastN software from the National Center for Biotechnology Information (NCBI), using standard parameters. According to some embodiments of the invention, identity is a global identity, i.e., an identity over all amino acid or nucleic acid sequences of the invention and not over parts thereof.

[0097] According to some embodiments of the invention, the term “homology” or “homologous” refers to the identity of two or more nucleic acid sequences; or the identity of two or more amino acid sequences; or the identity of an amino acid sequence with one or more nucleic acid sequences. According to some embodiments of the invention, the homology is a global homology, that is, a homology over all amino acid or nucleic acid sequences of the invention and not over parts thereof. The degree of homology or identity between two or more sequences can be determined using various known sequence comparison tools. FTO Applicants Petition 870250109327, dated 11 / 28 / 2025, p. 30 / 107 27 / 78

[0098] In some modalities, FTO refers to any gene FTO selected from a species comprising Homo sapiens, Acanthaster planci, Patiria miniata, Asterias rubens, Saccoglossus kowalevskii, Patiria miniata, Lytechinus variegatus, Aphanomyces invadans, Aphanomyces astaci, Aphanomyces stellatus, Ostreococcus tauri, Micromonas commoda, Aphanomyces cochlioides, Aphanomyces euteiches, Aphanomyces invadans, Ostreococcus lucimarinus CCE9901, Micromonas pusilla CCMP1545, Ostreococcus tauri, Chaetoceros tenuissimus, Pseudo-nitzschia multistriata, Fragilariopsis cylindrus CCMP1102, Thalassiosira pseudonana CCMP1335, Lytechinus variegatus, Nitzschia inconspicua, Guillardia theta CCMP2712, Ectocarpus sp CCAP 1310 / 34, Ectocarpus siliculosus, Thecamonas trahens ATCC 50062, Polarella glacialis, Aphanomyces invadans, Tribonema minus, Seminavis robusta, Aphanomyces cochlioides, Fragilaria crotonensis, Mayamaea pseudoterrestris, Capsaspora owczarzaki ATCC 30864, ​​Cryptophyta sp CCMP2293, Fragilaria crotonensis, Chaetoceros tenuissimus,synthetic construct, Ectocarpus sp CCAP 1310 / 34, Aphanomyces astaci, Pseudo-nitzschia multistriata, Ochromonadaceae sp CCMP2298, Ectocarpus siliculosus, Seminavis robusta, Nitzschia inconspicua, S phaeroforma arctica JP610, Nitzschia inconspicua, Fragilariopsis cylindrus CCMP1102, Fragilaria crotonensis, Thalassiosira oceanica, Polarella glacialis, Lytechinus pictus, Corticium candelabrum, Cylindrotheca closterium, Skeletonema marinoi, Tetraparma gracilis, Triparma retinervis, Triparma columacea, Triparma laevis f. inornata, Cylindrotheca closterium, Triparma laevis f. longispina, Parmales sp. scaly parma, Triparma columacea, Triparma verrucosa, Triparma strigata, Triparma strigata, Chrysophaeum taylorii, Triparma retinervis, Triparma strigata, Parmales sp. scaly parma, Parmales sp. scaly parma, Alaria esculenta, Alaria esculenta, Cladosiphon okamuranus S strain, Cladosiphon okamuranus S strain, Cyclotella cryptica CCMP332, Macrocystis pyrifera, Macrocystis pyrifera,Minidiscus variabilis CCMP495, Monodopsis C141, Monodopsis C73, Nemacystus, Petition 870250109327, dated 11 / 28 / 2025, page 31 / 107 28 / 78 decipiens Onna-1, Nemacystus decipiens Onna-1, Nitzschia putrida NIES4239, Nitzschia putrida NIES-4239, Ochromonas CCMP139, Paraphysomonas imperforata CCMP1604, Phaeocystis antarctica CCMP1374, Prasinoderma coloniale CCMP1413, Proteomonas sulcata CCMP 1175, Saccharina japonica, Tetraselmis striata, Undaria pinnatifida M23, Undaria pinnatifida M23.

[0099] In some forms, the FTO gene is derived from vertebrates, invertebrates, algae, and / or oomycetes.

[0100] In some embodiments, the FTO gene is identified by the selected accession number of the group comprising “NP_001073901.1, AAI48443.1, XP_022097537.1, XP_038078604.1, XP_033625687.1, XP_002738337.1, ALR88588.1, XP_038078612.1, XP_038049512.1, XP_041454524.1, XP_041454523.1, XP_008874274.1, RHZ09857.1, RHY54141.1, RHY97650.1, KAF0719040.1, RHY17698.1, RHY56488.1, XP_022840077.1, RQM30574.1, XP_009841386.1, XP_002502764.1, KAG9401836.1, KAF0726784.1, RHY29166.1, XP_001420808.1, XP_003060875.1, OUS43030.1, KAH9102871.1, KAH9187704.1, GFH54458.1, VEU39385.1, OEU13050.1, XP_002287957.1, XP_041454525.1, KAG7347203.1, XP_009841387.1, XP_005838228.1, CAB1105374.1, KAG7364155.1, CBJ31658.1, XP_013753876.1, CAE8644684.1, CAE8611733.1, CAE8644257.1, RHZ18017.1, RHY92689.1, RHY56741.1, RHY00398.1, XP_008874276.1, KAG5193084.1, CAB9501471.1, KAG9401837.1, KAI2507824.1, GKY90516.1, XP_004347827.1, KAJ1495429.1, KAI2498707.1, GFH54956.1, AIC63558.1, CAB1117017.1, KAF0752546.1, VEU37468.1, KAJ1427805.1, RLO12103.1, CBJ31645.1, CAB9497616.1, RLO00695.1, KAG7357268.1, XP_014148728.1, KAG7360647.1, OEU10178.1, KAI2500024.1, RHY56738.1, RHY00397.1, RHY92688.1, EJK46618.1, CAE8633430.1, XP_054763800.1, XP_054769608.1, XP_062505117.1, XP_062505118.1, CAJ1967500.1, KAK1734719.1, GMI22625.1, GMH73802.1, GMI46646.1, MDA9098936.1, Petição 870250109327, de 28 / 11 / 2025, pág. 32 / 107 29 / 78 GMH59919.1, CAJ1942268.1, GMH75049.1, GMI53731.1, GMI45036.1, GMH96566.1, GMH91706.1, GMH91703.1, KAJ8609712.1, GMI10743.1, GMH93314.1, GMI61299.1, esubft_2221_2, esubft_644_2, g17329.t1, g9595.t1, CCRYP_016297-RB, estExt_fgenesh2_pg.C_8_t10199, fgenesh2_pm.17_#_163, estExt_Genewise1.C_190154, Monodopsis_C141_contig_8_g54760.t1, Monodopsis_C73_contig_18_g112030.t1, g1621.t1, g11573.t1, NAA08P02010.m1, NAA05P01980.m1, CE122323_2229, fgenesh1_kg.94_#_167_#_TRINITY_DN11056_c0_g1_i3, Phant.0041s0181.1, PRCOL_00005642-RA, fgenesh1_kg.8_#_1316_#_TRINITY_DN4680_c0_g1_i6, SJ15771.1, SJ04115.1, TSEL_005271.t1, TSEL_005270.t1, TSEL_010033.t1, HiC_scaffold_21.g09429.m1, HiC_scaffold_11.g04962.m1.

[0101] In some embodiments, the FTO gene comprises an exogenous polynucleotide or nucleic acid sequence comprising SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37 or SEQ ID NO: 38.In some embodiments, the exogenous polynucleotide or nucleic acid sequence comprises at least about 80%, at least about 81%, at least about 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, for example, 100% identical to the nucleic acid sequence selected from the group consisting of SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37 or SEQ ID NO: 38.

[0102] According to some embodiments of the invention, the exogenous polynucleotide encodes a polypeptide consisting of the amino acid sequence presented by any of the SEQ IDs. Petition 870250109327, dated 11 / 28 / 2025, p. 33 / 107 30 / 78 Nos: 6-30 or 39-58. According to some embodiments of the invention, the exogenous polynucleotide of the disclosure comprises a nucleic acid sequence encoding a polypeptide with an amino acid sequence that is at least about 80%, at least about 81%, at least about 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99% or more, say, 100% homologous (e.g., identical) to the amino acid sequence selected from the group consisting of the SEQ IDs. Nos: 6-30 or 39-58.

[0103] In some embodiments, the exogenous polynucleotide or nucleic acid sequence further comprises at least one nuclear localization signal (NLS) sequence. In some embodiments, the exogenous polynucleotide or nucleic acid sequence comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 NLS sequences or intervals between them. In some embodiments, the exogenous polynucleotide or nucleic acid sequence comprises one NLS sequence. In some embodiments, the exogenous polynucleotide or nucleic acid sequence comprises two NLS sequences. In some embodiments, the exogenous polynucleotide or nucleic acid sequence comprises three NLS sequences.

[0104] In some embodiments, the NLS sequence is approximately or at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, or 100 nucleotides long, or varies between these two extremes. In some embodiments, the NLS sequence is approximately 10 nucleotides long. In some embodiments, the NLS sequence is approximately 11 nucleotides long. In some embodiments, the NLS sequence is approximately 15 nucleotides long. In some embodiments, the sequence Petition 870250109327, dated 11 / 28 / 2025, p. 34 / 107 31 / 78 NLS is approximately 16 nucleotides long. In some embodiments, the NLS sequence is approximately 21 nucleotides long. In some embodiments, the strength of the NLS is weak. In some embodiments, the strength of the NLS is strong. In some embodiments, the strength of the NLS is weak, strong, or a combination thereof.

[0105] Table 1 below illustrates the amino acid locations of the NLS or predicted NLS and their relative strength, when present. Table 1 SEQ ID Nr. Name Predicted NLS Strength 1 hFTO 8-18 Weak 2 mhFTO 8-18 Weak 3 olFTO 338-352 Very weak 4 ssFTO 8-18 Weak 5 btFTO 10-18 Weak 6 apFTO 18-33 Strong 7 mcFTO 3-45 Strong 8 niFTO 8-20, 537-551 Strong 9 acFTO 4-60, 495-512 Strong 10 ctFTO 2-38, 523-557 Strong 11 crtFTO 2-19 Strong 12 cmFTO None None 13 mecFTO 3-17 Strong 14 anaFTO 2-17 Strong 15 pmFTO 4-28 Strong 16 arFTO 51-67 Strong 17 skFTO 3-13 Weak 18 otFTO None None 19 mpFTO 10-40, 527-541 Strong 20 esFTO-1 40-53, 85-95, 383-398, Strong Petition 870250109327, dated 11 / 28 / 2025, page 35 / 107 32 / 78 652-715 21 esFTO-2 5-31, 187-209, 486-502 Forte 22 tpFTO 3-31, 581-613 Forte 23 pgFTO 661-674 Forte 24 pnmFTO 4-18, 316-326, 514-549 Forte 25 tmFTO 78-84, 538-551, 588-610 Forte 26 aeFTO 4-56, 491-512 Forte 27 aaFTO 3-35, 505-518 Forte 28 aiFTO 4-25, 522-535 Forte 29 asFTO 4-70, 501-516 Forte

[0106] In the composition and methods of the present invention, the exogenous polynucleotide or nucleic acid sequence disclosed herein is cloned into transformation vectors. The vectors used for plant transformation may include any vector generally known in the art.

[0107] In some embodiments, the nucleic acid construct, according to some embodiments of the invention, comprises a promoter sequence and the isolated polynucleotide of some embodiments of the invention.

