Plants with modified fatty acid and / or oil content

Abstract

Plants and plant parts comprising altered level or activity of the bZIP transcription factor, one or more mutations in the upstream regulatory region of the bZIP transcription factor gene (e.g., bZIP SIRT element), or having increased fatty acid, oil, and / or sugar content are provided. Methods and compositions for producing such plants and plant parts are also provided.

Description

[0001] PLANTS WITH MODIFIED FATTY ACID AND / OR OIL CONTENT

[0002] RELATED APPLICATIONS

[0003] This application claims priority to U.S. Provisional Application No. 63 / 507,866, filed on June 13, 2023, the content of which is incorporated herein by reference in its entirety.

[0004] SEQUENCE LISTING

[0005] This application contains a Sequence Listing which is submitted herewith in electronically readable format. The Sequence Listing file was created on June 10, 2024, is named “B88552_1570_SL.xml” and its size is 71.3 kb. The entire contents of the Sequence Listing in the sequencelisting.xml file are incorporated by reference herein.

[0006] FIELD OF THE INVENTION

[0007] The present disclosure relates to plants and plant parts having increased basic region leucine zipper motif (bZIP) transcription factor activity and / or an increased fatty acid / oil or sugar content, and associated methods and compositions thereof.

[0008] BACKGROUND OF THE INVENTION

[0009] With the increasing world population and the dwindling supply of arable land available for agriculture, nutrient rich, resilient plants are desired. High fatty acid / oil content is an exemplary desirable trait for plants and seeds. Plant oil, such as seed oil, is used for a variety of purposes including as ingredients for food, fuel (e.g., biodiesel), or industrial products (e.g., soaps, candles, cosmetic products, paints). Further, sugar is a primary driver of flavor in plants. Sweetness and enhanced sugar content are desirable traits and impact the overall flavor acceptance of plants (such as fruits and vegetables) by consumers. Modulating the function of the genes that regulate carbohydrate (e.g., sugar) transport in plants can modulate the fatty acid, oil, and / or sugar content of plants or plant parts, and can provide plants or plant parts (e.g., seeds) with increased fatty acid, oil, and / or sugar content that could have important commercial advantages.

[0010] SUMMARY OF THE INVENTION

[0011] The present disclosure provides plants, plant parts (e.g., seeds), and a population of plants or plant parts comprising altered function of the bZIP transcription factor, comprising one or more mutations in the upstream regulatory region of the bZIP transcription factor gene (e.g., bZIP SIRT element), or having increased fatty acid, oil, and / or sugar content. Also disclosed herein are methods for producing such plants, plant parts, and a population of plants or plant parts.

[0012] In one aspect, the present disclosure provides a plant or plant part comprising increased basic region / leucine zipper motif (bZIP) transcription factor activity compared to a control plant or plant part, wherein the plant or plant part comprises a genetic mutation that increases the bZIP transcription factor activity. The mutation comprises one or more insertions, substitutions, or deletions in an upstream open reading frame (uORF) of at least one native bZIP transcription factor gene or homolog thereof, the uORF comprises a sucrose induced repression of translation (SIRT) element, the mutation alters level or activity of the SIRT element, level or activity of the bZIP transcription factor gene or homolog thereof, or a bZIP transcription factor encoded by the bZIP transcription factor gene or homolog thereof is increased compared to a control plant or plant part, and carbohydrate transport in the plant or plant part is increased compared to a control plant or plant part.

[0013] In some embodiments, the plant or plant part comprises increased fatty acid, oil, and / or sugar content in the plant or plant part (e.g., seeds) compared to a control plant or plant part. In some embodiments, the plant or plant part comprises the oil content that is increased by about 1% or more dry weight (e.g., 0.5-2.5%, 0.8-1.8%, 1-1.6%, e.g., 1.3% or 1.4% dry weight) and / or the sugar content that is increased by about 0.5% or more dry weight (e.g., 0.4%-0.6%) as compared to a control plant or plant part.

[0014] In some embodiments, the mutation is located in an uORF of a bZIP transcription factor gene or homolog thereof that: comprises a nucleic acid sequence having at least 80% sequence identity to any one of SEQ ID NOs: 1-6, wherein the nucleic acid sequence encodes a polypeptide that retains bZIP transcription factor activity; comprises the nucleic acid sequence of any one of SEQ ID NOs: 1-6; encodes a polypeptide comprising an amino acid sequence having at least 80% sequence identity to an amino acid sequence of any one of SEQ ID NOs: 7-12, wherein the polypeptide retains bZIP transcription factor activity; or encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 7-12.

[0015] In some embodiments, the mutation is located at least partially in an SIRT element in the uORF, wherein the SIRT element: comprises a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 13-18, wherein the nucleic acid sequence encodes a polypeptide that retains SIRT activity; comprises the nucleic acid sequence of any one of SEQ ID NOs: 13-18; encodes a polypeptide comprising an amino acid sequence having at least 90% sequence identity to an amino acid sequence of any one of SEQ ID NOs: 19-24, wherein the polypeptide retains SIRT activity; or encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 19-24, before the mutation is located. In some embodiments, the mutation is located at least partially in an SIRT element in the uORF, wherein a 5’ untranslated region (5’ UTR) that contains the uORF comprises a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 48-53, or comprises the nucleic acid sequence of any one of SEQ ID NOs: 48-53, before the mutation is located.

[0016] In some embodiments, the plant or plant part comprises: a deletion of one or more nucleotides of SEQ ID NO: 13 in the SIRT element in the uORF of the Glycine max bZIP123 transcription factor gene; a deletion of one or more nucleotides of SEQ ID NO: 14 in the SIRT element in the uORF of the Glycine max bZIP125 transcription factor gene; a deletion of one or more nucleotides of SEQ ID NO: 15 in the SIRT element in the uORF of the Glycine max bZIP124 transcription factor gene; a deletion of one or more nucleotides of SEQ ID NO: 16 in the SIRT element in the uORF of the Glycine max bZIP126 transcription factor gene; a deletion of one or more nucleotides of SEQ ID NO: 17 in the SIRT element in the uORF of the Glycine max bZIP18 transcription factor gene; and / or a deletion of one or more nucleotides of SEQ ID NO: 18 in the SIRT element in the uORF of the Glycine max bZIP17 transcription factor gene.

[0017] In some embodiments, the mutation is located at least partially in a 5’ end region of the SIRT element and / or the mutation comprises a deletion of 7-25 nucleotides located at least partially in the SIRT element.

[0018] In some embodiments, the plant or plant part comprises a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 30-34 and 54-56, a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 35-39 and 57-58, and / or a mutated 5’ UTR of the Glycine max bZIP17 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 59-62.

[0019] In some embodiments, the plant or plant part comprises (i) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 32, or (ii) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NO: 54 and a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 57.

[0020] In some embodiments, the mutation comprises an out-of-frame mutation of the SIRT element. In some embodiments, the mutation comprises a nonsense mutation of the SIRT element. In some embodiments, the plant or plant part comprises 2 or more genes encoding a bZIP transcription factor. In some embodiments, the 2 or more genes have less than 100% sequence identity to one another.

[0021] In some embodiments, level or activity of one or more molecules regulated by the bZIP transcription factor is modulated, wherein the modulation comprises: an increased baseline level or activity in the absence of sucrose; a decreased suppression or an increased increase of the level or activity in the presence of sucrose relative to the absence of sucrose; and / or an increased level or activity in the presence of sucrose, relative to the control plant or plant part.

[0022] In some embodiments, the one or more molecules regulated by the bZIP transcription factor comprises a sugar transporter and / or a cell-wall invertase. In some embodiments, the sugar transporter is encoded by Glyma.10G217900, G!yma.02G075000, Glyma.02G218600, Glyma.l6G157100, Glyma.l6G 156800, Glyma.02G 124100, Glyma.01G067600, or homolog of any thereof, and / or the cell wall invertase is encoded by Glyma.13G349300, Glyma.15G024600, Glyma. 14G096600, Glyma.16G 175800, or homolog of any thereof.

[0023] In some embodiments, the one or more molecule regulated by the bZIP transcription factor further comprises asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, and / or trehalose-6-phosphate phosphatase (TPP6).

[0024] In some embodiments, the plant or plant part is a legume. In some embodiments, the plant or plant part is selected from the group consisting of The plant or plant part of claim 16, wherein said plant or plant part is selected from the group consisting of soybean (Glycine max), beans (Phaseolus spp.), common bean (Phaseolus vulgaris), fava bean (Vicia faba , mung bean (Vigna radiata), pea (Pisum sativum), chickpea (Cicer arietinum), peanut (Arachis hypogaea), lentils (Lens culinaris, Lens esculenta), lupins (Lupinus spp.), white lupin (Lupinus albus), mesquite (Prosopis spp.), carob (Ceratonia siliqua), tamarind (Tamarindus indica), alfalfa (Medicago sativd), barrel medic (Medicago truncatuld), birdsfood trefoil (Lotus japonicus), licorice (Glycyrrhiza glabra , and clover (Trifolium spp.). In some embodiments, the plant or plant part is Glycine max.

[0025] In some embodiments, the plant or plant part is selected from the group consisting of com (Zea mays), Brassica species, Brassica napus, Brassica rapa, Brassica juncea, rice (Oryza sativd), rye (Secale cereale), sorghum (Sorghum bicolor, Sorghum vulgare), millet, pearl millet (Pennisetum glaucum), proso millet (Panicum miliaceum), foxtail millet (Setaria italic ), finger millet (Eleusine coracan ), sunflower (Helianthus annuus), safflower (Carthamus tinctorius), wheat (Triticum aestivum), tobacco (Nicotiana tabacum), potato (Solanum tuberosum), peanuts (Arachis hypogaea), cotton (Gossypium barbadense, Gossypium hirsutum), sweet potato (Ipomoea balaliis). cassava (Manihot esculenta), coffee (Coffea spp.), coconut (Cocos nucifera), pineappleAnanas comosus). citrus trees Citrus spp.), cocoa (Theobroma cacao), tea (Camellia sinensis), banana Musa spp.), avocado Per sea americana), fig Ficus casica), guava (Psidium guajava), mango (Mangifera indica), olive (Olea europaea), papaya (Carica papaya), cashew (Anacardium occidentale), macadamia (Macadamia integrifolia), almond (Prunus amygdalus), sugar beets (Beta vulgaris), sugarcane (Saccharum spp.), oats, barley, vegetables, ornamentals, and conifers.

[0026] In some embodiments, the plant or plant part is a seed.

[0027] In one aspect, the present disclosure provides a population of plants or plant parts comprising the plant or plant part provided herein, wherein the population comprises increased bZIP transcription factor activity, increased carbohydrate transport, and / or increased fatty acid, oil, and / or sugar content in seeds compared to a control population. In some embodiments, the plant or plant part is a seed, and the population is a population of seeds.