[0108] According to some embodiments of the invention, the isolated polynucleotide is operatively linked to the promoter sequence. A coding nucleic acid sequence is “operatively linked” to a regulatory sequence (e.g., promoter) if the regulatory sequence is capable of exerting a regulatory effect on the coding sequence linked to it. As used herein, the term “promoter” refers to a region of DNA that lies upstream of the transcriptional initiation site of a gene to which RNA polymerase binds to initiate RNA transcription. The promoter controls where (e.g., which part of a plant) and / or when (e.g., at what stage or condition of an organism’s life) the gene is expressed.

[0109] According to some embodiments of the invention, the Petition 870250109327, dated 11 / 28 / 2025, p. 36 / 107 33 / 78 The promoter is heterologous to the isolated polynucleotide and / or the host cell. As used herein, the term “heterologous promoter” refers to a promoter from a different species or from the same species but from a different gene locus than the isolated polynucleotide sequence. According to some embodiments of the invention, the isolated polynucleotide is heterologous to the plant cell (e.g., the polynucleotide is derived from a different plant species compared to the plant cell, therefore the isolated polynucleotide and the plant cell are not of the same plant species).

[0110] Any suitable promoter sequence can be used by the nucleic acid construct of the present invention. Preferably, the promoter is a constitutive promoter, a tissue-specific promoter, or an abiotic stress-inducible promoter. According to some embodiments of the invention, the promoter is a plant promoter, which is suitable for the expression of the exogenous polynucleotide in a plant cell.

[0111] Promoters suitable for expression in wheat include, but are not limited to, wheat SPA promoter (SEQ ID NO: 6590; Albani et al., Plant Cell, 9: 171-184, 1997, which is fully incorporated herein by reference), wheat LMW (SEQ ID NO: 6591 (longer LMW promoter) and SEQ ID NO: 6592 (LMW promoter) and glutenin-1 HMW (SEQ ID NO: 6593 (longer wheat glutenin-1 HMW promoter); and SEQ ID NO: 6594 (wheat glutenin-1 HMW promoter); Thomas and Flavell, Plant Cell 2:1171-1180; Furtado et al., 2009 Plant Biotechnology Journal 7: 240-253, each of which is fully incorporated herein by reference), wheat alpha, beta and gamma gliadins [e.g., SEQ ID NO: 6595 (wheat alpha gliadin, B genome, promoter); SEQ ID NO: 6596 (wheat gamma gliadin promoter); EMBO 3: 1409-15, 1984, which is fully incorporated herein by reference], wheat TdPR60 [SEQ ID NO: 6597 (longer wheat TdPR60 promoter) or SEQ ID NO: 6598 (wheat TdPR60 promoter); Kovalchuk et al., Plant Mol Biol 71:81-98, 2009, which is fully incorporated herein by reference], Ub1 promoter. Petition 870250109327, dated 11 / 28 / 2025, p. 37 / 107 34 / 78 maize [cultivar Nongda 105 (SEQ ID NO: 6599); GenBank: DQ141598. 1; Taylor et al., Plant Cell Rep 1993 12: 491-495, which is fully incorporated herein by reference; and cultivar B73 (SEQ ID NO: 6600); Christensen, AH, et al. 18 (4), 675-689 (1992), which is fully incorporated herein by reference]; rice actin 1 (SEQ ID NO: 6601; Mc Elroy et al. 1990, The Plant Cell, Vol. 2, 163-171, which is fully incorporated herein by reference), rice GOS2 [SEQ ID NO: 6602 (longer promoter of rice GOS2) and SEQ ID NO: 6603 (promoter of rice GOS2); De Pater et al. Planta J. 1992; 2: 837-44, which is fully incorporated herein by reference], Arabidopsis Pho1 [SEQ ID NO: 6604 (promoter Arabidopsis Pho1); Hamburger et al., Plant Cell.2002; 14: 889-902, which is fully incorporated herein by reference], ExpansinB promoters, for example, ExpB5 from rice [SEQ ID NO: 6605 (longer promoter of ExpB5 from rice) and SEQ ID NO: 6606 (promoter of ExpB5 from rice)] and ExpB1 from barley [SEQ ID NO: 6607 (promoter of ExpB1 from barley), Won et al. Mol Cells. 2010; 30: 369-76, which is fully incorporated herein by reference], SS2 from barley (sucrose synthase 2) [(SEQ ID NO: 6608), Guerin and Carbonero, Plant Physiology May 1997 vol. 114 No. 1 55-62, which is fully incorporated herein by reference], and PG5a rice [SEQ ID NO: 6609, U.S. Patent No. 7,700,835, Nakase et al., Plant Mol Biol. 32:621-30, 1996, each of which is fully incorporated herein by reference].

[0112] Suitable constitutive promoters include, for example, CaMV 35S promoter [SEQ ID NO: 6610 (CaMV 35S Promoter (pQXNc)); SEQ ID NO: 6611 (Brachypodium PJJ 35S); SEQ ID NO: 6612 (Promoter CaMV 35S (OLD)) (Odell et al., Nature 313:810-812, 1985)], promoter Arabidopsis At6669 (SEQ ID NO: 6613 (Promoter Arabidopsis At6669 (OLD)); see PCT Publication No. WO04081173A2 or the new promoter At6669 (SEQ ID NO: 6614 (Promoter Arabidopsis At6669 (NEW))); maize Ub1 promoter [cultivar Nongda 105 (SEQ ID NO: 6599); GenBank: DQ141598.1; Taylor et al., Plant Cell Rep 1993 12: 491-495, which is fully incorporated herein by reference; and cultivar B73 (SEQ ID Petition 870250109327, dated 11 / 28 / 2025, page 38 / 107 35 / 78 NO: 6600); Christensen, A.H., et al. Plant Mol. Biol. 18 (4), 675-689 (1992), which is fully incorporated here by reference]; actin 1 de arroz (SEQ ID NO: 6601, McElroy et al., Plant Cell 2:163-171, 1990); pEMU (Last et al., Theor. Appl. Genet. 81:581-588, 1991); CaMV 19S (Nilsson et al., Physiol. Plant 100:456-462, 1997); GOS2 de arroz [SEQ ID NO: 6602 (promotor mais longo de GOS2 de arroz) e SEQ ID NO: 6603 (promotor de GOS2 de arroz), de Pater et al., Plant J November; 2(6):837-44, 1992]; Promotor RBCS (SEQ ID NO: 6615); Cyclophilin de arroz (Bucholz et al, Plant Mol Biol. 25(5):837-43, 1994); Histona H3 de milho (Lepetit et al, Mol. Gen. Genet. 231: 276-285, 1992); Actin 2 (An et al, Plant J. 10(1); 107-121, 1996) and Super MAS sintético (Ni et al., The Plant Journal 7: 66176, 1995). Outros constitutivos promoters incluem aqueles nas patentes dos EUA n°s 5.659.026, 5.608.149; 5,608,144; 5,604,121; 5,569,597: 5,466,785; 5,399,680; 5,268,463; and 5,608,142.

[0113] Suitable tissue-specific promoters include, but are not limited to, leaf-specific promoters [e.g., AT5G06690 (Thioredoxin) (high expression, SEQ ID NO: 6616), AT5G61520 (AtSTP3) (low expression, SEQ ID NO: 6617) described in Buttner et al. 2000 Plant, Cell and Environment 23, 175-184, or the promoters described in Yamamoto et al., Plant J. 12:255-265, 1997; Kwon et al., Plant Physiol. 105:357-67, 1994; Yamamoto et al., Plant Cell Physiol. 35:773-778, 1994; Gotor et al., Plant J. 3:509-18, 1993; Orozco et al., Plant Mol. Biol. 23:1129-1138, 1993; and Matsuoka et al., Proc. EUA 90:9586-9590, 1993; as well as the Arabidopsis STP3 promoter (AT5G61520) (Buttner et al., Plant, Cell and Environment 23: 175-184, 2000)], seed-preferred promoters [e.g., Napin (derived from Brassica napus which is characterized by a seed-specific promoting activity; Stuitje AR et al. Plant Biotechnology Journal 1 (4): 301-309; SEQ ID NO: 6618 (NAPIN promoter from Brassica napus) of seed-specific genes (Simon, et al., Plant Mol. Biol. 5. 191, 1985; Scofield, et al., J. Biol. Chem. 262: 12202, 1987; Baszczynski, et al., Plant Mol. Biol. 14: 633, 1990), rice PG5a (SEQ ID NO: 6609; US Patent No. Petition 870250109327, dated 11 / 28 / 2025, p. 39 / 107 36 / 78 7.700.835), Arabidopsis early seed development BAN (AT1G61720) (SEQ ID NO: 6619, US 2009 / 0031450 A1), Arabidopsis seed late development ABI3 (AT3G24650) (SEQ 62 Promotor ID NO: May Arabidopsis ABI3 (AT3G24650)) or 6621 (Promotor of Arabidopsis ABI3 (AT3G24650))) (Ng et al., Plant Molecular Biology 54: 25-38, 2004), albumin de castanha-do-pará (Pearson' et al., Plant Biol. 18: Mol. 235-245, 1992), legumina (Ellis, et al. Plant Mol. Biol. 10: 203-214, 1988), Glutelina (rice) (Takaiwa, et al., Mol. Gen. Genet. 208: 15-22, 1986; Takaiwa, et al., FEBS, 21:21). 43-47, 1987), Zein (Matzke et al. Plant Mol Biol, 143). 323-32 1990), napA (Stalberg, et al., Planta 199: 515-519, 1996), Wheat SPA (SEQ ID NO: 6590; Albanietal, Plant Cell, 9: 171-184, 1997), oleosin de gyrassol (Plant et al., Mol. Biol.19: 873-876, 1992)], specific endosperm promoters [e.g., wheat LMW (SEQ ID NO: 6591 (Wheat LMW longer promoter) and SEQ ID NO: 6592 (Wheat LMW promoter) and glutenin-1 HMW [(SEQ ID NO: 6593 (Wheat HMW glutenin-1 longer promoter)); and SEQ ID NO: 6594 (Wheat HMW glutenin-1 promoter), Thomas and Flavell, The Plant Cell 2:11711180, 1990; Mol Gen Genet 216:81-90, 1989; NAR17:461-2), wheat alpha, beta, and gamma gliadins (SEQ ID NO: 6595 (wheat alpha gliadin promoter (B genome)); SEQ ID NO: 6596 (wheat gamma gliadin promoter); EMBO 3: 1409-15, 1984), barley ltrl promoter, barley B1, C, D hordein (Theor Appl Gen 98: 1253-62, 1999; Plant J 4: 343-55, 1993; Mol Gen Genet 250:750-60, 1996), barley DOF (Mena et al. The Plant Journal, 116(1): 53-62, 1998), Biz2 (EP99106056.7), Barley SS2 (SEQ ID NO: 6608 (Barley SS2 Promoter); Guerin and Carbonero Plant Physiology 114: 1 55-62, 1997), Wheat Tarp60 (Kovalchuk et al., Plant Mol Biol 71:81-98, 2009), barley D-hordein (D-Hor) and B-hordein (BHor) (Agnelo Furtado, Robert J. Henry and Alessandro Pellegrineschi (2009)], synthetic promoter (Vicente-Carbajosa et al., Plant J. 13: 629640, 1998), rice prolamin NRP33, rice globulin Glb-1 (Wu et al., Plant Cell Physiology 39(8) 885-889, 1998), rice alpha-globulin. Petition 870250109327, dated 11 / 28 / 2025, p. 40 / 107 37 / 78 REB / OHP-1 (Nakase et al. Plant Mol. Biol. 33: 513-S22, 1997), rice ADPglucose PP (Trans Res 6:157-68, 1997), maize ESR gene family (Plant J 12:235-46, 1997), sorghum gamma-caphyrin (PMB 32:102935, 1996)], specific embryo promoters [e.g., rice OSH1 (Sato et al., Proc. Natl. Acad. Sci. USA, 93: 8117-8122), KNOX (PostmaHaarsma et al., Plant Mol. Biol. 39:257-71, 1999), rice oleosin (Wu et al., J. Biochem., 123:386, 1998)] and flower-specific promoters [e.g., AtPRP4, chalene synthase (chsA) (Van der Meer, et al., Plant Mol. Biol. 15, 95-109, 1990), LAT52 (Twell et al. Mol. Gen Genet. 217:240-245; 1989), Arabidopsis apetala—3 (Tilly et al., Development. 125:1647-57, 1998), Arabidopsis APETALA 1 (AT1G69120, API) (SEQ ID NO: 6622 (Arabidopsis (AT1G69120) APETALA 1)) (Hempel et al., Development 124:3845-3853, 1997)] and root promoters [e.g., the ROOTP promoter [SEQ ID NO: 6623]; rice ExpB5 (SEQ ID NO: 6606 (Rice ExpB5 promoter); or SEQ ID NO: 6605 (Longer rice ExpB5 promoter)) and barley ExpB1 promoters (SEQ ID NO: 6607) (Won et al. Mol. Cells 30: 369-376, 2010); Arabidopsis ATTPS-CIN (AT3G25820) promoter (SEQ ID NO: 6624; Chen et al., Plant Phys 135:1956-66, 2004); Arabidopsis Pho1 promoter (SEQ ID NO: 6604, Hamburger et al., Plant Cell. 14: 889-902, 2002), which is also slightly induced by stress].

[0114] Suitable abiotic stress-inducible promoters include, but are not limited to, salt-inducible promoters such as RD29A (Yamaguchi-Shinozalei et al., Mol. Gen. Genet. 236:331-340, 1993); drought-inducible promoters, such as the maize rab17 gene promoter (Pla et al., Plant Mol. Biol. 21:259-266, 1993), the maize rab28 gene promoter (Busk et al., Plant J. 11:1285-1295, 1997) and the maize Ivr2 gene promoter (Pelleschi et al., Plant Mol. Biol. 39:373-380, 1999); Heat-inducible promoters, such as the hsp80 tomato heat promoter (US Patent No. 5,187,267).

[0115] According to some embodiments of the invention, the expression of the exogenous polynucleotide of the invention in the plant is effected Petition 870250109327, dated 11 / 28 / 2025, p. 41 / 107 38 / 78 by transforming one or more plant cells with the exogenous polynucleotide, followed by generating a mature plant from the transformed cells and cultivating the mature plant under conditions suitable for the expression of the exogenous polynucleotide in the mature plant. According to some embodiments of the invention, the transformation is effected by introducing into the plant cell one or more nucleic acid constructs that include the exogenous polynucleotide of some embodiments of the invention and at least one promoter to direct the transcription of the exogenous polynucleotide in a host cell (e.g., a plant cell).

[0116] The nucleic acid construct of some embodiments of the invention may further include an appropriate selectable marker and / or an origin of replication. According to some embodiments of the invention, the nucleic acid construct used is a transport vector, which can propagate both in E. coli (where the construct comprises an appropriate selectable marker and an origin of replication) and be compatible with propagation in cells. The construct according to the present invention may be, for example, a plasmid, a bacmid, a phagemid, a cosmid, a phage, a virus, or an artificial chromosome.

[0117] The nucleic acid construct of some embodiments of the invention may further include appropriate localization signals, such as nuclear localization signals (NLS). According to some embodiments of the invention, the nucleic acid construct used has a nuclear localization signal as described in Raikhel, Plant Physiol. 100: 1627-1632 (1992) and references mentioned therein, such as the NLS of the SV40 long T antigen [Kalderon et al. Cell 39: 499-509 (1984)]. The nuclear localization signal can be located anywhere in the protein, but is conveniently located at the N-terminal end of the protein. As illustrated in Table 1, there are predicted sites on FTO homologs for the NLS>

[0118] The construction of nucleic acid in some forms of Petition 870250109327, dated 11 / 28 / 2025, page 42 / 107 39 / 78 The invention can be used to transform plant cells in a stable or transient manner. In stable transformation, the exogenous polynucleotide is integrated into the plant genome and, as such, represents a stable and inherited trait. In transient transformation, the exogenous polynucleotide is expressed by the transformed cell but is not integrated into the genome and, as such, represents a transient trait.

[0119] There are several methods of introducing foreign genes into monocotyledonous and dicotyledonous plants (Potrykus, I., Annu. Rev. Plant. Physiol., Plant. Mol. Biol. (1991) 42:205-225; Shimamoto et al., Nature (1989) 338:274-276).

[0120] The main methods for causing stable integration of exogenous DNA into plant genomic DNA include two main approaches:

[0121] (i) Gene transfer mediated by Agrobacterium: Klee et al. Klee and Rogers in Cell Culture and Somatic Cell Genetics of Plants, Vol. 6, Molecular Biology of Plant Nuclear Genes, eds. Schell, J. and Vasil, LK, Academic Publishers, San Diego, California (1989), p. 2-25; Gatenby, in Plant Biotechnology, eds. Kung, S. and Arntzen, CJ, Butterworth Publishers, Boston, Massachusetts (1989), p. 93-112.

[0122] (ii) Direct DNA uptake: Paszkowski et al., in Cell Culture and Somatic Cell Genetics of Plants, Vol. 6, Molecular Biology of Plant Nuclear Genes eds. Schell, J., and Vasil, LK, Academic Publishers, San Diego, Calif. (1989) p. 52-68; including methods for direct DNA uptake in protoplasts, Toriyama, K. et al. (1988) Bio / Technology 6:1072-1074. DNA uptake induced by brief electric shock in plant cells: Zhang et al. (1988) 7:379-384. Fromm et al. Nature (1986) 319:791-793. Injection of DNA into plant cells or tissues by particle bombardment, Klein et al. Bio / Technology (1988) 6:559-563; McCabe et al. Bio / Technology (1988) 6:923-926; Sanford, Physiol. Plant. (1990) 79:206-209; by the use of micropipette systems: Neuhaus et al., Theor. Appl. Genet. (1987) 75:30-36; Neuhaus and Spangenberg, Physiol. Petition 870250109327, dated 11 / 28 / 2025, page 43 / 107 40 / 78 Plant. (1990) 79:213-217; transformation of glass fibers or silicon carbide fibers into cell cultures, embryos or callus tissue, U.S. Patent No. 5,464,765 or by direct incubation of DNA with germinating pollen, DeWet et al. in Experimental Manipulation of Ovule Tissue, eds. Chapman, GP and Mantell, SH and Daniels, W. Longman, London, (1985) p. 197-209; and Ohta, Proc. Natl. Acad. Sci. USA (1986) 83:715-719.

[0123] The Agrobacterium system involves the use of plasmid vectors containing defined DNA segments that integrate into the plant's genomic DNA. Plant tissue inoculation methods vary depending on the plant species and the Agrobacterium delivery system. A widely used approach is the leaf disc procedure, which can be performed with any tissue explant that provides a good source for initiating whole-plant differentiation. See, for example, Horsch et al. in Plant Molecular Biology Manual AS, Kluwer Academic Publishers, Dordrecht (1988) p. 1-9. A complementary approach employs the Agrobacterium delivery system in combination with vacuum infiltration. The Agrobacterium system is especially viable in the creation of transgenic dicotyledonous plants.

[0124] There are several methods of direct DNA transfer into plant cells. In electroporation, protoplasts are briefly exposed to a strong electric field. In microinjection, DNA is mechanically injected directly into cells using very small micropipettes. In microparticle bombardment, DNA is adsorbed onto microprojectiles, such as magnesium sulfate crystals or tungsten particles, and the microprojectiles are physically accelerated into plant cells or tissues.

[0125] According to some embodiments of the invention, transgenic plants are generated by transient transformation of leaf cells, meristematic cells or the whole plant. The transient transformation can be affected by any of the methods of Petition 870250109327, dated 11 / 28 / 2025, page 44 / 107 41 / 78 direct DNA transfer as described above or by viral infection using modified plant viruses.

[0126] Viruses that have proven useful for transforming host plants include, among others, CaMV, tobacco mosaic virus (TMV), bromine mosaic virus (BMV), and bean common mosaic virus (BV or BCMV). Plant transformation using plant viruses is described in U.S. Patent No. 4,855,237 (bean golden mosaic virus; BGV), EP-A 67,553 (TMV), Japanese Publication Application No. 63-14693 (TMV), EPA 194,809 (BV), EPA 278,667 (BV); and Gluzman, Y. et al., Communications in Molecular Biology: Viral Vectors, Cold Spring Harbor Laboratory, New York, pp. 172-189 (1988). Pseudovirus particles for use in expressing foreign DNA in many hosts, including plants, are described in document WO 87 / 06261.

[0127] According to some embodiments of the invention, the virus used for transient transformations is avirulent and therefore incapable of causing severe symptoms such as reduced growth rate, mosaic, ring spots, leaf curling, yellowing, streaking, pox formation, tumor formation, and corrosion. A suitable avirulent virus may be a naturally occurring avirulent virus or an artificially attenuated virus. Viral attenuation may be achieved by the use of methods well known in the art, including, but not limited to, sublethal heating, chemical treatment, or by targeted mutagenesis techniques, such as those described, for example, by Kurihara and Watanabe (Molecular Plant Pathology 4:259-269, 2003), Gal-on et al. (1992), Atreya et al. (1992), and Huet et al. (1994).

[0128] Suitable viral strains can be obtained from available sources, such as the American Type Culture Collection (ATCC), or by isolation from infected plants. Isolation of viruses from infected plant tissues can be performed using well-known techniques in the field, such as those described, for example, by Foster and Taylor, Eds. “Plant Virology Protocols: From Virus Isolation to Transgenic Petition 870250109327, dated 11 / 28 / 2025, page 45 / 107 42 / 78 Resistance (Methods in Molecular Biology (Humana Pr), Vol. 81)”, Humana Press, 1998. In short, tissues from an infected plant believed to contain a high concentration of a suitable virus, preferably young leaves and flower petals, are ground in a buffer solution (e.g., phosphate buffer solution) to produce a virus-infected sap that can be used in subsequent inoculations.

[0129] The construction of plant RNA viruses for the introduction and expression of exogenous nonviral polynucleotide sequences in plants is demonstrated by the references above, as well as by Dawson, WO et al., Virology (1989) 172:285-292; Takamatsu et al. EMBO J. (1987) 6:307-311; French et al. Science (1986) 231:1294-1297; Takamatsu et al. FEBS Letters (1990) 269:73-76; and U.S. Patent No. 5,316,931.

[0130] When the virus is a DNA virus, appropriate modifications can be made to the virus itself. Alternatively, the virus can first be cloned into a bacterial plasmid to facilitate the construction of the desired viral vector with the foreign DNA. The virus can then be excised from the plasmid. If the virus is a DNA virus, a bacterial origin of replication can be attached to the viral DNA, which is then replicated by the bacteria. Transcription and translation of this DNA will produce the coat protein that will encapsulate the viral DNA. If the virus is an RNA virus, it is usually cloned as a cDNA and inserted into a plasmid. The plasmid is then used to make all the constructs. The RNA virus is then produced by transcribing the viral sequence from the plasmid and translating the viral genes to produce the coat protein(s) that encapsulate the viral RNA.