[0028] In one aspect, the present disclosure provides a method for increasing basic region / leucine zipper motif (bZIP) transcription factor activity in a plant or plant part, the method comprising introducing a genetic mutation that increases the bZIP transcription factor activity into the plant or plant part. The mutation comprises one or more insertions, substitutions, or deletions in an upstream open reading frame (uORF) of at least one native bZIP transcription factor gene or homolog thereof, the uORF comprises a sucrose induced repression of translation (SIRT) element, the mutation alters level or activity of the SIRT element, wherein the mutation increases level or activity of the bZIP transcription factor gene or homolog thereof, or a bZIP transcription factor encoded by the bZIP transcription factor gene or homolog thereof, and the mutation increases carbohydrate transport in the plant or plant part.

[0029] In some embodiments, the method further comprises introducing the genetic mutation that increases the bZIP transcription factor activity into a plant cell, and regenerating the plant or plant part from the plant cell.

[0030] In some embodiments, fatty acid, oil, and / or sugar content is increased in the plant or plant part (e.g., seeds) compared to a control plant or plant part. In some embodiments, the oil content is increased by about 1% or more dry weight (e.g., 0.5-2.5%, 0.8-1.8%, 1-1.6%, e.g., 1.3% or 1.4% dry weight) and / or the sugar content is increased by about 0.5% or more dry weight (e.g., about 0.3-0.7%, e.g., 0.5% dry weight) in the plant or plant part compared to a control plant or plant part.

[0031] In some embodiments, the mutation is introduced into an uORF of a bZIP transcription factor gene or homolog thereof that: comprises a nucleic acid sequence having at least 80% sequence identity to any one of SEQ ID NOs: 1-6, wherein the nucleic acid sequence encodes a polypeptide that retains bZIP transcription factor activity; comprises the nucleic acid sequence of any one of SEQ ID NOs: 1-6; encodes a polypeptide comprising an amino acid sequence having at least 80% sequence identity to an amino acid sequence of any one of SEQ ID NOs: 7-12, wherein the polypeptide retains bZIP transcription factor activity; or encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 7-12.

[0032] In some embodiments, the mutation is introduced at least partially in an SIRT element in the uORF, wherein the SIRT element: comprises a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 13-18, wherein the nucleic acid sequence encodes a polypeptide that retains SIRT activity; comprises the nucleic acid sequence of any one of SEQ ID NOs: 13-18; encodes a polypeptide comprising an amino acid sequence having at least 90% sequence identity to an amino acid sequence of any one of SEQ ID NOs: 19-24, wherein the polypeptide retains SIRT activity; or encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 19-24, before the mutation is introduced. In some embodiments, the mutation is introduced at least partially in an SIRT element in the uORF, wherein a 5’ untranslated region (5’ UTR) that contains the uORF: comprises a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 48-53, wherein said nucleic acid sequence encodes a polypeptide that retains SIRT activity; or comprises the nucleic acid sequence of any one of SEQ ID NOs: 48-53, before the mutation is introduced.

[0033] In some embodiments, the mutation comprises: a deletion of one or more nucleotides of SEQ ID NO: 13 in the SIRT element in the uORF of the Glycine max bZIP 123 transcription factor gene; a deletion of one or more nucleotides of SEQ ID NO: 14 in the SIRT element in the uORF of the Glycine max bZIP 125 transcription factor gene; a deletion of one or more nucleotides of SEQ ID NO: 15 in the SIRT element in the uORF of the Glycine max bZIP 124 transcription factor gene; a deletion of one or more nucleotides of SEQ ID NO: 16 in the SIRT element in the uORF of the Glycine max bZIP 126 transcription factor gene; a deletion of one or more nucleotides of SEQ ID NO: 17 in the SIRT element in the uORF of the Glycine max bZIP 18 transcription factor gene; and / or a deletion of one or more nucleotides of SEQ ID NO: 18 in the SIRT element in the uORF of the Glycine max bZIP 17 transcription factor gene.

[0034] In some embodiments, the mutation is introduced at least partially in a 5’ end region of the SIRT element, and / or the mutation comprises a deletion of 7-25 nucleotides located at least partially in the SIRT element.

[0035] In some embodiments, the plant or plant part comprises a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 30-34 and 54-56, a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 35-39 and 57-58, and / or a mutated 5’ UTR of the Glycine max bZIP17 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 59-62, after the mutation is introduced.

[0036] In some embodiments, the plant or plant part comprises (i) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 32, or (ii) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NO: 54 and a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 57, after the mutation is introduced.

[0037] In some embodiments, introducing the mutation comprises introducing an out-of-frame mutation into the SIRT element.

[0038] In some embodiments, the method further comprises introducing editing reagents or a nucleic acid construct encoding the editing reagents into the plant, plant part, or plant cell.

[0039] In some embodiments, the editing reagents comprise at least one nuclease, wherein the nuclease cleaves a target site in the uORF of the at least one native bZIP transcription factor or homolog thereof in the plant, plant part, or plant cell, and the mutation is introduced at the cleaved target site.

[0040] In some embodiments, the at least one nuclease comprises a CRISPR nuclease. In some embodiments, the CRISPR nuclease is a Type II CRISPR system nuclease, a Type V CRISPR system nuclease, a Cas9 nuclease, a Cast 2a (Cpfl) nuclease, or a Cmsl nuclease. In some embodiments, the CRISPR nuclease is a Casl2a nuclease or an ortholog thereof.

[0041] In some embodiments, the editing reagents comprise one or more guide RNAs (gRNAs). In some embodiments, the one or more gRNAs comprise a nucleic acid sequence complementary to a region of a genomic DNA sequence of the uORF or the SIRT element in the at least one native bZIP transcription factor or homolog thereof in the plant or plant part.

[0042] In some embodiments, at least one of the one or more gRNAs comprise a nucleic acid sequence encoded by: a nucleic acid sequence that shares at least 80% sequence identity with the nucleic acid sequence of any one of SEQ ID NOs: 25-29; or a nucleic acid sequence of SEQ ID NOs: 25-29.

[0043] In some embodiments, level or activity of one or more molecules regulated by the bZIP transcription factor in the plant or plant part is modulated, wherein the modulation comprises: an increased baseline level or activity in the absence of sucrose; a decreased suppression or an increased increase of the level or activity in the presence of sucrose relative to the absence of sucrose; and / or an increased level or activity in the presence of sucrose, relative to the control plant or plant part.

[0044] In some embodiments, the one or more molecules regulated by the bZIP transcription factor comprises a sugar transporter and / or a cell-wall invertase. In some embodiments, the sugar transporter is encoded by Glyma.10G217900, G!yma.02G075000, Glyma.02G218600, Glyma.l6G157100, Glyma.l6G 156800, Glyma.02G124100, Glyma.01G067600, or homolog of any thereof, and / or the cell wall invertase is encoded by Glyma.13G349300, Glyma.15G024600, Glyma. 14G096600, Glyma.16G 175800, or homolog of any thereof.

[0045] In some embodiments, the one or more molecule regulated by the bZIP transcription factor further comprises asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, and / or trehalose-6-phosphate phosphatase (TPP6).

[0046] In some embodiments, the plant or plant part is a legume. In some embodiments, the plant or plant part is selected from the group consisting of soybean (Glycine max), beans (Phaseolus spp.), common bean (Phaseolus vulgaris), fava bean (Vicia faba , mung bean (Cigna radiata), pea (Pisum sativum), chickpea (Cicer arietinum), peanut (Arachis hypogaea), lentils (Lens culinaris, Lens esculenta), lupins (Lupinus spp.), white lupin (Lupinus albus), mesquite (Prosopis spp.), carob (Ceratonia siliqua), tamarind (Tamarindus indica), alfalfa (Medicago saliva), barrel medic (Medicago truncatula), birdsfood trefoil (Lotus japonicus), licorice (Glycyrrhiza glabra , and clover (Trifolium spp.). In some embodiments, the plant or plant part is Glycine max.

[0047] In some embodiments, the plant or plant part is selected from the group consisting of com (Zea mays), Brassica species, Brassica napus, Brassica rapa, Brassica juncea, rice (Oryza saliva , rye (Secale cereale), sorghum (Sorghum bicolor, Sorghum vulgare), millet, pearl millet (Pennisetum glaucum), proso millet (Panicum miliaceum), foxtail millet (Setaria italica), finger millet (Eleusine coracana), sunflower (Helianthus annuus), safflower (Carthamus tinctorius), wheat (Triticum aestivum), tobacco (Nicotiana tabacum), potato (Solanum tuberosum), peanuts (Arachis hypogaea , cotton (Gossypium barbadense, Gossypium hirsutum), sweet potato (Ipomoea batatus), cassava (Manihot esculenta , coffee (Coffea spp.), coconut (Cocos nucifera), pineapple (Ananas comosus), citrus trees (Citrus spp.), cocoa (Theobroma cacao), tea (Camellia sinensis), banana (Musa spp.), avocado (Per sea americana), fig (Ficus casica), guava (Psidium guajava), mango (Mangifera indica , olive (Olea europaea), papaya (Carica papaya , cashew (Anacardium occidental , macadamia (Macadamia integrifolia), almond (Prunus amygdalus), sugar beets (Beta vulgaris , sugarcane (Saccharum spp.), oats, barley, vegetables, ornamentals, and conifers. In one aspect, the present disclosure provides a plant or plant part produced by the method provided herein, wherein the plant or plant part comprises increased bZIP transcription factor activity compared to a control plant or plant part.

[0048] In some embodiments, the plant or plant part comprises increased fatty acid, oil, and / or sugar content in seeds compared to a control plant or plant part. In some embodiments, the plant or plant part is a seed.

[0049] In one aspect, the present disclosure provides a population of plants or plant parts produced by the method provided herein, wherein the population comprises increased bZIP transcription factor activity, increased carbohydrate transport, and / or increased fatty acid, oil, and / or sugar content in seeds compared to a control population.

[0050] In one aspect, the present disclosure provides a population of plants or plant parts of claim 51, wherein the plant or plant part is a seed, and the population is a population of seeds.

[0051] In one aspect, the present disclosure provides a seed composition produced from the plant or plant part or the population of plants or plant parts of provided herein.

[0052] In one aspect, the present disclosure provides an oil composition produced from the plant or plant part or the population of plants or plant parts provided herein.