[0131] According to one aspect of some embodiments of the invention, the method for increasing the yield, biomass, growth rate, vigor, oil content, fiber yield, fiber quality, fiber length, photosynthetic capacity, tolerance to abiotic stress and / or nitrogen use efficiency of a plant. Petition 870250109327, dated 11 / 28 / 2025, page 46 / 107 43 / 78 is effected by the expression, in the plant, of an exogenous polynucleotide comprising a nucleic acid sequence encoding a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 6-30 and 39-58, thereby increasing the yield, biomass, growth rate, vigor, oil content, fiber yield, fiber quality, fiber length, photosynthetic capacity, tolerance to abiotic stress and / or nitrogen use efficiency of the plant.

[0132] According to one aspect of some embodiments of the invention, a method is provided for increasing the yield, biomass, growth rate, vigor, oil content, fiber yield, fiber quality, fiber length, photosynthetic capacity, abiotic stress tolerance and / or nitrogen use efficiency of a plant, comprising the expression within the plant of an exogenous polynucleotide comprising a nucleic acid sequence of at least about 80%, at least about 81%, at least about 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 93%, at least approximately 94%, at least approximately 95%, at least approximately 96%, at least approximately 97%, at least approximately 98%,at least approximately 99%, for example, 100% identical to the selected nucleic acid sequence from the group consisting of SEQ ID NOs: 34-38, thus increasing the yield, biomass, growth rate, vigor, oil content, fiber yield, fiber quality, fiber length, photosynthetic capacity, tolerance to abiotic stress and / or nitrogen use efficiency of the plant.

[0133] In some embodiments, the methods of the invention include all plants belonging to the superfamily Viridiplantae, in particular monocotyledonous and dicotyledonous plants, including a Petition 870250109327, dated 11 / 28 / 2025, p. 47 / 107 44 / 78 leguminosa forrageira or forrageira, ornamental plant, alimentary culture, árvore ou arbusto selected from lista spp., Agathis australis, Albizia amara, Alsophila tricolor, Andropogon spp., Arachis spp., Areca catechu, Astelia fragrans, Astragalus cicer, Baikiaea plurijuga, Betula spp., Brassica spp farinosa, Calliandra spp., Camellia sinensis, Canna indica, Cannabis ssp., Capsicum spp., Cassia spp., Centroema pubescens, Chacoomeles spp., Cinnamomum cassia, Coffea arabica, Colophospermum mopane, Coronillia varia, Cotoneaster serotina, Crataegus spp., Cucumis spp., Cupressus spp., Cyathea whitebata, Cydonia oblonga, Cryptomeria japonica, Cymbopogon spp., Cynthea whitebata, Cydonia oblonga, Dalbergia monataria, Davallia divaricata, Desmodium spp., Dicksonia squarosa, embracing Dibeteropogon, Dioclea spp spp., Dorycnium rectum, Echinochloa pyramidalis, Ehraffia spp., Eleusine coracana, Eragrestis spp., Erythrina spp., Eucalyppus spp., Euclea schimperi, Eulalia villosa, Pagopyrum spp., Feijoa sellowlana, Fragaria spp., Flemingia spp., Freycinetia banksli, Geranium thunbergii, GinAgo biloba, Glycine javanica, Gliricidia spp., Gossypium hirsutum, Gossypium spp., Grevillea spp., Guibourtia coleosperma, Hedysarum spp., Hemaffhia altissima, Heteropogon contoffus, Hordeum vulgare, Hyparrhenia rufa, Hypericum erectum, Hypefhelia dissolute, Indigo incamata, Iris spp., Leptarrhena pyrolifolia, Lespediza spp., Lettuca spp., Leucaena leucocephala, Linum spp., Linum usitatissimum, Loudetia simplex, Lotonus bainesli, Lotus spp., Macrotyloma axillare, Malus spp. Saliva of Medicago, Metasequoia glyptostroboides, Musa sapientum, Nicotianum spp., Onobrychis spp., Ornithopus spp., Oryza spp., Peltophorum africanum, Pennisetum spp., Persea gratissima, Petunia spp., Phaseolus spp., Phoenix canariensis, Phormium cookianum spp., Photinia spp., Picea glauca, Pinus spp., Pisum sativam, Podocarpus totara, Pogonarthria fleckii, Pogonaffhria squarrosa, Populus spp., Prosopis cineraria, Pseudotsuga menziesii, Pterolobium stellatum, Pyrus communis, Quercus spp., Rhaphiolepsis umbellata, Rhopalostylis sapida, Rhus natalensis, Ribes grossularia, Ribes spp., Robinia pseudoacacia, Rosa spp., Rubus spp., Salix spp., Schyzachyrium. Petition 870250109327, dated 28 / 11 / 2025, p. 48 / 107 45 / 78 sanguineum, Sciadopitys vefficillata, Sequoia sempervirens, Sequoiadendron giganteum, Sorghum bicolor, Spinacia spp., Sporobolus fimbriatus, Stiburus alopecuroides, Stylosanthos humilis, Tadehagi spp., Taxodium distichum, Themeda triandra, Trifolium spp., Triticum spp., Tsuga heterophylla, Vaccinium spp., Vicia spp., Vitis vinifera, Watsonia piramidata, Zantedeschia aethiopica, Zea mays, amaranth, artichoke, asparagus, broccoli, Brussels sprouts, cabbage, canola, carrot, cauliflower, celery, kale, flax, cabbage, lentil, rapeseed, okra, onion, potato, rice, soybean, straw, beet, sugarcane, sunflower, tomato, pumpkin tea, corn, wheat, barley, rye, oats, peanut, pea, lentil and alfalfa, cotton, Rapeseed, canola, pepper, sunflower, tobacco, eggplant, eucalyptus, a tree, an ornamental plant, a perennial grass, and a forage crop. In some embodiments, algae and other non-viridifoliate plants may be used for the methods of the present invention.

[0134] According to some embodiments of the invention, the plant used by the method of the invention is a cultivated plant, such as rice, corn, wheat, barley, peanut, potato, sesame, olive, palm oil, banana, soybean, sunflower, canola, sugarcane, alfalfa, millet, legumes (beans, peas), flax, lupin, rapeseed, tobacco, poplar and cotton.

[0135] In some embodiments, the plant used by the method is selected from the group consisting of canola, wheat, soybean, millet, sorghum, cotton, tomato, grape, peanut, rice, lettuce, melon, sweet potato, carrot, cucumber, sugar cane, alfalfa, Camelina or PennyCress.

[0136] In some embodiments, the FTO gene comprising an exogenous polynucleotide or a nucleic acid sequence comprising SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37 or SEQ ID NO: 38 is expressed in a nucleus of said plant, plant cell, tissue or seed.

[0137] In some embodiments, the exogenous polynucleotide encodes an FTO polypeptide comprising the amino acid sequence Petition 870250109327, dated 11 / 28 / 2025, page 49 / 107 46 / 78 established by either of the SEQ ID Nos: 6-30 or 39-71 and is expressed in a nucleus of said plant, plant cell, tissue or seed.

[0138] According to one aspect of some embodiments of the invention, a method is provided for producing a plant with increased yield, growth rate, biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency, drought tolerance and / or abiotic stress tolerance compared with a wild plant of the same species grown under the same growing conditions, the method comprising: (a) transforming the plant with an exogenous polynucleotide encoding a polypeptide comprising an amino acid sequence of at least about 80%, at least about 81%, at least about 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 93%, into at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, for example, 100% homologous (e.g., with sequence similarity or identity) to the selected amino acid sequence from the group that consists of SEQ ID NOs: 6-30 or 39-71, and (b) selecting from the plants in step (a) a plant with the highest yield, growth rate, biomass, vigor, oil content, seed yield,fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency, drought tolerance and / or abiotic stress tolerance (by, Petition 870250109327, dated 11 / 28 / 2025, page 50 / 107 47 / 78 example, selecting plants for the enhanced characteristic).

[0139] According to one aspect of some embodiments of the invention, a method is provided for selecting a transformed plant with higher yield, growth rate, biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency, drought tolerance and / or abiotic stress tolerance compared with a wild plant of the same species grown under the same growing conditions, the method comprising: (a) provide the transformed plants with an exogenous polynucleotide encoding a polypeptide comprising an amino acid sequence that is at least about 80%, at least about 81%, at least about 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, for example, 100% homologous (e.g., with sequence similarity or identity) to the selected amino acid sequence of group consisting of SEQ ID NOs: 6-30 or 39-71, (b) select from among the plants in step (a) a plant with the highest yield, growth rate, biomass, vigor, oil content, seed yield,Fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency, drought tolerance and / or abiotic stress tolerance (e.g., selecting plants for the enhanced trait), thus selecting the plant with the highest yield, growth rate, Petition 870250109327, dated 11 / 28 / 2025, page 51 / 107 48 / 78 biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency and / or tolerance to abiotic stress compared to wild plants of the same species grown under the same growing conditions.

[0140] According to one aspect of some embodiments of the invention, a method is provided for selecting a transformed plant with increased yield, growth rate, biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency and / or tolerance to abiotic stress compared with a wild plant of the same species grown under the same growing conditions, the method comprising: (a) provide the transformed plants with an exogenous polynucleotide that is at least about 80%, at least about 81%, at least about 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, for example, 100% identical to the nucleic acid sequence selected from the group consisting of SEQ ID NOs: 34-38, (b) select from the plants from stage (a) a plant with higher yield, growth rate, biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity,efficiency in nitrogen use and / or tolerance to abiotic stress, Petition 870250109327, dated 11 / 28 / 2025, page 52 / 107 49 / 78 thus selecting the plant with the highest yield, growth rate, biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency and / or tolerance to abiotic stress compared to the wild plant of the same species grown under the same growing conditions.

[0141] According to some embodiments of the invention, the plant is a dicotyledonous plant.

[0142] According to some embodiments of the invention, the plant is a monocotyledonous plant.

[0143] According to some embodiments of the invention, a plant cell is provided that exogenously expresses the polynucleotide of some embodiments of the invention, the nucleic acid construct of some embodiments of the invention and / or the polypeptide of some embodiments of the invention.

[0144] In some embodiments, after stable transformation, plant propagation is carried out. In some embodiments, propagation by seeds is used. In some embodiments, the transformed plant is regenerated by micropropagation, which provides rapid and consistent reproduction of the transformed plants.

[0145] Micropropagation is a process of cultivating new-generation plants from a single piece of tissue that has been excised from a mother plant or selected cultivar. This process allows for the mass reproduction of plants with the preferred tissue that expresses the fusion protein. The new-generation plants produced are genetically identical to the original plant and possess all its characteristics. Micropropagation allows for the mass production of quality plant material in a short period of time and offers rapid multiplication of selected cultivars, preserving the characteristics of the original transgenic or transformed plant. The advantages of plant cloning are the speed of multiplication of Petition 870250109327, dated 11 / 28 / 2025, page 53 / 107 50 / 78 plants and the quality and uniformity of the plants produced.

[0146] Micropropagation is a multi-stage procedure that requires altering the culture medium or growing conditions between stages. Thus, the micropropagation process involves four basic stages: Stage one, initial tissue culture; stage two, tissue culture multiplication; stage three, plant differentiation and formation; and stage four, greenhouse cultivation and hardening. During stage one, initial tissue culture, the tissue culture is established and certified as free of contaminants. During stage two, the initial tissue culture is multiplied until a sufficient number of tissue samples are produced from the seedlings to meet production targets. During stage three, the tissue samples grown in stage two are divided and grown into individual seedlings.In the fourth phase, the transformed seedlings are transferred to a hardening greenhouse, where the plants' tolerance to light is gradually increased so that they can be grown in their natural environment.