[0053] In one aspect, the present disclosure provides a nucleic acid molecule comprising:

[0054] In one aspect, the present disclosure provides a nucleic acid molecule comprising a nucleic acid sequence of a 5’ untranslated region (5’ UTR) of a bZIP transcription factor gene comprising a mutated sucrose induced repression of translation (SIRT) element, wherein said bZIP transcription factor gene comprises a nucleic acid sequence of any one of SEQ ID NOs: 1-6, or wherein the 5’ UTR comprises the nucleic acid sequence of any one of SEQ ID NOs: 48-53 comprising one or more insertions, substitutions, or deletions therein; or a nucleic acid sequence of a mutated SIRT element, wherein the nucleic acid sequence comprises any one of SEQ ID NOs: 13-18 comprising one or more insertions, substitutions, or deletions therein, wherein the nucleic acid molecule increases level or activity of an operably-linked polynucleotide of interest compared to a control nucleic acid molecule without the mutation. In some embodiments, the nucleic acid sequence of the 5’ UTR comprising a mutated SIRT element comprises any one of SEQ ID NOs: 30-39 and 54-62.

[0055] In one aspect, the present disclosure provides a DNA construct comprising, in operable linkage: a promoter that is functional in a plant cell; the nucleic acid molecule of claim 55 or 56; and a polynucleotide of interest.

[0056] In one aspect, the present disclosure provides a cell comprising the nucleic acid molecule or the DNA construct provided herein. In some embodiments, the cell is a plant cell or a bacterial cell. BRIEF DESCRIPTION OF THE DRAWINGS

[0057] FIG. 1 depicts an expression profile of a soybean bZIP gene copy GmZIP123 (Glyma.06G010200) in various tissues of two varieties of soybean plants based on a soy expression database.

[0058] FIG. 2 depicts an expression profile of a soybean bZIP gene copy GmZIP125 (Glyma. 04G010300) in various tissues of two varieties of soybean plants based on a soy expression database.

[0059] FIG. 3 depicts an expression profile of a soybean bZIP gene copy GmZIP124 (Glyma.12G040600) in various tissues of two varieties of soybean plants based on a soy expression database.

[0060] FIG. 4 depicts an expression profile of a soybean bZIP gene copy GmZIP126 (Glyma.llG 114800) in various tissues of two varieties of soybean plants based on a soy expression database.

[0061] FIG. 5 depicts an expression profile of a soybean bZIP gene copy GmZIP18 (Glyma.14G071400) in various tissues of two varieties of soybean plants based on a soy expression database.

[0062] FIG. 6 depicts an expression profile of a soybean bZIP gene copy GmZIP17 (Glyma.17G253200) in various tissues of two varieties of soybean plants based on a soy expression database. In FIGs. 1-6, FPKM stands for fragments per kilobase of exon per million reads.

[0063] FIG. 7 depicts expression levels of firefly luciferase in soybean protoplasts that have been transfected with a construct containing GmbZIP123 5’ UTR with a mutated SIRT element (182- lObp, 182-l lbp, 183-10bp, 183-8bp, or 183-7bp), or wild-type GmbZIP123 5’ UTR (WT), operably linked to a polynucleotide encoding firefly luciferase.

[0064] FIG. 8 depicts expression levels of firefly luciferase in soybean protoplasts that have been transfected with a construct containing GmbZIP125 5’ UTR with a mutated SIRT element (206- lObp, 206-1 Ibp, 207-10bp, 207-8bp, or 207-7bp), or wild-type GmbZIP125 5’ UTR (WT), operably linked to a polynucleotide encoding firefly luciferase.

[0065] FIG. 9 depicts expression levels of firefly luciferase in soybean protoplasts that have been transfected with a construct containing GmbZIP123 5’ UTR with a mutated SIRT element (183- 16bp, 183-1 Ibp, 183-7bp, 207-10bp), or wild-type GmbZIP123 5’ UTR (WT), operably linked to a polynucleotide encoding firefly luciferase.

[0066] FIG. 10 depicts expression levels of firefly luciferase in soybean protoplasts that have been transfected with a construct containing GmbZIP125 5’ UTR with a mutated SIRT element (207- 19bp, 207-7bp, 207-5bp, 207-10bp), or wild-type GmbZIP125 5’ UTR (WT), operably linked to a polynucleotide encoding firefly luciferase.

[0067] FIG. 11 depicts expression levels of firefly luciferase in soybean protoplasts that have been transfected with a construct containing GmbZIP175’ UTR with a mutated SIRT element (73-5bp, 73-7bpA, 73-7bpB, 73-25bp), or wild-type GmbZIP17 5’ UTR (WT), operably linked to a polynucleotide encoding firefly luciferase. In FIGs. 7-11, the luciferase expression levels indicate the translation initiation activity of the bZIP promoter / 5’ UTR.FIG. 12 depicts oil content (%) as measured by NIR in seeds from T1 soybean plants containing (i) GmbZIP 123 5’ UTR with a mutated SIRT element (183-10 bp); (ii) GmbZIP123 5’ UTR with a mutated SIRT element (183-16 bp) and GmbZIP125 5’ UTR with a mutated SIRT element (207-19 bp); or (iii) no SIRT mutation (null).

[0068] FIG. 13 depicts sucrose content (%) as measured by NIR in soybean plants containing (i) GmbZIP 123 5’ UTR with a mutated SIRT element (183-10 bp); (ii) GmbZIP 123 5’ UTR with a mutated SIRT element (183-16 bp) and GmbZIP125 5’ UTR with a mutated SIRT element (207-19 bp); or (iii) no SIRT mutation (null).

[0069] DETAILED DESCRIPTION OF THE INVENTION

[0070] The present disclosure now will be described more fully hereinafter. The disclosure may be embodied in many different forms and should not be construed as limited to the aspects set forth herein; rather, these aspects are provided so that this disclosure will satisfy applicable legal requirements.

[0071] I. Definitions

[0072] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.

[0073] As used herein, “a,” “an,” or “the” can mean one or more than one. For example, “a” cell can mean a single cell or a multiplicity of cells. Further, the term “a plant” may include a plurality of plants.

[0074] As used herein, unless specifically indicated otherwise, the word “or” is used in the inclusive sense of “and / or” and not the exclusive sense of “either / or.”

[0075] The term “about” or “approximately” usually means within 5%, or more preferably within 1%, of a given value or range. The terms “comprises”, “comprising”, “includes”, “including”, “having” and their conjugates mean “including but not limited to”.

[0076] Various embodiments of this disclosure may be presented in a range format. It should be noted that whenever a value or range of values of a parameter are recited, it is intended that values and ranges intermediate to the recited values are also part of this disclosure. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the disclosure. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range. For example, description of a range such as from 1- 10 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 1 to 6, from 1 to 7, from 1 to 8, from 1 to 9, from 2 to 4, from 2 to 6, from 2 to 8, from 2 to 10, from 3 to 6, etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10. This applies regardless of the breadth of the range.

[0077] Whenever a numerical range is indicated herein, it is meant to include any cited numeral (fractional or integral) within the indicated range. The phrases “ranging / ranges between” a first indicate number and a second indicate number and “ranging / ranges from” a first indicate number “to” a second indicate number are used herein interchangeably and are meant to include the first and second indicated numbers and all the fractional and integral numerals there between. The recitation of a numerical range for a variable is intended to convey that the present disclosure may be practiced with the variable equal to any of the values within that range. Thus, for a variable which is inherently discrete, the variable can be equal to any integer value within the numerical range, including the end-points of the range. Similarly, for a variable which is inherently continuous, the variable can be equal to any real value within the numerical range, including the end-points of the range. As an example, and without limitation, a variable which is described as having values between 0 and 2 can take the values 0, 1 or 2 if the variable is inherently discrete, and can take the values 0.0, 0.1, 0.01, 0.001, or any other real values =0 and =2 if the variable is inherently continuous.

[0078] A plant refers to a whole plant, any part thereof, or a cell or tissue culture derived from a plant, comprising any of: whole plants, plant components or organs (e.g., leaves, stems, roots, embryos, pollen, ovules, seeds, leaves, flowers, branches, fruit, pulp, juice, kernels, ears, cobs, husks, stalks, root tips, anthers, etc.), plant tissues, seeds, plant cells, protoplasts and / or progeny of the same. A plant cell is a biological cell of a plant, taken from a plant or derived through culture of a cell taken from a plant. By grain is intended the mature seed produced by commercial growers for purposes other than growing or reproducing the species. Progeny, variants, and mutants of the regenerated plants are also included within the scope of the invention.

[0079] As used herein, a “subject plant or plant cell” is one in which genetic alteration, such as a mutation, has been effected as to a gene of interest, or is a plant or plant cell which is descended from a plant or cell so altered and which comprises the alteration. As used herein, the term “mutated” or “genetically modified” or “transgenic” or “transformed” or “edited” plants, plant cells, plant tissues, plant parts or seeds refers plants, plant cells, plant tissues, plant parts or seeds that have been mutated by the methods of the present disclosure to include one or more mutations (e.g., insertions, substitutions, or deletions) in the genomic sequence.

[0080] As used herein, a “control plant” or “control plant part” or “control cell” or “control seed” refers to a plant or plant part or plant cell or seed that has not been subject to the methods and compositions described herein. A “control” or “control plant” or “control plant part” or “control cell” or “control seed” provides a reference point for measuring changes in phenotype of the subject plant or plant cell. A control plant or plant cell may comprise, for example: (a) a wild-type plant or cell, i.e., of the same genotype as the starting material for the genetic alteration which resulted in the subject plant or cell; (b) a plant or plant cell of the same genotype as the starting material but which has been transformed with a null construct (i.e. with a construct which has no known effect on the trait of interest, such as a construct comprising a marker gene); (c) a plant or plant cell which is a non-transformed segregant among progeny of a subject plant or plant cell; (d) a plant or plant cell genetically identical to the subject plant, or plant cell but which is not exposed to conditions or stimuli (e.g., sucrose) that would induce expression of the gene of interest; or (e) the subject plant or plant cell itself, under conditions in which the gene of interest is not expressed. In certain instances, a control plant of the present disclosure is grown under the same environmental conditions (e.g., same or similar temperature, humidity, air quality, soil quality, water quality, and / or pH conditions) as a subject plant described herein. Similarly, control fatty acid / oil or sugar content, or control fatty acid / oil or sugar composition can refer to fatty acid / oil or sugar content, or fatty acid / oil or sugar composition that is isolated or derived from a control plant. In specific embodiments, a control plant, plant part, or plant cell is a plant cell that does not have a mutated nucleotide sequence encoding a SIRT element.

[0081] Plant cells possess nuclear, plastid, and mitochondrial genomes. Accordingly, by “chromosome” or “chromosomal” is intended the nuclear, plastid, or mitochondrial genomic DNA. “Genome” as it applies to plant cells encompasses not only chromosomal DNA found within the nucleus, but organelle DNA found within subcellular components (e.g., mitochondria or plastids) of the cell. The compositions and methods disclosed herein are not limited to mutations made in the genomic DNA of the plant nucleus, but may be used to modify the sequence of the nuclear, plastid, and / or mitochondrial genome, or to modulate the expression of a gene or genes encoded by the nuclear, plastid, and / or mitochondrial genome. In certain embodiments, a mutation is created in the genomic DNA of an organelle (e.g. a plastid and / or a mitochondrion). In certain embodiments, a mutation is created in extrachromosomal nucleic acids (including RNA) of the plant, cell, or organelle of a plant. Nonlimiting examples include creating mutations in supernumerary chromosomes (e.g. B chromosomes), plasmids, and / or vector constructs used to deliver nucleic acids to a plant. It is anticipated that new nucleic acid forms will be developed and yet fall within the scope of the claimed invention when used with the teachings described herein.

[0082] As used herein, the term “gene” or “coding sequence”, herein used interchangeably, refers to a functional nucleic acid unit encoding a protein, polypeptide, or peptide. As will be understood by those in the art, this functional term includes genomic sequences, cDNA sequences, and smaller engineered gene segments that express, or may be adapted to express proteins, polypeptides, domains, peptides, fusion proteins, and mutants. A gene may include a regulatory region, e.g., a promoter region or a 5 ’untranslated region, that regulates transcription or translation of the encoded gene. For example, a “bZIP transcription factor gene” includes the coding region of the bZIP transcription factor gene, and may also include the regulatory region (e.g., promoter, 5’ UTR, uORF, SIRT element) of the bZIP transcription factor gene. Further, a “bZIP transcription factor gene” as used herein includes a homolog of a known a bZIP transcription factor gene.

[0083] As used herein, the term a “nucleic acid”, used interchangeably with a “nucleotide”, refers to a molecule consisting of a nucleoside and a phosphate that serves as a component of DNA or RNA. For instance, nucleic acids include adenine, guanine, cytosine, uracil, and thymine.

[0084] As used herein, a “mutation” is any change in a nucleic acid sequence. Nonlimiting examples comprise insertions, deletions, duplications, substitutions, inversions, and translocations of any nucleic acid sequence, regardless of how the mutation is brought about and regardless of how or whether the mutation alters the functions or interactions of the nucleic acid. For example and without limitation, a mutation may produce altered enzymatic activity of a ribozyme, altered base pairing between nucleic acids (e.g. RNA interference interactions, DNA-RNA binding, etc.), altered mRNA folding stability, and / or how a nucleic acid interacts with polypeptides (e.g. DNA- transcription factor interactions, RNA-ribosome interactions, gRNA-endonuclease reactions, etc.). A mutation might result in the production of proteins with altered amino acid sequences (e.g. missense mutations, nonsense mutations, frameshift mutations, etc.) and / or the production of proteins with the same amino acid sequence (e.g. silent mutations). Certain synonymous mutations may create no observed change in the plant while others that encode for an identical protein sequence nevertheless result in an altered plant phenotype (e.g. due to codon usage bias, altered secondary protein structures, etc.). Mutations may occur within coding regions (e.g., open reading frames) or outside of coding regions (e.g., within promoters, terminators, untranslated elements, or enhancers), and may affect, for example and without limitation, gene expression levels, gene expression profiles, protein sequences, and / or sequences encoding RNA elements such as tRNAs, ribozymes, ribosome components, and microRNAs.

[0085] Accordingly, “plant with mutation” or “plant part with mutation” or “plant cell with mutation” or “plant genome with mutation” refers to a plant or plant part or plant cell or plant genome that contains a mutation (e.g., an insertion, a substitution, or a deletion) described in the present disclosure, such as a mutated nucleotide sequence encoding a SIRT element and / or a mutated SIRT element (e g., bZIP123 SIRT, bZIP125 SIRT, bZIP124 SIRT, bZIP126 SIRT, bZIP18 SIRT, bZIP17 SIRT). For example, as used herein, a plant, plant part or plant cell with mutation may refer to a plant, plant part or plant cell in which, or in an ancestor of which, a SIRT element has been deliberately mutated such that the plant, plant part or plant cell expresses a mutated (e.g., truncated, having one or more insertions, substitutions, or deletions) SIRT peptide. The mutated SIRT peptide can have altered function, e.g., reduced function or loss-of-function, compared to a wild-type, or control, SIRT peptide comprising no mutation.

[0086] “Genome editing” or “gene editing” as used herein refers to a type of genetic engineering by which one or more mutations (e.g., insertions, substitutions, deletions, modifications) are introduced at a specific location of the genome.

[0087] As used herein, the terms “nuclease” and “endonuclease” are used interchangeably to refer to naturally-occurring or engineered enzymes, which cleave a phosphodiester bond within a polynucleotide chain.

[0088] As used herein, the term “recombinant DNA construct,” “recombinant construct,” “expression cassette,” “expression construct,” “chimeric construct,” “construct,” and “recombinant DNA fragment” are used interchangeably herein and are single or double-stranded polynucleotides. A recombinant construct comprises an artificial combination of nucleic acid fragments, including, without limitation, regulatory and coding sequences that are not found together in nature. For example, a recombinant DNA construct may comprise regulatory sequences and coding sequences that are derived from different sources, or regulatory sequences and coding sequences derived from the same source and arranged in a manner different than that found in nature. Such a construct may be used by itself or may be used in conjunction with a vector.

[0089] An expression construct can permit transcription of a particular polynucleic acid sequence in a host cell (e.g., a bacterial cell or a plant cell). An expression cassette may be part of a plasmid, viral genome, or nucleic acid fragment. Typically, an expression cassette includes a polynucleotide to be transcribed, operably linked to a promoter. “Operably linked” is intended to mean a functional linkage between two or more elements. For example, an operable linkage between a promoter of the present invention and a heterologous nucleotide is a functional link that allows for expression of the heterologous nucleic acid molecule. Operably linked elements may be contiguous or noncontiguous. When used to refer to the joining of two protein coding regions, by operably linked is intended that the coding regions are in the same reading frame. The cassette may additionally contain at least one additional gene to be co-transformed into the plant. Alternatively, the additional gene(s) can be provided on multiple expression cassettes or DNA constructs. The expression cassette may additionally contain selectable marker genes. Other elements that may be present in an expression cassette include those that enhance transcription (e.g., enhancers) and terminate transcription (e.g., terminators), as well as those that confer certain binding affinity or antigenicity to the recombinant protein produced from the expression cassette.

[0090] As used herein, “function” of a gene, a peptide, a protein, or a molecule refers to activity of a gene, a peptide, a protein, or a molecule.

[0091] “Introduced” in the context of inserting a nucleic acid molecule (e.g., a recombinant DNA construct) into a cell, means “transfection” or “transformation” or “transduction” and includes reference to the incorporation of a nucleic acid fragment into a plant cell where the nucleic acid fragment may be incorporated into the genome of the cell (e.g., nuclear chromosome, plasmid, plastid chromosome or mitochondrial chromosome), converted into an autonomous replicon, or transiently expressed (e.g., transfected mRNA).

[0092] As used herein with respect to a parameter, the term “increased” or “increasing” or “increase” refers to a detectable (e.g., at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 100%, 120%, 150%, 200%, 300%, 400%, 500%, or more) positive change in the parameter from a comparison control, e.g., an established normal or reference level of the parameter, or an established standard control. Accordingly, the terms “increased”, “increase”, and the like encompass both a partial increase and a significant increase compared to a control.

[0093] As used herein with respect to a parameter, the term “decreased” or “decreasing” or “decrease” or “reduced” or “reducing” or “reduce” or “lower” refers to a detectable (e.g., at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100%) negative change in the parameter from a comparison control, e.g., an established normal or reference level of the parameter, or an established standard control. Accordingly, the terms “decreased”, “reduced”, and the like encompass both a partial reduction and a complete reduction compared to a control.

[0094] When reference is made to particular sequence listings, such reference is to be understood to also encompass sequences that substantially correspond to its complementary sequence as including minor sequence variations, resulting from, e.g., sequencing errors, cloning errors, or other alterations resulting 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.

[0095] As used herein, the term “polypeptide” refers to a linear organic polymer containing a large number of amino-acid residues bonded together by peptide bonds in a chain, forming part of (or the whole of) a protein molecule. The amino acid sequence of the polypeptide refers to the linear consecutive arrangement of the amino acids comprising the polypeptide, or a portion thereof.

[0096] As used herein the term “polynucleotide” refers to a single or double stranded nucleic acid sequence which is isolated and provided in the form of an RNA sequence (e.g., an mRNA sequence), a complementary polynucleic acid sequence (cDNA), a genomic polynucleic acid sequence and / or a composite polynucleic acid sequences (e.g., a combination of the above).

[0097] The term “isolated” refers to at least partially separated from the natural environment e.g., from a plant cell.

[0098] As used herein, the term “expression” or “expressing” refers to the transcription and / or translation of a particular nucleic acid sequence driven by a promoter.

[0099] As used herein, the terms “exogenous” or “heterologous” in reference to a nucleic acid sequence or amino acid sequence are intended to mean a sequence that is purely synthetic, that originates from a foreign species, or, if from the same species, is substantially modified from its native form in composition and / or genomic locus by deliberate human intervention. Thus, a heterologous nucleic acid sequence may not be naturally expressed within the plant (e.g., a nucleic acid sequence from a different species) or may have altered expression when compared to the corresponding wild type plant. An exogenous polynucleotide may be introduced into the plant in a stable or transient manner, 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 which is identical or partially homologous to an endogenous nucleic acid sequence of the plant. As used herein, the term “endogenous” in reference to a gene or nucleic acid sequence or protein is intended to mean a gene or nucleic acid sequence or protein that is naturally comprised within or expressed by a cell. Endogenous genes can include genes that naturally occur in the cell of a plant, but that have been modified in the genome of the cell without insertion or replacement of a heterologous gene that is from another plant species or another location within the genome of the modified cell.

[0100] As used herein, “fertilization” and / or “crossing” broadly includes bringing the genomes of gametes together to form zygotes but also broadly may include pollination, syngamy, fecundation and other processes related to sexual reproduction. Typically, a cross and / or fertilization occurs after pollen is transferred from one flower to another, but those of ordinary skill in the art will understand that plant breeders can leverage their understanding of fertilization and the overlapping steps of crossing, pollination, syngamy, and fecundation to circumvent certain steps of the plant life cycle and yet achieve equivalent outcomes, for example, a plant or cell of a soybean cultivar described herein. In certain embodiments, a user of this innovation can generate a plant of the claimed invention by removing a genome from its host gamete cell before syngamy and inserting it into the nucleus of another cell. While this variation avoids the unnecessary steps of pollination and syngamy and produces a cell that may not satisfy certain definitions of a zygote, the process falls within the definition of fertilization and / or crossing as used herein when performed in conjunction with these teachings. In certain embodiments, the gametes are not different cell types (i.e. egg vs. sperm), but rather the same type and techniques are used to effect the combination of their genomes into a regenerable cell. Other embodiments of fertilization and / or crossing include circumstances where the gametes originate from the same parent plant, i.e. a “self’ or “self-fertilization”. While selfing a plant does not require the transfer pollen from one plant to another, those of skill in the art will recognize that it nevertheless serves as an example of a cross, just as it serves as a type of fertilization. Thus, methods and compositions taught herein are not limited to certain techniques or steps that must be performed to create a plant or an offspring plant of the claimed invention, but rather include broadly any method that is substantially the same and / or results in compositions of the claimed invention.