[0147] According to one aspect of some embodiments of the invention, a method is provided for cultivating a crop comprising sowing and / or planting seedlings of a plant transformed with the exogenous polynucleotide of the invention, for example, the polynucleotide encoding the polypeptide of some embodiments of the invention, wherein the plant is derived from plants that have been transformed with the exogenous polynucleotide and that have been selected for at least one selected characteristic of the group consisting of greater tolerance to abiotic stress, greater water use efficiency, greater growth rate, greater vigor, greater biomass, greater oil content, greater yield, greater seed yield, greater fiber yield, greater fiber quality, greater fiber length, greater photosynthetic capacity and / or greater fertilizer use efficiency (e.g., greater nitrogen use efficiency) compared with an untransformed plant. Petition 870250109327, dated 11 / 28 / 2025, p. 54 / 107 51 / 78

[0148] According to some embodiments of the invention, the increase in corn productivity may manifest itself as one or more of the following: increase in the number of plants per cultivated area, increase in the number of ears per plant, increase in the number of rows per ear, number of grains per row of ears, grain weight, weight of one thousand grains (weight of 1000), ear length / diameter, increase in oil content per grain and increase in starch content per grain.

[0149] According to some embodiments of the invention, the increase in plant productivity may be determined by several parameters.For example, increased rice productivity can manifest itself through an increase in one or more of the following: number of plants per cultivated area, number of panicles per plant, number of spikelets per panicle, number of flowers per panicle, increased seed filling rate, increased thousand-grain weight (1000-grain weight), increased oil content per seed, increased starch content per seed, increased number of tillers, among others. An increase in productivity can also result from modified architecture or can occur due to modified architecture.

[0150] According to some disclosure methods, increased soybean productivity can be manifested by an increase in one or more of the following: number of plants per growing area, number of pods per plant, number of seeds per pod, increased seed filling rate, increased thousand-seed weight (1000-seed weight), reduced pod breakage, increased oil content per seed, increased protein content per seed, among others. An increase in productivity can also result from modified architecture or can occur due to this modified architecture.

[0151] According to some disclosure methods, increased canola productivity can be manifested by an increase in one or more of the following: number of plants per growing area, number of pods per plant, number of seeds per pod, increase in seed filling rate, increase in thousand-seed weight Petition 870250109327, dated 11 / 28 / 2025, p. 55 / 107 52 / 78 seeds (weight of 1000), reduced pod breakage, increased oil content per seed, among others. An increase in productivity can also result from modified architecture or can occur due to this modified architecture.

[0152] According to some disclosure methods, increased cotton yield may be manifested by an increase in one or more of the following: number of plants per growing area, number of bolls per plant, number of seeds per boll, increased seed filling rate, increased thousand-seed weight (1000-seed weight), increased oil content per seed, improved fiber length, fiber strength, among others. An increase in yield may also result from modified architecture or may occur due to modified architecture.

[0153] In some embodiments, the increase in oil content can be determined by extracting the oil from the seed or vegetative part of the plant. Briefly, lipids (oil) can be removed from the plant (e.g., seed) by grinding the plant tissue in the presence of specific solvents (e.g., hexane or petroleum ether) and extracting the oil in a continuous extractor. Indirect analysis of oil content can be performed using several known methods, such as Nuclear Magnetic Resonance (NMR) Spectroscopy, which measures the resonance energy absorbed by hydrogen atoms in the liquid state of the sample [See, for example, Conway T F. and Earle F R., 1963, Journal of the American Oil Chemists' Society; Springer Berlin / Heidelberg, ISSN: 0003-021X (Print) 1558-9331 (Online)]; Near-Infrared (NI) Spectroscopy, which utilizes the absorption of near-infrared energy (1100-2500 nm) by the sample; and a method described in WO / 2001 / 023884, which is based on extracting oil from a solvent, evaporating the solvent in a gas stream that forms oil particles, and directing a light onto the gas stream and oil particles that forms a detectable reflected light.

[0154] Non-limiting examples of abiotic stress conditions Petition 870250109327, dated 11 / 28 / 2025, pp. 56 / 107 53 / 78 include salinity, osmotic stress, drought, water deprivation (e.g., drought), excess water (e.g., flooding), etiolation, low temperature (e.g., cold stress), high temperature, heavy metal toxicity, anaerobiosis, nutrient deficiency (e.g., nitrogen deficiency or limitation), excess nutrients, air pollution, and UV radiation.

[0155] According to some embodiments of the disclosure, the method further comprises cultivating the plant expressing the exogenous polynucleotide under limiting fertilizer conditions (e.g., limiting nitrogen conditions). Non-limiting examples include cultivating the plant in soils with low nitrogen content (40-50% of the nitrogen content present under normal or optimum conditions), or even under severe nitrogen deficiency (0-10% of the nitrogen content present under normal or optimum conditions), where normal or optimum conditions include approximately 6-15 mM of nitrogen, e.g., 6-10 mM of nitrogen).

[0156] According to some embodiments of the disclosure, the method further comprises cultivating the plant expressing the exogenous polynucleotide under drought conditions (e.g., water deprivation conditions). In some embodiments, drought conditions occur when the plant is not watered for about 2 days, about 4 days, about 6 days, about 7 days, about 10 days, about 20 days, about 30 days, about 60 days, or any number of days between 2 and 60 days. In some embodiments, plants that exogenously express polynucleotide(s), nucleic acid constructs, and / or polypeptide(s) of the invention exhibit greater drought tolerance compared to plants that do not exogenously express polynucleotide(s), nucleic acid constructs, and / or polypeptide(s) of the invention. In some embodiments, methods for determining the plant's drought tolerance include, among others, measuring root length and leaf wilting.

[0157] In some forms, once expressed in the cell Petition 870250109327, dated 11 / 28 / 2025, pp. 57 / 107 54 / 78 vegetable or whole plant, the level of the polypeptide encoded by the exogenous polynucleotide can be determined by well-known methods in the art, such as activity assays, Western blots using antibodies capable of specifically binding to the polypeptide, enzyme-linked immunosorbent assay (ELISA), radioimmunoassays (RIA), immunohistochemistry, immunocytochemistry, immunofluorescence and similar methods, or combinations thereof.

[0158] Methods for determining the level in the plant of RNA transcribed from exogenous polynucleotides are well known in the art and include, for example, Northern blot analysis, reverse transcription polymerase chain reaction (RT-PCR) analysis (including quantitative, semi-quantitative or real-time RT-PCR) and situ mRNA hybridization or combinations thereof.

[0159] The sequence information and annotations revealed by the present teachings can be used in favor of classical genetic improvement. Thus, subsequence data of the polynucleotides described above can be used as markers for marker-assisted selection (MAS), in which a marker is used for indirect selection of a genetic determinant or determinants of a trait of interest (e.g., biomass, growth rate, oil content, yield, tolerance to abiotic stress, water use efficiency, nitrogen use efficiency and / or fertilizer use efficiency).The nucleic acid data from these teachings (DNA or RNA sequence) may contain or be linked to polymorphic sites or genetic markers in the genome, such as restriction fragment length polymorphism (RFLP), microsatellite and single nucleotide polymorphism (SNP), DNA fingerprinting (DFP), amplified fragment length polymorphism (AFLP), expression-level polymorphism, encoded polypeptide polymorphism, and any other polymorphism in the DNA or RNA sequence.

[0160] Examples of selections assisted by markers include, Petition 870250109327, dated 11 / 28 / 2025, page 58 / 107 55 / 78 but without limitation, selection for a morphological trait (e.g., a gene affecting shape, coloration, male sterility, or resistance, such as the presence or absence of arbor, leaf sheath coloration, height, grain color, rice aroma); selection for a biochemical trait (e.g., a gene encoding a protein that can be extracted and observed; e.g., isoenzymes and storage proteins); selection for a biological trait (e.g., races of pathogens or biotypes of insects based on pathogen-host or parasite-host interaction) can be used as a marker, since the genetic makeup of an organism can affect its susceptibility to pathogens or parasites).

[0161] In some embodiments, the polynucleotides and polypeptides described herein can be used in a wide range of economically viable plants, in a safe and cost-effective manner.

[0162] In some embodiments, plant lines that exogenously express the polynucleotide or polypeptide of the invention are screened to identify those that exhibit the greatest increase in the desired plant trait.

[0163] Thus, according to a further embodiment of the present disclosure, a method is provided for evaluating a plant characteristic, the method comprising: (a) expressing in a plant or a part thereof the nucleic acid construct of some embodiment of the invention; and (b) evaluating a plant characteristic in comparison with a wild-type plant of the same type (e.g., a plant not transformed with the claimed biomolecules); thereby evaluating the plant characteristic.

[0164] According to one aspect of some embodiments of the disclosure, a method is provided for producing a culture comprising cultivating a plant culture expressing an exogenous polynucleotide comprising a nucleic acid sequence encoding a polypeptide at least about 80%, at least about 81%, at least about 82%, at least about 83%, Petition 870250109327, dated 11 / 28 / 2025, p. 59 / 107 56 / 78 at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99% or more, say 100% homologous (i.e., identical) to the selected amino acid sequence from the group consisting of SEQ ID NOs: 34-38, wherein the plant is derived from a plant (mother plant) that has been transformed to express the exogenous polynucleotide and that has been selected for greater tolerance to abiotic stress, greater water use efficiency, greater growth rate, greater vigor, greater biomass, higher oil content, higher yield, higher seed yield, higher fiber yield, higher fiber quality, greater fiber length.greater photosynthetic capacity and / or greater efficiency in the use of fertilizers (e.g., greater efficiency in the use of nitrogen) compared with a control plant, thus producing the crop.

[0165] According to one aspect of some embodiments of the disclosure, a method of cultivating a crop is provided comprising sowing seeds and / or planting seedlings of a plant transformed with the exogenous polynucleotide of the invention, for example, the polynucleotide encoding the polypeptide of some embodiments of the invention, wherein the plant is derived from plants that have been transformed with the exogenous polynucleotide and that have been selected for at least one selected characteristic of the group consisting of greater tolerance to abiotic stress, greater efficiency in water use, greater growth rate, greater vigor, greater biomass, greater oil content, greater yield, greater seed yield, greater fiber yield, greater fiber quality,Greater fiber length, greater photosynthetic capacity, and / or greater fertilizer use efficiency (e.g., greater nitrogen use efficiency) compared to an unprocessed plant. Petition 870250109327, dated 11 / 28 / 2025, pp. 60 / 107 57 / 78

[0166] It is recognized that certain features of the disclosure, which are, for greater clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, several features of the invention, which are, for greater brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination or as appropriate in any other described embodiment of the invention. Certain features described in the context of several embodiments should not be considered essential features of those embodiments unless the embodiment is inoperative without those elements.

[0167] Several embodiments and aspects of the present invention, as outlined above and as claimed in the Claims section below, find experimental support in the following examples. EXAMPLES

[0168] Reference is now made to the following examples, which, together with the descriptions above, illustrate some embodiments of the invention in a non-limiting manner.