[0101] “Homolog” or “homologous sequence” may refer to both orthologous and paralogous sequences. Paralogous sequence relates to gene-duplications within the genome of a species. Orthologous sequence relates 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 given two species and therefore have great likelihood of having the same function. One option to identify homologs (e.g., orthologs) in monocot plant species is by performing a reciprocal BLAST search. This may be done by a first blast involving blasting the sequence-of-interest against any sequence database, such as the publicly available NCBI database which may be found at: ncbi.nlm.nih.gov. If orthologs in rice were sought, the sequence-of-interest would be blasted against, for example, the 28,469 full-length cDNA clones from Oryza sativa Nipponbare available at NCBI. The blast results may be filtered. The full-length sequences of either the filtered results or the non-filtered results are then blasted back (second blast) against the sequences of the organism from which the sequence-of-interest is derived. The results of the first and second blasts are then compared. An ortholog is identified when the sequence resulting in the highest score (best hit) in the first blast identifies in the second blast the query sequence (the original sequence-of-interest) as the best hit. Using the same rational a paralog (homolog to a gene in the same organism) is found. In case of large sequence families, the ClustalW program may be used [ebi.ac.uk / Tools / clustalw2 / index.html], followed by a neighbor-joining tree (wikipedia.org / wiki / Neighbor-joining) which helps visualizing the clustering.

[0102] In some embodiments, the term “homolog” as used herein, refers to functional homologs of genes. A functional homolog is a gene encoding a polypeptide that has sequence similarity to a polypeptide encoded by a reference gene, and the polypeptide encoded by the homolog carries out one or more of the biochemical or physiological function(s) of the polypeptide encoded by the reference gene. In general, it is preferred that functional homologs and / or polypeptides encoded by functional homologs share at least some degree of sequence identity with the reference gene or polypeptide encoded by the reference gene.

[0103] Homology (e.g., percent homology, sequence identity+sequence similarity) can be determined using any homology comparison software computing a pairwise sequence alignment.

[0104] As used herein, “sequence identity,” “identity,” “percent identity,” “percentage similarity,” “sequence similarity” and the like refer to a measure of the degree of similarity of two sequences based upon an alignment of the sequences that maximizes similarity between aligned amino acid residues or nucleotides, and which is a function of the number of identical or similar residues or nucleotides, the number of total residues or nucleotides, and the presence and length of gaps in the sequence alignment. A variety of algorithms and computer programs are available for determining sequence similarity using standard parameters. As used herein, sequence similarity is measured using the BLASTp program for amino acid sequences and the BLASTn program for nucleic acid sequences, both of which are available through the National Center for Biotechnology Information, and are described in, for example, Altschul et al. (1990), J. Mol. Biol. 215:403-410; Gish and States (1993), Nature Genet. 3:266-272; Madden et al. (1996), Meth. EnzymoL 266: 131-141; Altschul et al. (1997), Nucleic Acids Res. 25:3389-3402); Zhang et al. (2000), J. Comput. Biol. 7(l-2):203-14. As used herein, percent similarity of two amino acid sequences is the score based upon the following parameters for the BLASTp algorithm: word size=3; gap opening penalty=-l 1; gap extension penalty=-l; and scoring matrix=BLOSUM62. As used herein, percent similarity of two nucleic acid sequences is the score based upon the following parameters for the BLASTn algorithm: word size=l l; gap opening penalty=-5; gap extension penalty=-2; match reward=l; and mismatch penalty=-3. When percentage of sequence identity is used in reference to proteins it is recognized that residue positions which are not identical often differ by conservative amino acid substitutions, where amino acid residues are substituted for other amino acid residues with similar chemical properties (e.g. charge or hydrophobicity) and therefore do not change the functional properties of the molecule. Where sequences differ in conservative substitutions, the percent sequence identity may be adjusted upwards to correct for the conservative nature of the substitution. Sequences which differ by such conservative substitutions are considered to have “sequence similarity” or “similarity”. Means for making this adjustment are well-known to those of skill in the art. Typically this involves scoring a conservative substitution as a partial rather than a full mismatch, thereby increasing the percentage sequence identity. Thus, for example, where an identical amino acid is given a score of 1 and a non-conservative substitution is given a score of zero, a conservative substitution is given a score between zero and 1. The scoring of conservative substitutions is calculated, e.g., according to the algorithm of Henikoff S and Henikoff J G. (Proc Natl Acad Sci 89: 10915-9 (1992)). Identity (e.g., percent homology) can be determined using any homology comparison software, including for example, the BlastN software of the National Center of Biotechnology Information (NCBI) such as by using default parameters.

[0105] According to some embodiments, the identity is a global identity, i.e., an identity over the entire amino acid or nucleic acid sequences of the invention and not over portions thereof.

[0106] According to some embodiments, the term “homology” or “homologous” refers to identity of two or more nucleic acid sequences; or identity of two or more amino acid sequences; or the identity of an amino acid sequence to one or more nucleic acid sequence. According to some embodiments, the homology is a global homology, e.g., a homology over the entire amino acid or nucleic acid sequences of the invention and not over portions thereof. The degree of homology or identity between two or more sequences can be determined using various known sequence comparison tools which are described in WO2014 / 102774.

[0107] As used herein, the term “method” refers to manners, means, techniques and procedures for accomplishing a given task including, but not limited to, those manners, means, techniques and procedures either known to, or readily developed from known manners, means, techniques and procedures by practitioners of the chemical, pharmacological, biological, biochemical and medical arts.

[0108] As used herein, the term “population” refers to a set comprising any number, including one, of individuals, objects, or data from which samples are taken for evaluation, e.g., estimating quantitative trait locus (QTL) effects and / or disease tolerance. Most commonly, the terms relate to a breeding population of plants from which members are selected and crossed to produce progeny in a breeding program. A population of plants can include the progeny of a single breeding cross or a plurality of breeding crosses and can be either actual plants or plant derived material, or in silico representations of plants. The member of a population need not be identical to the population members selected for use in subsequent cycles of analyses, nor does it need to be identical to those population members ultimately selected to obtain a final progeny of plants. Often, a plant population is derived from a single biparental cross but can also derive from two or more crosses between the same or different parents. Although a population of plants can comprise any number of individuals, those of skill in the art will recognize that plant breeders commonly use population sizes ranging from one or two hundred individuals to several thousand, and that the highest performing 5-20% of a population is what is commonly selected to be used in subsequent crosses in order to improve the performance of subsequent generations of the population in a plant breeding program.

[0109] As used herein, the term “crop performance” is used synonymously with “plant performance” and refers to of how well a plant grows under a set of environmental conditions and cultivation practices. Crop performance can be measured by any metric a user associates with a crop’s productivity (e.g., yield), appearance and / or robustness (e.g., color, morphology, height, biomass, maturation rate, etc.), product quality (e.g., fiber lint percent, fiber quality, seed protein content, seed carbohydrate content, etc.), cost of goods sold (e.g., the cost of creating a seed, plant, or plant product in a commercial, research, or industrial setting) and / or a plant’s tolerance to disease (e.g., a response associated with deliberate or spontaneous infection by a pathogen) and / or environmental stress (e.g., drought, flooding, low nitrogen or other soil nutrients, wind, hail, temperature, day length, etc.). Crop performance can also be measured by determining a crop’s commercial value and / or by determining the likelihood that a particular inbred, hybrid, or variety will become a commercial product, and / or by determining the likelihood that the offspring of an inbred, hybrid, or variety will become a commercial product. Crop performance can be a quantity (e.g., the volume or weight of seed or other plant product measured in liters or grams) or some other metric assigned to some aspect of a plant that can be represented on a scale (e.g., assigning a 1-10 value to a plant based on its disease tolerance). A “microbe” will be understood to be a microorganism, i.e. a microscopic organism, which can be single celled or multicellular. Microorganisms are very diverse and include all the bacteria, archaea, protozoa, fungi, and algae, especially cells of plant pathogens and / or plant symbionts. Certain animals are also considered microbes, e.g. rotifers. In various embodiments, a microbe can be any of several different microscopic stages of a plant or animal. Microbes also include viruses, viroids, and prions, especially those which are pathogens or symbionts to crop plants. A “pathogen” as used herein refers to a microbe that causes disease or harmful effects on plant health.

[0110] A “fungus” includes any cell or tissue derived from a fungus, for example whole fungus, fungus components, organs, spores, hyphae, mycelium, and / or progeny of the same. A fungus cell is a biological cell of a fungus, taken from a fungus or derived through culture of a cell taken from a fungus.

[0111] A “pest” is any organism that can affect the performance of a plant in an undesirable way. Common pests include microbes, animals (e.g. insects and other herbivores), and / or plants (e.g. weeds). Thus, a pesticide is any substance that reduces the survivability and / or reproduction of a pest, e.g. fungicides, bactericides, insecticides, herbicides, and other toxins.

[0112] “Tolerance” or “improved tolerance” in a plant to disease conditions (e.g. growing in the presence of a pest) will be understood to mean an indication that the plant is less affected by the presence of pests and / or disease conditions with respect to yield, survivability and / or other relevant agronomic measures, compared to a less tolerant, more “susceptible” plant. Tolerance is a relative term, indicating that a “tolerant” plant survives and / or performs better in the presence of pests and / or disease conditions compared to other (less tolerant) plants (e.g., a different soybean cultivar) grown in similar circumstances. As used in the art, “tolerance” is sometimes used interchangeably with “resistance”, although resistance is sometimes used to indicate that a plant appears maximally tolerant to, or unaffected by, the presence of disease conditions. Plant breeders of ordinary skill in the art will appreciate that plant tolerance levels vary widely, often representing a spectrum of more-tolerant or less-tolerant phenotypes, and are thus trained to determine the relative tolerance of different plants, plant lines or plant families and recognize the phenotypic gradations of tolerance.

[0113] “Yield” as used herein is defined as the measurable produce of economic value from a crop. This may be defined in terms of quantity and / or quality. Yield is directly dependent on several factors, for example, the number and size of the organs, plant architecture (for example, the number of branches), seed production, leaf senescence and more. Root development, nutrient uptake, stress tolerance, photosynthetic carbon assimilation rates, and early vigor may also be important factors in determining yield. Optimizing the abovementioned factors may therefore contribute to increasing crop yield. Yield can be measured and expressed by any means known in the art. In specific embodiments, yield is measured by seed weight or volume in a given harvest area.