[0169] In general, the nomenclature used herein and the laboratory procedures employed in the present invention include molecular, biochemical, microbiological, and recombinant DNA techniques. Such techniques are exhaustively explained in the literature. See, for example, “Molecular Cloning: A Laboratory Manual” Sambrook et al., (1989); “Current Protocols in Molecular Biology” Volumes I-III Ausubel, RM, ed. (1994); Perbal, “A Practical Guide to Molecular Cloning”, John Wiley & Sons, New York (1988); Watson et al., “Recombinant DNA”, Scientific American Books, New York; Birren et al. (eds) “Genome Analysis: A Laboratory Manual Series”, Vols. 1-4, Cold Spring Harbor Laboratory Press, New York (1998); methodologies as established. Petition 870250109327, dated 11 / 28 / 2025, pp. 61 / 107 58 / 78 in US patents nos. 4,666,828; 4,683,202; 4,801,531; 5,192,659 and 5,272,057; “Cell Biology: A Laboratory Handbook”, volumes I-III Cellis, JE, ed. (1994); “Current Protocols in Immunology”, volumes I-III Coligan JE, ed. (1994); Stites et al. (eds); available immunoassays are extensively described in the scientific and patent literature; “Nucleic Acid Hybridization”, Hames, BD and Higgins SJ, eds. (1985); “Transcription and Translation”, Hames, BD and Higgins SJ, eds. (1984); “Immobilized Cells and Enzymes” IRL Press, (1986); “A Practical Guide to Molecular Cloning” Perbal, B., (1984); “PCR Protocols: A Guide to Methods and Applications”, Academic Press, San Diego, California (1990); Marshak et al., “Strategies for Protein Purification and Characterization — A Laboratory Course Manual” CSHL Press (1996); all of which are incorporated by reference as if fully described herein.Other general references are provided throughout this document. The procedures contained herein are believed to be well known in the art and are provided for the reader's convenience. All information contained herein is incorporated herein by reference. Example 1: Obtaining cDNA from the FTO gene

[0170] FTO genes were identified from the National Center for Biotechnology Information (NCBI) database. FTO nucleic acid sequences (SEQ ID NOs: 31-38) and amino acid sequences (SEQ ID NOs: 1-30) were obtained. The corresponding cDNAs were acquired or synthesized by GenScript. Example 2: FTO expression in E. coli

[0171] FTO cDNAs (SEQ ID NOs: 1-9) were amplified after being cloned into the pET28a(+) vector using NdeI / NotI restriction sites. The gene of interest is under the regulation of the T7 promoter / lacI operator and is inducible with IPTG. The average sequence size of the inserted gene Petition 870250109327, dated 11 / 28 / 2025, pp. 62 / 107 The 59 / 78 sample size was 510 bp. The plasmid used for cloning is illustrated in FIG. 1, and the pET-28a plasmid containing acFTO, SEQ ID No. 9, is illustrated in FIG. 2. E. coli transformed with different FTO constructs disclosed here were cultured overnight at 37°C. At OD600, the culture was induced from 0.6 to 1.0 with 200 g / L of 100 mM isopropyl e-d-1-thiogalactopyranoside (IPTG) in each flask and moved to 16°C with agitation at 80 rpm and incubated overnight. After approximately 18 hours, the culture was harvested by centrifugation at 4000 K for 10 minutes. The supernatant was discarded, and the pellets were harvested and frozen at -20°C. 1 ml samples were collected at induction and harvest.

[0172] The pellets from the saved samples were resuspended in running buffer equivalent to DO (1 gl for each DO unit) and boiled for 10 minutes. The samples were run on an acrylamide gel with a gradient of 4-20% at a constant 140 mAmps. The resulting gel was stained with 50 ml of LabSafe™ GEL Blue for 20 minutes and decolorized overnight with water, as exemplified in FIG. 3. As shown in FIG. 3, the expression of several FTO homologs was observed after induction with IPTG. Example 3: Protein purification

[0173] Frozen cell pellets were resuspended in 10 ml of lysis buffer and lysed by passing them through a Frech press three times. The lysis buffer was prepared by adding, immediately before use, 5 mM beta-mercaptoethanol, lysozyme, and protease inhibitor to 50 mM Tris-HCl, pH 8.0, 300 mM NaCl. Debris was removed by centrifugation at 4000 rpm for 10 min. The supernatant was transferred to a new tube and the pellet was discarded. Ni NEBExpress® Spin Columns (S1427L) were used to purify the FTO protein according to the manufacturer's instructions. For each sample, five columns were run and the eluates were pooled. Petition 870250109327, dated 11 / 28 / 2025, pp. 63 / 107 60 / 78 resulting in a total of 1 ml of each. To concentrate and change the buffer to one more suitable for activity assays, Amicon Ultra-4 Centifugal Filter devices with a molecular weight cutoff of 10000K were used. The entire 1 ml sample of each purified enzyme was passed through the column and centrifuged at 4000 rpm for 20 minutes. Five milliliters of 50 mM Tris-HCl buffer, pH 7.5, were added to the column and centrifuged again at 4000 rpm for 15 minutes. The concentrated sample, with approximately 200 µL of each, was recovered from the column and frozen until use. Example 4: Detection of full-length FTO protein in soybeans

[0174] Constructs expressing hFTO homologs were transformed into soybean plants. Samples of soybean plants expressing hFTO (SEQ ID NO:1), acFTO (SEQ ID NO:9), niFTO (SEQ ID NO:8) were extracted and the FTO protein was detected by Western Blots shown in FIG. 13. Example 5: Screening of the A. thaliana ecotype

[0175] Constructs expressing hFTO homologs were transformed into several A. thaliana ecotypes to determine if the response differs between ecotypes. The following ecotypes were selected: Col-0 (standard), Ws-2 (standard), Lan-0 (Lansberg erecta) (standard). Some ecotypes with low freeze tolerance, as defined by Maximilian Boinot et al. 2022, were also selected: C24 (Portugal 41°15'00.0”N 8°27'00.0”W), Can-0 (Canary Islands, low freeze tolerance), different M6A (Guan-Zheng Luo, Alice MacQueen et al. 2015 Nat. Commun.), Sah-0 (Sierra Alhambra - Spain). Other ecotypes with high freeze tolerance, as defined by Maximilian Boinot et al. 2022 plants were selected: Hen-16 (North Sweden, M6A different Guan-Zheng Luo, Alice MacQueen et al. 2015 Nat. Commun.), Ms-0 Petition 870250109327, dated 11 / 28 / 2025, pp. 64 / 107 61 / 78 Moscow (climate similar to Hen-16 - Maximilian Boinot et al. 2022. Plant). Seeds of all ecotypes were stratified for 7 days at 4°C in the dark. Eight (8) days later, the seeds were transferred to the soil. Approximately two weeks later, plant growth was observed. Most ecotypes were observed germinating well (except Col-0 and Sah-0). Two days later, new Col-0 seeds were stratified for sowing two days later. Three weeks after observation, La-0 and C24 plants were cut in preparation for floral immersion. Agro (GV301) was seeded with hFTO and mhFTO constructs. Two days later, single colonies of hFTO and mhFTO were harvested to grow on a small scale. One day later, the plants were transferred from a small-scale agricultural culture to a large-scale one (125 mL flask with 75 mL of LB).One day later, the crop culture was centrifuged and resuspended in buffer with 1% sucrose and 500 pL / L of Silwet-77 (see Clough and Bent, 1997 for details). This culture was placed in a Tupperware container (plastic container) and the flower parts were immersed in it, shaking off the excess. One day later, all the plants were tied together with string and skewers.

[0176] After problems with the Hen-16 and Can-0 plants, the plants were restarted in 1 / 2 MS plates in a tissue culture chamber. The plants were exposed to 24 hours of continuous light in the growth chamber, resulting in stressed seedlings. The seedlings were transferred to soil in another chamber with standard A. thaliana growth conditions (16 hours of light / 8 hours of darkness, 22°C during the day and 19°C at night). Due to the error in the light cycles, the Hen-16 plants, which would normally require several weeks of vernalization at 4°C and short days (8 hours of light / 16 hours of darkness), ended up flowering prematurely under standard conditions.

[0177] This allowed floral immersion of Hen-16 plants with hFTO and the hFTO mutant in Agro. The plants were then tied, staked, and placed in seed collection bags to continue Petition 870250109327, dated 11 / 28 / 2025, pp. 65 / 107 62 / 78 growing. The primary stems were pruned and immersed 5 days later. To help the plants recover from the stem pruning, a weak mixture of water and fertilizer was applied to aid in rapid recovery.

[0178] Unlike the Hen-16 plants, the Can-0 plants were not affected by the erroneous growing conditions and took longer to form primary stems. They were pruned and fertilized like the Hen-16 plants, but the Can-0 plants took longer to recover and produce lateral shoots with inflorescences.

[0179] The harvested seeds were placed at 30°C for 2 days in magenta boxes with dri-rite, with the lid open. After two days of drying, a 5% liquid smoke solution was prepared to add to the seeds (Wright pecan liquid smoke). The seeds were placed in mesh bags (NYLON BIOPSY BAG, 35X50MM, 100 - Electron Microscopy Services via VWR) and sealed by folding and securing them with adhesive tape and a standard paper clip. The seed bags were then placed vertically in magenta boxes and ~75 mL of 5% liquid smoke was added. They were then placed in the fume hood and left to soak overnight. The seeds were then removed from the solution and placed on a benchtop base to absorb the liquid in the fume hood. After allowing the seeds to dry for 3 hours, they were placed in a ziplock plastic bag and moistened with sphagnum moss.The moss was moist but not saturated, and most of the air was removed, but not completely. The seeds were then placed in a box, sealed with tape to prevent light from entering, and kept at 4°C for at least 2 weeks. Afterwards, the seeds could be planted in soil or kept in a seed dryer at 30°C for 2 to 3 days, followed by storage in envelopes for a few more months before planting. The seeds maintained a germination rate of 95% after several years of storage. The seeds were categorized by copy number and weight, as shown in FIG. Petition 870250109327, dated 11 / 28 / 2025, pp. 66 / 107 63 / 78 5. Example 6: Weight of Seteria viridis seeds transformed with FTO

[0180] Constructs expressing hFTO homologs were transformed into Seteria viridis plants. To analyze FTO expression in S. viridis, S. viridis plants expressing GFP-labeled hFTO were processed for visualization by DAPI and GFP. As illustrated in FIG. 14, hFTO-GFP was detected in nuclei. Example 7: Root assay in Seteria viridis

[0181] Next, the root growth of transgenic Seteria virdis plants was determined by comparing each of the mcFTO-expressing plants with null and void vectors. Seteria seeds, which were placed in pots with soil, were returned to 28°C in magenta boxes with dri-rite to prevent spurious germination of the seeds inside the mesh bags. Four days later, 10 to 12 seeds were placed in 1.5 mL epi tubes with 0.1% Triton-X and 20% bleach by volume for surface sterilization. This was done. The seeds were placed in the shaker for 20 minutes for sterilization. The Triton-X and bleach solutions were washed off and followed by 8 washes with sterile autoclaved diH2O. As the seeds were washed with water, they began to absorb water and produce mucilage and project their radicals. This allowed individual seeds to be removed from the water and placed in the bags.The bags were moistened with 15 mL of 1 / 2MS medium and compressed to ensure all the bag material was moist. Using tweezers, each seed was individually placed in a notch to allow germination and plant growth. Once completed, the bags were placed in the Conviron growth chamber with the following settings: light intensity of 450 µmol / s, 12 hours of light / 12 hours of darkness, 30 °C during the day and 22 °C at night. The bags were... Petition 870250109327, dated 11 / 28 / 2025, pp. 67 / 107 64 / 78 samples were checked daily to determine if an additional 1 / 2 MS should be added to the samples for irrigation. This was usually ~5 mL every few days. On day 14, the primary root was traced with a black Sharpie marker. The bag was then backlit with a white light box and photographed for measurement using ImageJ software. The root length was determined and plotted as illustrated in FIGS. 4 and 7. Example 8: ATACseq Analysis (Transposase Accessible Chromatin Assay with Sequencing)