[0114] A plant (e.g., a plant provided herein having increased bZIP activity and / or increased fatty acid / oil or sugar (e.g., sucrose) content), or its environment, can be contacted with a wide variety of “agriculture treatment agents.” As used herein, an “agriculture treatment agent”, or “treatment agent”, or “agent” can refer to any exogenously provided compound that can be brought into contact with a plant tissue (e.g. a seed) or its environment that affects a plant’s growth, development and / or performance, including agents that affect other organisms in the plant’s environment when those effects subsequently alter a plant’s performance, growth, and / or development (e.g. an insecticide that kills plant pathogens in the plant’s environment, thereby improving the ability of the plant to tolerate the insect’s presence, an herbicide that kills weeds in the plant’s environment, thereby improving the ability of the plant to grow). Agriculture treatment agents also include a broad range of chemicals and / or biological substances that are applied to seeds, in which case they are commonly referred to as seed treatments and / or seed dressings. Seed treatments are commonly applied as either a dry formulation or a wet slurry or liquid formulation prior to planting and, as used herein, generally include any agriculture treatment agent including growth regulators, micronutrients, nitrogen-fixing microbes, and / or inoculants. Agriculture treatment agents include pesticides (e.g. fungicides, insecticides, bactericides, etc.) hormones (abscisic acids, auxins, cytokinins, gibberellins, etc.) herbicides (e.g. glyphosate, atrazine, 2,4-D, dicamba, etc.), nutrients (e.g. a plant fertilizer), and / or a broad range of biological agents, for example a seed treatment inoculant comprising a microbe that improves crop performance, e.g. by promoting germination and / or root development. In certain embodiments, the agriculture treatment agent acts extracellularly within the plant tissue, such as interacting with receptors on the outer cell surface. In some embodiments, the agriculture treatment agent enters cells within the plant tissue. In certain embodiments, the agriculture treatment agent remains on the surface of the plant and / or the soil near the plant. In certain embodiments, the agriculture treatment agent is contained within a liquid. Such liquids include, but are not limited to, solutions, suspensions, emulsions, and colloidal dispersions. In some embodiments, liquids described herein will be of an aqueous nature. However, in various embodiments, such aqueous liquids that comprise water can also comprise water insoluble components, can comprise an insoluble component that is made soluble in water by addition of a surfactant, or can comprise any combination of soluble components and surfactants. In certain embodiments, the application of the agriculture treatment agent is controlled by encapsulating the agent within a coating, or capsule (e.g. microencapsulation). In certain embodiments, the agriculture treatment agent comprises a nanoparticle and / or the application of the agriculture treatment agent comprises the use of nanotechnology.

[0115] In certain embodiments, plants disclosed herein can be modified to exhibit at least one desired trait, and / or combinations thereof. The disclosed innovations are not limited to any set of traits that can be considered desirable, but nonlimiting examples include high protein content, male sterility, herbicide tolerance, pest tolerance, disease tolerance, modified fatty acid metabolism, modified carbohydrate metabolism, modified seed yield, modified seed oil, modified seed protein, modified lodging resistance, modified shattering, modified iron-deficiency chlorosis, modified water use efficiency, and / or combinations thereof. Desired traits can also include traits that are deleterious to plant performance, for example, when a researcher desires that a plant exhibits such a trait in order to study its effects on plant performance.

[0116] In certain embodiments, a user can combine the teachings herein with high-density molecular marker profiles spanning substantially the entire genome of the plant (e.g., soybean, pea) to estimate the value of selecting certain candidates in a breeding program in a process commonly known as genomic selection.

[0117] The patent and scientific literature referred to herein establishes knowledge that is available to those of skill in the art. The issued US patents, allowed applications, published foreign applications, and references, including GenBank database sequences, which are cited herein are hereby incorporated by reference to the same extent as if each was specifically and individually indicated to be incorporated by reference.

[0118] All publications, patent applications, patents, and other references mentioned herein are incorporated by reference herein in their entirety.

[0119] II. Overview of the Invention

[0120] Increased oil / fatty acid content or increased sugar (e.g., sucrose) content in plants or plant parts (e.g., seeds), without a negative impact on plant agronomics or acid / volatile levels, is an advantageous trait in the growing markets of food and industrial use. Increasing the activity of a basic region leucine zipper (bZIP) transcription factor gene can be one approach to generate advantageous traits, such as increased fatty acids / oil content or increased sugar (e.g., sucrose) content. The terms “basic leucine zipper transcription factor” and “bZIP,” as used herein, also refer to naturally occurring DNA sequence variations of the bZIP transcription factor gene.

[0121] Basic region leucine zipper (bZIP) transcription factor serves a vital role as a signaling molecule to regulate gene expression and pathways, including those involved in carbohydrate transport and / or metabolism in plants. bZIP transcription factor genes of various plants commonly have a conserved sucrose-sensitive sequence upstream open reading frame (uORF), termed “sucrose-induced repression of translation (SIRT)”, that can be translated into a polypeptide and represses the downstream main ORF. The bZIP transcription factor gene is normally under regulation by the upstream SIRT element through feedback inhibition by sucrose accumulation. “Upstream open reading frame” or “uORF” as used herein refers to an open reading frame (a span of DNA or mRNA between the start and stop codons) encoded within the 5’ untranslated region (5’ UTR) of a gene or an mRNA. An uORF can be a regulator of translation in plants. A “5’ untranslated region” or “5’ UTR” refers to a genetic region upstream (5’ side) of a coding sequence of DNA or mRNA, and includes translation regulatory elements such as uORFs, internal ribosome entry sites, microRNA binding sites, and structural components involved in the regulation of mRNA stability, pre-mRNA splicing, and translation initiation.

[0122] Table 1 depicts a multiple sequence alignment of the exemplary amino acid sequences of bZIP SIRT elements from Glycine max (GmbZIP 123, GmbZIP125, GmbZIP124, GmbZIP126, GmbZIP 18, and GmbZIP 17), Arabidopsis thaliana (AtbZIPl, AtbZIP125, AtbZIPl 1, AtbZIP44, AtbZIP53), Solanum lycopersicum (bZIPl, bZIP125), and Capsicum annuum (CabZIPl, CabZIP 125) showing high conservation of the C-terminal end of the SIRT amino acid sequence. Alignment was completed using the MUSCLE align program on the Molecular Evolutionary Genetics Analysis-X (MEGA-X) software. Conserved amino acids are denoted by a in the top row.

[0123] TABLE 1. Alignment of the bZIP SIRT amino acid sequences from multiple species In soybean plants (Glycine max'), several bZIP family transcription factor homologs have been identified, including GmbZIP123, GmbZIP 125, GmbZIP 124, GmbZIP 126, GmbZIP 18, and GmbZIP 17. bZIP123, bZIP125, bZIP124, bZIP126, bZIP18, and bZIP17 are all expressed throughout the soybean tissue, including high expression in the soybean seed pod and seed coat. bZIP17 is also highly expressed in the soybean nodule. As used herein, a bZIP123 transcription factor refers to the bZIP123 transcription factor encoded by the nucleic acid sequence set forth in SEQ ID NO: 1, or a fragment or variant thereof that encodes a functional bZIP123 transcription factor. Variants of the bZIP123 transcription factor retain bZIP123 activity and are encoded by a nucleic acid molecule with a sequence having at least 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 1. As used herein, a bZIP125 transcription factor refers to the bZIP125 transcription factor encoded by the nucleic acid sequence set forth in SEQ ID NO: 2, or a fragment or variant thereof that encodes a functional bZIP125 transcription factor. Variants of the bZIP125 transcription factor retain bZIP125 activity and are encoded by a nucleic acid molecule with a sequence having at least 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 2. As used herein, a bZIP124 transcription factor refers to the bZIP124 transcription factor encoded by the nucleic acid sequence set forth in SEQ ID NO: 3, or a fragment or variant thereof that encodes a functional bZIP124 transcription factor. Variants of the bZIP124 transcription factor retain bZIP124 activity and are encoded by a nucleic acid molecule with a sequence having at least 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 3. As used herein, a bZIP126 transcription factor refers to the bZIP126 transcription factor encoded by the nucleic acid sequence set forth in SEQ ID NO: 4, or a fragment or variant thereof that encodes a functional bZIP126 transcription factor. Variants of the bZIP126 transcription factor retain bZIP126 activity and are encoded by a nucleic acid molecule with a sequence having at least 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 4. As used herein, a bZIP18 transcription factor refers to the bZIP18 transcription factor encoded by the nucleic acid sequence set forth in SEQ ID NO: 5, or a fragment or variant thereof that encodes a functional bZIP18 transcription factor. Variants of the bZIP18 transcription factor retain bZIP18 activity and are encoded by a nucleic acid molecule with a sequence having at least 75%, 1 A>, n° / 0, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 5. As used herein, a bZIP17 transcription factor refers to the bZIP17 transcription factor encoded by the nucleic acid sequence set forth in SEQ ID NO: 6, or a fragment or variant thereof that encodes a functional bZIP17 transcription factor. Variants of the bZIP17 transcription factor retain bZIP17 activity and are encoded by a nucleic acid molecule with a sequence having at least 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 6. Transgenic expression of bZIP123 in Arabidopsis can enhance expression of genes involved in carbohydrate transport or metabolism, including sucrose transporters SUC1 and SUC5, cell wall invertases cwINVl, cwINV3, and cwINV6, asparagine synthetase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, and trehalose-6-phosphate phosphatase (TPP6); increase sugar translocation to siliques; and increase fatty acid content (including palmitic acid (C16:0), oleic acid (C18: l), linoleic acid (C18:2), and linolenic acid (Cl 8:3)) and oil content in seeds.

[0124] Soybean bZIP can contain an upstream SIRT element in its 5’ UTR. In specific embodiments, the bZIP123 SIRT element is encoded by a nucleic acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 13, or comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 19. In specific embodiments, the bZIP125 SIRT element is encoded by a nucleic acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 14, or comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 20. In specific embodiments, the bZIP124 SIRT element is encoded by a nucleic acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 15, or comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 21. In specific embodiments, the bZIP126 SIRT element is encoded by a nucleic acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 16, or comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 22. In specific embodiments, the bZIP18 SIRT element is encoded by a nucleic acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 17, or comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 23. In specific embodiments, the bZIP17 SIRT element is encoded by a nucleic acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the nucleic acid sequence set forth in SEQ ID NO: 18, or comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 24. Mutations in the SIRT element of a bZIP transcription factor gene, e.g. bZIP123, bZIP125, bZIP124, bZIP126, bZIP18, bZIP17, or a homolog thereof could alter (e.g., increase) the bZIP activity, alter (e.g., increase) activity of carbohydrate transport / metabolism genes, alter (e.g., increase) carbohydrate transport into seeds or siliques, and alter (e.g., increase) fatty acid / oil and / or sugar content in the plant or plant part (e.g., seeds). Novel gene traits with one or more bZIP SIRT mutations that increase fatty acid, oil, and / or sugar (e.g., sucrose) content could offer new plant varieties with added commercial value or improved consumer acceptance.

[0125] III. Plants with Increased Fatty Acid / Oil and / or Sugar Content

[0126] Disclosed herein are plants or plant parts comprising altered level or activity of a basic region / leucine zipper motif (bZIP) transcription factor gene or a homolog thereof. “bZIP transcription factor level” or “bZIP level” as used herein refers to the expression level, either in mRNA or protein, of the bZIP transcription factor. “bZIP transcription factor activity” or “bZIP activity” as used herein refers to the activity of the bZIP transcription factor to regulate expression of downstream target genes, such as genes involved in carbohydrate transport or metabolism, such as sugar transporter, sucrose transporter, cell wall invertase, asparagine synthetase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, and trehalose-6-phosphate phosphatase (TPP6).

[0127] In particular aspects, plants and plant parts (e.g., seeds) disclosed herein have a genetic mutation that alters (e.g., increases) the bZIP (e.g., bZIP123, bZIP125, bZIP124, bZIP126, bZIP18, bZIP 17) transcription factor activity. The plants or plant parts provided herein can have one or more insertions, substitutions, or deletions in a 5’ untranslated region (5’ UTR) of a bZIP transcription factor gene or a homolog thereof, wherein the 5’ UTR comprises an upstream open reading frame (uORF) with a sucrose induced repression of translation (SIRT) element. The one or more insertions, substitutions, or deletions can alter activity of the bZIP SIRT element (i.e., bZIP SIRT activity). “bZIP SIRT activity” as used herein refers to activity of the SIRT element of bZIP to regulate (e.g., suppress) bZIP transcription factor activity in the presence or absence or sucrose. For example, the mutated SIRT element may have reduced activity to regulate (e.g., reduce) the expression of a bZIP transcription factor in the presence or absence of sucrose. The plants or plant parts provided herein with decreased bZIP SIRT activity and / pr increased bZIP transcription factor activity can have increased activity of molecules regulated by bZIP (e.g., genes involved in carbohydrate transport or metabolism), and / or increased fatty acid, oil, and / or sugar (e.g., sucrose) content relative to a control plant or plant part (e.g., without mutation).

[0128] Also provided herein is a population of plants and plant parts comprising the plants and plant parts described herein having altered (e.g., increased) bZIP transcription factor level or activity. In such population of plants or plant parts, having altered bZIP transcription factor level or activity relative to a control population, not all individual plants or plant parts need to have altered (e.g., reduced) bZIP transcription factor level or activity, genetic mutation that cause altered (e.g., reduced) bZIP transcription factor level or activity, or phenotypes caused by the altered (e.g., reduced) activity of the bZIP transcription factor (e.g., bZIP123 transcription factor, bZIP125 transcription factor, bZIP 124 transcription factor, bZIP 126 transcription factor, bZIP 18 transcription factor, bZIP17 transcription factor). In specific embodiments at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more plants within a given plant population have a mutation that alters the bZIP transcription factor level or activity (e.g., a mutation in the bZIP SIRT element).

[0129] The teachings herein are not limited to certain plant species, and it is envisioned that they can be modified to be useful for monocots, dicots, and / or substantially any crop and / or valuable plant type, including plants that can reproduce by self-fertilization and / or cross fertilization, hybrids, inbreds, varieties, and / or cultivars thereof. A plant or plant part of the present disclosure can be a legume, i.e., a plant belonging to the family Fabaceae (or Leguminosae), or a part (e.g., fruit or seed) of such a plant. When used as a dry grain, the seed of a legume is also called a pulse. Examples of legume include, without limitation, soybean (Glycine max), beans (Phaseolus spp., Vigna spp.), common bean (Phaseolus vulgaris), mung bean (Vigna radiata), cowpea (Vigna unguiculata), adzuki bean (Vigna angularis), fava bean (Vicia faba), pea (Pisum sativum), chickpea (Cicer arietinum), peanut (Arachis hypogaea), lentils (Lens culinaris, Lens esculenta), lupins (Lupinus spp.), white lupin (Lupinus albus), mesquite (Prosopis spp.), carob (Ceratonia siliqua), tamarind (Tamarindus indica), alfalfa (Medicago sativa), barrel medic (Medicago truncatula), birdsfood trefoil (Lotus japonicus), licorice (Glycyrrhiza glabra), and clover (Trifolium spp.). For example, a plant or plant part of the present disclosure can be Glycine max or a part of Glycine max. Additionally, a plant or plant part of the present disclosure can be a crop plant or part of a crop plant, including legumes. Examples of crop plants include, but are not limited to, corn (Zea mays), Brassica sp. (e.g., B. napus, B. rapa, B.juncea), particularly those Brassica species useful as sources of seed oil, alfalfa (Medicago saliva), rice (Oryza saliva), rye (Secale cereale), sorghum Sorghum bicolor, Sorghum vulgar e), camelina (Camelina sativa), millet (e.g., pearl millet (Pennisetum glaucum), proso millet (Panicum miliaceum), foxtail millet (Setaria italica), finger millet Eleusine coracana)), sunflower (Helianthus annuus), quinoa (Chenopodium quinoa), chicory (Cichorium intybus), lettuce (Lactuca sativa), safflower (Carthamus tinctorius), wheat (Triticum aestivum), soybean (Glycine max), tobacco (Nicotiana spp., e.g., Nicotiana tabacum, Nicotiana sylvestris), potato (Solanum tuberosum), tomato (Solanum lycopersicum), peanuts (Arachis hypogaea), cotton (Gossypium barbadense, Gossypium hirsutum), sweet potato (Ipomoea batatus), cassava (Manihot esculenta), coffee (Coffea spp.), coconut (Cocos nucifera), pineapple (Ananas comosus), citrus trees (Citrus spp.), cocoa (Theobroma cacao), tea (Camellia sinensis), banana (Musa spp.), avocado (Persea americana), fig (Ficus casica), guava (Psidium guajava), mango (Mangifera indica), grapes (Vitis vinifera, Vitis riparia), olive (Olea europaea), papaya (Carica papaya), cashew (Anacardium occidentale), macadamia (Macadamia integrifolia), almond (Prunus amygdalus), sugar beets (Beta vulgaris), sugarcane (Saccharum spp.), oil palm (Elaeis guineensis), poplar (Populus spp.), pea (Pisum sativum), eucalyptus (Eucalyptus spp.), oats (Avena sativa), barley (Hordeum vulgare), vegetables, ornamentals, and conifers. Additionally, a plant or plant part of the present disclosure can be an oilseed plant (e.g., canola (Brassica napus), cotton (Gossypium sp.), camelina (Camelina sativa) and sunflower (Helianthus sp.)), or other species including wheat (Triticum sp., such as Triticum aestivum L. ssp. Aestivum (common or bread wheat), other subspecies of Triticum aestivum, Triticum turgidum L. ssp. Durum (durum wheat, also known as macaroni or hard wheat), Triticum monococcum L. ssp. Monococcum (cultivated einkorn or small spelt), Triticum timopheevi ssp. Timopheevi, Triticum turgigum L. ssp. Dicoccon (cultivated emmer), and other subspecies of Triticum turgidum (Feldman)), barley (Hordeum vulgare), maize (Zea mays), oats (Avena sativa), or hemp (Cannabis sativa). Additionally, a plant or plant part of the present disclosure can be a forage plant or part of a forage plant. Examples of forage plants include legumes and crop plants described herein as well as grass forages including Agrostis spp., Lolium spp., Festuca spp., Poa spp., n Bromus spp.

[0130] A. Plants with Altered Level or Activity of bZIP Transcription Factor

[0131] Provided herein are plants or plant parts (e.g., seeds) comprising altered (e.g., increased) activity of a bZIP transcription factor (e.g., bZIP123 transcription factor, bZIP125 transcription factor, bZIP 124 transcription factor, bZIP126 transcription factor, bZIP18 transcription factor, bZIP17 transcription factor) compared to a control plant or plant part. Also provided herein is a population of plants or plant parts (e.g., seeds) comprising altered (e.g., increased) activity of bZIP transcription factor compared to a control population provided herein.

[0132] The genetic mutation that alters (e.g., decreases) the bZIP transcription factor activity in the plants and plant parts provided herein can comprise one or more insertions, substitutions, or deletions in at least one native bZIP transcription factor gene or homolog thereof, or in a regulatory region of at least one native bZIP transcription factor gene or homolog thereof. The genetic mutation that alters (e.g., increases) the bZIP transcription factor activity can be located in at least one native bZIP transcription factor gene or homolog thereof; in a regulatory region of the native bZIP transcription factor gene or homolog thereof; a coding region, a non-coding region, or a regulatory region of any other gene; or at any other site in the genome of the plant or plant part. A bZIP transcription factor “gene”, as used herein, refers to any polynucleotide that encodes a polypeptide having bZIP transcription factor activity. In some embodiments, a bZIP transcription factor gene is bZIP123, bZIP125, bZIP124, bZIP126, bZIP18, or bZIP17. A bZIP transcription factor gene, as used herein, can refer to a polynucleotide including a regulatory region (e.g., promoter, 5’ UTR) of the bZIP transcription factor gene. A bZIP transcription factor gene can also include a homolog, ortholog, or variant, that retains bZIP transcription factor activity, of a known a bZIP transcription factor gene.

[0133] A “native” gene or nucleic acid sequence, as used herein, refers to any gene or nucleic acid sequence having a wild-type nucleic acid sequence, e.g., a nucleic acid sequence that can be found in the genome of a plant existing in nature, and need not naturally occur within the plant, plant part, or plant cell comprising such native gene. For example, a transgenic bZIP transcription factor gene (e.g., bZIP123, bZIP125, bZIP124, bZIP126, bZIP18, bZIP17) or a transgenic bZIP SIRT element located at a genomic site or in a plant in a non-naturally occurring matter is a “native” bZIP transcription factor gene or a “native” bZIP SIRT element if its nucleic acid sequence can be found in a plant existing in nature.

[0134] A “regulatory region” of a gene as used herein refers to a genomic site that modulate transcription or translation of the gene, e.g., where a RNA polymerase, a transcription factor, or other transcription or translation modulators bind, or where a regulatory structure or complex is formed, and include a promoter region, 5’ UTR, a binding site for transcription modulator proteins (e.g., transcription factors), and other genomic regions that contribute to regulation of transcription or translation of the gene. A regulatory region of the gene can be located in the 5’ region from the coding region of the gene. For example, one or more insertions, substitutions, and / or deletions can be introduced at least partially into a promoter region, 5’ UTR, a binding site (e.g., an enhancer sequence) for a transcription modulator protein (e.g., transcription factor), or other genomic regions that contribute to regulation of transcription or translation of at least one (e.g., one, more than one but not all, or all) bZIP transcription factor gene, to confer to the plant or plant part an increased bZIP transcription factor activity. In specific embodiments, a regulatory region of a bZIP transcription factor includes the SIRT element.