[0182] To determine the accessible chromatin regions, ATAC sequence analysis was performed. The nf-core ATACseq pipeline (Assay for Transposase-Accessible Chromatin using sequencing) doi: 10.5281 / zenodo.2634132 was used to analyze the ATACseq sequencing data following these steps: 1. Raw Sequencing Read Quality Control (FastQC) 2. Sequencing Adapter Adjustment (Trim Galore!) 3. Reference genome-aligned trimmed reads (BWA) 4. Duplicate readings counted and marked (Picard) 5. Identification of broad peaks or accessible chromatin regions based on stacked reads (MACS2) 6. Annotation of peaks related to genetic characteristics (HOMER) - Creation of a consensus peak set across all samples and creation of a tabular file to aid in data filtering (BEDTools) Petition 870250109327, dated 11 / 28 / 2025, pp. 68 / 107 65 / 78 - Counting reads at consensus peaks related to merged alignments at the library level (featureCounts) - Differential accessibility analysis, PCA and clustering (R, DESeq2) 7. Presentation of QC for raw reading, alignment, peak identification and differential accessibility results (ataqv, MultiQC, R)

[0183] The ATAC analysis results are provided in Table 2 below. As shown in Table 2, most of the unique peaks of mcFTO plants are in intergenic regions (between protein-coding genes, which may contain regulatory elements). Table 2 mcFTO hFTOmu t Shared Intergenic 2633 620 6948 Exon 484 952 5732 Promoter - Transcription Start Location 461 742 10006 Transcription End Location 303 573 4549 Intron 216 511 2174 Other 2 0 0 Total 4099 3398 29409 Example 9: Soybean Drying Trial #1 Petition 870250109327, dated 11 / 28 / 2025, pp. 69 / 107 66 / 78

[0184] For this essay, the construction containing AFN008 (Seq. ID No. 9) was transformed into a vector for use, pAFB070 - [pSOLIS21-070]. The empty vector was the binary vector pSOLIS21 (a vector specific for Solis). The positive transformants were transferred to the soil and seeds (T1 seeds) were collected from the plants. The T1 seeds were then planted and genotyped for the presence of AFN008 using a copy number assay. The GOI-positive plants were transplanted into larger pots to generate seeds (T2 seeds). From the events shown above, the mother plant T1 was used to generate T2 seeds. The T2 seeds were used in the drought assay. The plant growth conditions were (14 hours of light / 10 hours of darkness, 25°C during the day, 23°C at night). The plants were grown in Berger BM7-35% soil with high porosity bark and fertilized on Mondays, Wednesdays, and Fridays of each week with Nitrogen:Phosphorus:Potassium (NPK) fertilizer in a 15-16-17 ratio.

[0185] For the trial, 1,801 trays were filled with soil. BM7-35 and saturated. One seed per cell was planted at a depth of approximately 2.5 to 4 cm. Each cell received a stake label with individual seed information. Simultaneously, WT Illini-3025N seeds were sown to create a border / buffer line around all experimental soybean layers.

[0186] Between days 12 and 13, the newest leaf was cut for tissue collection for DNA extraction and copy number analysis. The plants were allowed to grow until day 17, and drying was initiated for half of the plants. The well-irrigated plants and the drought-treated plants were placed in 1801 trays, arranged in a checkerboard pattern to allow for spacing between plants, as shown below. Ten trays were generated using a randomized block design (RCD) for each of the events, along with the controls. Each event had 15 plants per treatment distributed among the 10 trays. Ten trays were well-irrigated, and ten trays were placed under total drought conditions, with all water retained during Petition 870250109327, dated 11 / 28 / 2025, pp. 70 / 107 67 / 78 throughout the process. The exact positions were determined for the plants, and each map was used to position the correct plant in the correct location for the experiment, as shown in FIG. 15A. FIG. 15B shows the well-irrigated plants and the drought-treated plants surrounded by soybean borders to avoid edge effects and additional water losses (unrelated to drought). After placing the plants in the DCC, two photos were taken of the pots in the trays. There was an aerial photo showing the entire tray for each tray. A side view was also taken, where each pot was aligned by genotype to capture the sprout. Aerial and side photos were taken daily for the next 7 days. After the last aerial photo of the plants on the 7th day of drought treatment, all roots were washed to remove as much soil as possible. Five plants were arranged for an aerial photo with a scale and color correction card to capture root morphology.The total length of the roots was also measured and recorded manually. Determining the wilting score

[0187] Aerial photographs from the last day of drought were then independently examined by two people. The scoring system was as follows: good - 4, low - 3, medium - 2, high - 1, crispy - 0, as illustrated in FIG. 16A. Using the RCBD map, the score for each individual plant was recorded based on appearance. The higher the score, the less wilting; the lower the score, the greater the degree of wilting. Individual scores were calculated for each plant and then combined to obtain an average wilting score specific to the event. The standard deviation was also calculated for each average event. To determine the significance of wilting, a t-test was used to compare the wilting values ​​with the empty vector plant values. Measuring root length Petition 870250109327, dated 11 / 28 / 2025, pp. 71 / 107 68 / 78

[0188] Next, the length of each individual root was measured and recorded per plant. The results for each event were calculated for the treatment with good irrigation or drought. The standard derivation was determined. To determine significance, a t-test was used to compare plants with empty vectors with AFN008 plants. The root length results are shown in FIG. 16B. The asterisk indicates a statistically significant event for plants with empty vectors with a p-value less than 0.05. Example 10: Soybean Drying Trial #2

[0189] For this essay, constructions containing AFN008 (acFTO, Seq ID No. 9) were transformed into a vector for use, pAFB070 [pSOLIS21-070]. The empty vector was the binary vector pSOLIS21 (a Solis-specific vector). Positive transformers were moved into the soil and seeds (T1 seeds) were collected from the plants. T1 seeds were then planted and genotyped for the presence of AFN008 using a copy number assay. GOI-positive plants were transplanted into larger pots to generate seeds (T2 seeds). The T1 mother plant was used to generate T2 seeds. T2 seeds were used in the drought assay. For AFN008m, the initial plants for the drought assay are shown planted in apartments 1801 on the day of sowing. The plant growing conditions were (14 hours of light / 10 hours of darkness, 25°C during the day, 23°C at night).The plants were grown in Berger BM7-35% soil with high porosity bark and fertilized on Mondays, Wednesdays, and Fridays of each week with Nitrogen:Phosphorus:Potassium (NPK) fertilizer in a 15-16-17 ratio.

[0190] For this test, 1801 trays were filled with soil BM7-35 and saturated. One seed per cell was planted at a depth of approximately 2.5 to 4 cm. Each cell received a stake label with individual seed information. Between days 16 and 18, the leaf Petition 870250109327, dated 11 / 28 / 2025, pp. 72 / 107 The youngest plant, 69 / 78, was cut to collect tissue for DNA extraction and copy number analysis. The plants were allowed to grow until day 18. Drying was initiated for half of the plants. The well-irrigated plants and the dry-period plants were placed in 1801 trays and arranged in a checkerboard pattern to allow for spacing between plants, as shown here. A total of 10 trays were generated using a randomized complete block design (RCBD) for each of the events, along with the controls. Each event had 5 plants per treatment distributed among the 10 trays. 10 trays were well-irrigated and 10 trays were placed under total dry conditions, with all water retained throughout the process. The exact positions were determined for the plants and each map used to place the correct plant in the correct position for the experiment, as shown in FIG. 17A. After placing the plants in the RCBD, two photos were taken of the pots in the trays.An aerial photograph was taken showing the entire tray for each plant. A side view was also taken, where each plant was aligned by genotype to capture the sprout. Photographs were then taken daily, both an aerial and side view, for the next 7 days. After taking a final aerial photograph of the plants on day 7 of the drought treatment, all roots were washed to remove as much soil as possible. Five plants were placed for an aerial photograph with a scale and color correction card to capture root morphology. The total root length was also measured and recorded manually. Determining the wilting score

[0191] Aerial photographs from the last day of drought were examined independently by two people. The scoring system was as follows: good - 4, low - 3, medium - 2, high - 1, crispy - 0, as illustrated in FIG. 16A. Using the RCBD map, the score for each individual plant was recorded based on appearance. The higher the score, the less wilting; the lower the score, the greater the degree. Petition 870250109327, dated 11 / 28 / 2025, pp. 73 / 107 70 / 78 wilting. Individual scores were calculated for each plant and then combined to obtain a specific average wilting score for the event. The standard deviation was also calculated for each average event. To determine the significance of wilting, a t-test was used to compare the wilting values ​​of the plants with an empty vector. The wilting score results are shown in FIG. 17B. Measuring root length

[0192] The length of each individual root was measured and recorded per plant. The results for each event were calculated for the treatment with good irrigation or drought. The standard derivation was also determined. To determine significance, a t-test was used to compare plants with empty vectors with AFN008 plants. The root length results are shown in FIG. 17C. The asterisk indicates a statistically significant event for plants with empty vectors with a p-value less than 0.05. Example 11: Soybean drying trial #3 For this experiment, constructs containing AFN004 (mcFTO, Seq. ID #7), AFN007 (niFTO, Seq. ID #8), and AFN011 (mutant control for AFN004) were transformed into a vector for use by Solis. At the time of transformation, empty vector controls were created for each transformation instance. The mutant control of AFN004 has R321Q and R327Q substitutions.

[0193] Positive transformants were transferred to the soil and seeds (T1 seeds) were collected from the plants. The T1 seeds were then planted and genotyped for the presence of AFN008 using a copy number assay. GOI-positive plants were transplanted into larger pots to generate seeds (T2 seeds). The T1 mother plant was used to generate T2 seeds. The T2 seeds Petition 870250109327, dated 11 / 28 / 2025, pp. 74 / 107 71 / 78 were used in the drought trial. For AFN008m, the initial plants for the drought trial are shown planted in the 1801 apartments on the day of sowing. Plant growing conditions were (14 hours of light / 10 hours of darkness, 25°C during the day, 23°C at night). Plants were grown in Berger BM7-35% soil with high porosity bark and were fertilized on the second, fourth, and Friday of each week with Nitrogen:Phosphorus:Potassium (NPK) fertilizer in the 15-16-17 propartement. Seeds were planted and genotyped for the presence of the construct using a copy number assay specific to the promoter region or the antibiotic resistance region. Plants positive for GOI were selected to continue in the drought trial – with the exception of event 570 of AFN004, which did not contain plants containing the construct but was carried forward nonetheless.Table 3 contains information on the number of plants per treatment carried forward in the drought experiment. For the controls AFN004, AFN011, and AFN004_EV, only T1 seeds were available; therefore, they were used in this experiment. For AFN007 and AFN007_EV, T2 seeds were available for use in this experiment; therefore, information on the mother plant is observed only for events AFN007 and EV controls. Table 3 Species Genotype Event Mother Plant # of seeds planted for drought #germinated # positive #null Glycine maximum AFN004 567 na(T1) 20 20 17 3 Glycine AFN004 570 na(T1) 20 20 0 17 Petition 870250109327, dated 11 / 28 / 2025, pp. 75 / 107 72 / 78 maximum glycine maximum AFN004 575 na(T1) 20 18 16 2 maximum glycine AFN004 582 na(T1) 20 19 15 4 maximum glycine AFN004_EV 535 na(T1) 20 18 Na Na maximum glycine AFN011 935 na(T1) 20 20 15 5 Maximum glycine AFN011 939 na(T1) 20 14 13 1 Maximum glycine AFN007 11B25 9 45 45 29 16 Maximum glycine AFN007 11B19B 25 45 40 24 15 Maximum glycine AFN007 11B19A 20 45 43 29 12 Maximum glycine AFN007_EV 11A2 33 40 38 na na Maximum glycine AFN007_EV 11A2 36 40 37 na na Species Genotype Field copy # rxn #no germination % germination success % positive % null Total # plant of the # in the experiment by treatment* Glycine AFN004 0 0 100 85.0 15. 20 7 Petition 870250109327, dated 11 / 28 / 2025, pp. 76 / 107 73 / 78 maximum 0 Maximum glycine AFN004 3 0 100 0.0 100 .0 20 7 Maximum glycine AFN004 0 2 90 88.9 11. 1 20 7 Maximum glycine AFN004 0 1 95 78.9 21. 1 20 7 Maximum glycine AFN004 _EV na 2 90 na na 20 7 Maximum glycine AFN011 0 0 100 75.0 25. 0 20 7 Maximum glycine AFN011 0 6 70 92.9 7.1 20 7 (dry) 6 (well watered) Maximum glycine AFN007 0 0 100.0 64.4 35. 6 45 14 Maximum Glycine AFN007 1 5 88.9 61.5 38. 5 45 12 Maximum glycine AFN007 2 2 95.6 70.7 29. 3 45 14 Maximum glycine AFN007 _EV na 2 95 na na 40 7 Maximum glycine AFN007 _EV na 3 92.5 na na 40 8

[0194] For planting, 1801 trays were filled with soil. BM7-35 and saturated with water. One seed per cell was planted at a depth of 1 to 1.5 inches. Each cell was labeled with a tag indicating individual information about the seed. At the same time Petition 870250109327, dated 11 / 28 / 2025, pp. 77 / 107 74 / 78 at the time the experimental plants were sown, WT IL3025N seeds were planted to create a WT border to surround the experimental mounds and reduce the positional effects and desiccation of the experimental plants at the edges.

[0195] Copy number assays were performed to determine which plants contain the gene of interest. Between days 12 and 13, a portion of the newest leaf was collected for DNA extraction and copy number analysis. Copy number analysis was used to determine null plants (i.e., plants that do not contain the gene of interest) and positive plants (i.e., plants that contain the gene of interest). This information was used to discard the null plants, and only the positive plants advanced to the drying experiment (for this experiment, the exception was event 570 of AFN004, which contained only null plants and was carried forward).

[0196] The plants for the experiment were grown for 17 days under normal conditions (i.e., normal irrigation and fertilization). After 17 days of growth under normal conditions, the plants were divided into two treatments: dry (i.e., total water suspension) and well-irrigated (i.e., normal irrigation schedule).

[0197] Well-irrigated and drought-stricken plants for each event were separated and placed in 1801 compartments in a grid pattern to allow for spacing between plants, as shown above. Plants within each treatment were arranged in a randomized block design (RCD), along with controls to reduce positional effects. In addition, a border of WT plants surrounded the greenhouse bench to prevent border plants from drying out more than plants in the inner position. The number of plants per treatment for each construct / event is shown in Table 3.

[0198] Half of the plants per event were well irrigated and half per event were subjected to total drought conditions (all water was retained) for an additional 6 or 7 days. The dried plants were grown for 6 days without water. Petition 870250109327, dated 11 / 28 / 2025, pp. 78 / 107 Plants with 75 / 78 or fertilizer were measured and sampled on the 6th day of drought. Well-irrigated plants were grown for 7 days under normal conditions and measured and sampled on the 7th day (see measurement and sampling protocols below). Wilt scores were determined after measurement and sampling from photographs.

[0199] After the start of the dry period, photos of the experimental plants (dry and well-irrigated) were taken daily. This included an aerial photo of each tray, as well as a side view photo of all plants per tray, as illustrated in FIG. 18B. Examples of each type of photo are illustrated in FIGS. 18A and 18B. Two photos were taken of the pots from the trays. The photos were taken during the 6th and 7th day for the dry and well-irrigated plants, respectively.

[0200] Measurement and Sampling: On day 6, dried plants were collected for measurement and sampling. Roots were washed to remove soil. Root length was measured from the base of the plant to the root tip and recorded in cm. Total shoot height was measured and recorded for each plant as the length (cm) from the top of the root to the top of the stem. These values ​​were summed to provide the total plant size for analysis. Plants were placed on black velvet for an aerial photograph with a scale and color correction card to capture root morphology, as illustrated in FIG. 18C. The photographs were used to determine the total root area (cm2), as described below. In addition, 4 plants per construct / event per treatment were sampled in liquid nitrogen and stored in dry ice for subsequent molecular analyses.

[0201] The total area of ​​the root was determined using the program PlantCV. The software was used to divide the image into color spaces and then a subsequent mask for the root tissue. The final root mask after cleaning was performed using the “pcv.analze.shape” function. The “pcv.analze.shape” is capable of measuring outputs in pixels such as total area, longest path, width, height, and other metrics. The main measurement used was the “main measurement”. Petition 870250109327, dated 11 / 28 / 2025, pp. 79 / 107 76 / 78 images were processed as .csv file outputs with subsequent analyses performed in R. Area measurements were converted from pixels to cm2 and allowed the data to be normalized to compensate for differences between individual images. The normalized data were then analyzed using an ANOVA to compare the differences in mean root area between groups with a Tukey HSD correction. Determining the wilting score

[0202] Aerial photographs from the last day of drought were independently examined by two people using the following scoring system: good - 4, low - 3, medium - 2, high - 1, crispy - 0, as illustrated in FIG. 16A. Using the RCBD map, the score for each individual plant was recorded based on appearance. The higher the score, the less wilting; the lower the score, the greater the degree of wilting. Individual scores were calculated for each plant and then combined to provide an average wilting score for each event for dried plants. The standard deviation was also calculated for each average event. The wilting score results are presented in FIG. 18E.

[0203] Measurements, including root length (cm), root area (cm2), shoot height (cm), total plant height (cm), as well as wilting score, were calculated for an event for data analysis. The standard deviation for each measurement within an event was calculated. Drought events were compared with their well-irrigated equivalents. Events in both treatments were compared to controls with empty vector and mutant (for ANF004) to determine the effect of the presence of the FTO homolog. The results are presented in Table 4. Plants were photographed as shown in FIG. 18D. The overall results were summarized in a decision table, as illustrated in Table 4, to compare drought indifference, the Petition 870250109327, dated 11 / 28 / 2025, pp. 80 / 107 77 / 78 performance under drought conditions, performance under well-irrigated (“normal”) conditions, and wilting score. Green indicates a favorable outcome for the event. Table 4 Decision Table Plant Information Drought Indifference Performance Under Drought Conditions Genotype Event Root Length (cm) DI Shoot Height (cm) DI Total Plan (cm) DI Root Surface Area (cm2) DI Roots D longer than controls D shoots higher than controls s D Total Plan greater than controls D Root Surface Area (cm2) ni 11B19A NNNNNYNN 11B19B YNNNYNYN 11B25 NNNNYYYY mc 567 YYYNYYYN 570 (all nulls) NNNNNNNN 575 NYNNYYYN 582 NYNNYYYN Decision Table Plant Information Performance Under “Normal” Conditions Genotype Event Roots ND longer than controls ND shoots higher than controls ND Total Plant larger than controls Root Surface Area ND (cm2) Image A wilted plant scores better than controls ni 11B19A NNNNY 11B19B NNNNY 11B25 YYYNY mc 567 NNNNY* 570 (all YYYNY* Petition 870250109327, dated 11 / 28 / 2025, pp. 81 / 107 78 / 78 (nulls) 575 YNYNY* 582 YNYNY* *mut 939 showed the highest wilting score; the assessment does not include 939

Claims

1. A plant comprising plant cells containing a nucleic acid molecule encoding a fat mass and obesity-associated factor (FTO) gene, characterized in that the expression of said nucleic acid molecule results in one or more increases in yield, biomass, growth rate, drought tolerance, and abiotic stress tolerance compared to a plant of the same variety lacking the nucleic acid molecule encoding the FTO gene.

2. A plant, according to Claim 1, characterized in that the said FTO nucleic acid molecule is from vertebrates, invertebrates, algae, or oomycetes.

3. Plant, according to Claim 1, characterized in that the nucleic acid molecule encoding an FTO has at least 80% identity with any of the SEQ ID Nos: 34-38.

4. A plant, according to Claim 1, characterized in that the plant exhibits at least a two-fold increase in biomass compared to a plant that no longer contains the nucleic acid molecule.

5. A plant, according to Claim 1, characterized in that the plant exhibits at least a two-fold increase in productivity compared to a plant that no longer contains the nucleic acid molecule.

6. A plant, according to Claim 1, characterized in that the plant exhibits increased drought tolerance compared to a plant that no longer contains the nucleic acid molecule.

7. Plant, according to Claim 1, characterized in that said plant is selected from the group consisting of Arabadopsis, canola, wheat, corn, soybean, millet, sorghum, cotton, tomato, grape, Petition 870250090461, dated 03 / 10 / 2025, p. 93 / 112 2 / 3 peanut, rice, lettuce, melon, sweet potato, carrot, cucumber, sugarcane, alfalfa, Camelina or PennyCress.

8. A plant according to Claim 1, characterized in that the nucleic acid molecule further comprises a promoter, a nuclear localization signal, a coding sequence, and a termination sequence.

9. Plant according to Claim 1, characterized in that the FTO gene is expressed in a nucleus of said plant cells.

10. Plant, according to Claim 1, characterized in that the FTO gene encodes a polypeptide comprising either of the SEQ IDs 6-30 or 39-71 or a polypeptide comprising at least 90% identity therewith.

11. Method for Generating a Modified Plant, comprising increased yield, growth rate, biomass, vigor, oil content, seed yield, fiber yield, fiber quality, fiber length, photosynthetic capacity, nitrogen use efficiency, drought tolerance and / or abiotic stress tolerance compared with a wild plant of the same variety, characterized in that the method comprises: (a) genetically modifying a plant cell with an exogenous polynucleotide encoding a polypeptide comprising an amino acid sequence with at least 80% identity with the selected sequence from the group comprising any of the SEQ ID NOs: 6-30 and 39-71, and (b) regenerating the modified plant from said plant cell modified with an exogenous polynucleotide.

12. Method for Generating Modified Plant, according to Petition 870250090461, dated 10 / 03 / 2025, page 94 / 112 3 / 3 Claim 11, characterized in that the method further comprises the production of modified plants from the plant produced in step (b).

13. Method for Generating a Modified Plant, according to Claim 11, characterized in that said polypeptide is derived from vertebrates, invertebrates, algae or oomycetes.

14. Method for Generating a Modified Plant, according to Claim 11, characterized in that the modified plant exhibits at least a two-fold increase in biomass compared to a plant that no longer contains the nucleic acid molecule.

15. Method for Generating a Modified Plant, according to Claim 11, characterized in that the modified plant exhibits at least a two-fold increase in productivity compared to a plant that no longer contains the nucleic acid molecule.

16. Method for Generating a Modified Plant, according to Claim 11, characterized in that the modified plant exhibits increased drought tolerance compared to a plant that no longer contains the nucleic acid molecule.

17. Method for Generating a Modified Plant, according to Claim 11, characterized in that the plant is selected from the group consisting of Arabadopsis, canola, wheat, corn, soybean, millet, sorghum, cotton, tomato, grape, peanut, rice, lettuce, melon, sweet potato, carrot, cucumber, sugarcane, alfalfa, Camelina or PennyCress.

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