[0135] A control plant or plant part can be a plant or plant part to which a mutation provided herein has not been introduced, e.g., by methods of the present disclosure. Thus, a control plant or plant part (e.g., seeds, leaves) may express a native (e.g., wild-type) bZIP transcription factor gene or bZIP SIRT element endogenously or transgenically. A control plant of the present disclosure may be grown under the same environmental conditions (e.g., same or similar temperature, humidity, air quality, soil quality, water quality, and / or pH conditions) as a plant with the mutation described herein. A plant, plant part (e.g., seeds, leaves), or a population of plants or plant parts of the present disclosure may have altered (e.g., decreased) bZIP SIRT activity, altered (e.g., increased) bZIP transcription factor (e.g., bZIP123 transcription factor, bZIP125 transcription factor, bZIP124 transcription factor, bZIP 126 transcription factor, bZIP 18 transcription factor, bZIP 17 transcription factor) level or activity, and / or altered (e.g., increased) fatty acid, oil, and / or sugar content as compared to a control plant, plant part, or population, when the plant, plant part, or population of plants or plant parts of the present disclosure is grown under the same environmental conditions as the control plant or plant part.

[0136] 1. Plants with Mutation in Regulatory Region of bZIP Transcription Factor

[0137] In some aspects, the present disclosure provides plants or plant parts thereof, including fruits and seeds comprising one or more mutations (e.g., insertions, substitutions, or deletions) in a 5’ UTR of a bZIP transcription factor gene or a homolog thereof, wherein the 5’ UTR comprises an uORF, and wherein the uORF comprises a SIRT element. The one or more mutations can increase the bZIP transcription factor activity and / or increase carbohydrate (e.g., sugar transport) in the plant or plant part relative to a control plant or plant part. In some aspects, the plants or plant parts have one or more mutations in the uORF of bZIP coding sequence. For example, the plants or plant parts have one or more mutations in the SIRT element of the uORF of a bZIP coding sequence. In particular, described herein are plants or plant parts, in which an upstream regulatory region of the bZIP transcription factor gene (e.g., the bZIP SIRT element) has been mutated, e.g., by one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) insertions, substitutions, or deletions in the genome of cells or parts of the plants. For example, disclosed herein are plants or plant parts in which a gene encoding the SIRT peptide has been mutated, e.g., by one or more insertions, substitutions, or deletions in the SIRT coding sequence. In some embodiments, the mutation is located at least partially in a 5’ end region of said SIRT element.

[0138] In some embodiments, the mutation in the plants or plant parts of the present disclosure may comprise one or more insertions, substitutions, or deletions in a nucleotide region corresponding to the 5’ UTR of the bZIP transcription factor gene. For example, the plants or plant parts may comprise one or more insertions, substitutions, or deletions of about 1-20, 2-20, 3-20, 4-20, 5-20, 6- 20, 7-20, 8-20, 9-20, 10-20, or 11-20 (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) of bZIP 5’ UTR in the genome of a plant cell or plant part.

[0139] In some embodiments, the plants or plant parts of the present disclosure may comprise one or more insertions, substitutions, or deletions in a nucleotide region corresponding to the uORF of the bZIP transcription factor gene. For example, the plants or plant parts may comprise one or more insertions, substitutions, or deletions of about 1-20, 2-20, 3-20, 4-20, 5-20, 6-20, 7-20, 8-20, 9-20, 10-20, or 11-20 (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) nucleotides of bZIP uORF in the genome of a plant cell or plant part.

[0140] In some embodiments, the plants or plant parts of the present disclosure may comprise one or more insertions, substitutions, or deletions in a nucleotide region corresponding to the SIRT element of the bZIP transcription factor gene. For example, the plants or plant parts may comprise one or more insertions, substitutions, or deletions of about 1-20, 2-20, 3-20, 4-20, 5-20, 6-20, 7-20, 8-20, 9-20, 10-20, or 11-20 (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) nucleotides of bZIP SIRT element in the genome of a plant cell or plant part.

[0141] In specific embodiments, the mutation is located in an uORF of a bZIP transcription factor gene or homolog thereof that: (i) comprises a nucleic acid sequence having at least 80% (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to any one of SEQ ID NOs: 1-6, wherein said nucleic acid sequence encodes a polypeptide that retains bZIP transcription factor activity; (ii) comprises the nucleic acid sequence of any one of SEQ ID NOs: 1-6; (iii) encodes a polypeptide comprising an amino acid sequence having at least 80% (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to an amino acid sequence of any one of SEQ ID NOs: 7-12, wherein said polypeptide retains bZIP transcription factor activity; or (iv) encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 7-12. For example, the mutation can be located in an uORF of bZIP123, bZIP125, bZIP124, bZIP126, bZIP18, or bZIP17, e.g., GmbZIP123 (Glyma.06G010200), GmbZIP 125 (Glyma.04G010300), GmbZIP124 (Glyma.12G040600), GmbZIP 126 (Glyma.11G114800), GmbZIP 18 (Glyma.14G071400), or GmbZIP 17 (Glyma.17G253200). In some embodiments, the mutation is located at least partially in a SIRT element in the uORF of the bZIP transcription factor gene. As used herein, where an insertion, a substitution, or a deletion is “at least partially” in a certain nucleotide region, the whole part of the insertion, substitution, or deletion can be within the certain nucleotide region, or alternatively, can span across the certain nucleotide region and a region outside the nucleotide region. The SIRT element can (i) comprise a nucleic acid sequence having at least 90% (e.g., 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to any one of SEQ ID NOs: 13-18, wherein said nucleic acid sequence encodes a polypeptide that retains SIRT activity; (ii) comprise the nucleic acid sequence of any one of SEQ ID NOs: 13-18; (iii) encode a polypeptide comprising an amino acid sequence having at least 90% (e.g., 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to an amino acid sequence of any one of SEQ ID NOs: 19- 24, wherein said polypeptide retains SIRT activity; or (iv) encode a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 19-24, before the mutation is located. The 5’ UTR of the bZIP that contains the uORF can (v) comprise a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 48-53; or (vi) comprise the nucleic acid sequence of any one of SEQ ID NOs: 48-53, before the mutation is located. In specific embodiments, the plant or plant part comprises a deletion of one or more nucleotides of: SEQ ID NO: 13 in the SIRT element in the uORF of the Glycine max bZIP 123 transcription factor gene; SEQ ID NO: 14 in the SIRT element in the uORF of the Glycine max bZIP 125 transcription factor gene; SEQ ID NO: 15 in the SIRT element in the uORF of the Glycine max bZIP 124 transcription factor gene; SEQ ID NO: 16 in the SIRT element in the uORF of the Glycine max bZIP126 transcription factor gene; SEQ ID NO: 17 in the SIRT element in the uORF of the Glycine max bZIP 18 transcription factor gene; Sor EQ ID NO: 18 in the SIRT element in the uORF of the Glycine max bZIP17 transcription factor gene.

[0142] A plant or plant part provided herein can contain a mutation that is a deletion of 7-25 nucleotides located at least partially in the SIRT element of a bZIP transcription factor gene. The plant or plant part can contain a mutation at least partially in the SIRT element of two or more copies of a bZIP transcription factor gene. For example, a plant or plant part can contain a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 30-34 and 54-56, a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 35-39 and 57-58, and / or a mutated 5’ UTR of the Glycine max bZIP17 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 59-62. In specific embodiments, the plant or plant part contains (i) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 32, or (ii) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NO: 54 and a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 57. In some embodiments, the one or more mutations (e.g., insertions, substitutions, or deletions) comprise an out-of-frame mutation of the SIRT element. In some embodiments, the one or more insertions, substitutions, or deletions comprise an in-frame mutation, nonsense mutation, or missense mutation of the SIRT element.

[0143] A plant or plant part described herein can comprise 1-6, 2-4, 3-4, 2-5, or 3-5 (e.g., 1, 2, 3, 4, 5, or 6) copies of the bZIP transcription factor and / or the bZIP SIRT element. In particular, a plant or plant part described herein can comprise at least 2 genes encoding a bZIP transcription factor and / or a bZIP SIRT peptide, such as 2 genes that have less than 100% (e.g., less than 100%, 99%, 98%, 97%, 96%, 95%, 94%, 93%, 92%, 91%, 90%, 89%, 88%, 87%, 86%, or 85%) sequence identity to each other. In some embodiments, a plant or a plant part comprises two or more of a bZIP 123 transcription factor, a bZIP 125 transcription factor, a bZIP 124 transcription factor, a bZIP 126 transcription factor, a bZIP 18 transcription factor, and a bZIP 17 transcription factor, each of which comprise a SIRT element in its uORF that has less than 100% identity to each other.

[0144] Also disclosed are variants and fragments of sequences (e.g., bZIP transcription factor, bZIP SIRT) of the present disclosure. Such sequences include sequences that are orthologs of the disclosed sequences. By “Orthologs” is intended genes derived from a common ancestral gene and which are found in different species as a result of speciation. Genes found in different species are considered orthologs when their nucleic acid sequences and / or their encoded protein sequences share at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or greater sequence identity. Functions of orthologs are often highly conserved among species. Thus, isolated polynucleotides that have transcription activation or enhancer activities and which share at least 75% sequence identity to the sequences disclosed herein, or to variants or fragments thereof, are encompassed by the present disclosure. For example, orthologs of bZIP include, but not limited to Capsicum annuum bZIPl, Capsicum annuum bZIP2, Solanum lycopersicum bZIPl, Solanum lycopersicum bZIP2, Solanum melongena bZIPl, and Solanum melongena bZIP2, Arabidopsis thaliana bZIPl, Arabidopsis thaliana bZIP2, Arabidopsis thaliana bZIPl 1, Arabidopsis thaliana bZIP44, and Arabidopsis thaliana bZIP53. These bZIP orthologs, SIRT elements, and plants and plant parts comprising these bZIP orthologs and / or SIRT elements are encompassed by the present disclosure. Variant sequences can be isolated by PCR. Methods for designing PCR primers and PCR cloning are generally known in the art and are disclosed in Sambrook et al. (1989) Molecular Cloning: A Laboratory Manual (2d ed., Cold Spring Harbor Laboratory Press, Plainview, New York). See also Innis et al., eds. (1990) PCR Protocols: A Guide to Methods and Applications (Academic Press, New York); Innis and Gelfand, eds. (1995) PCR Strategies (Academic Press, New York); and Innis and Gelfand, eds. (1999) PCR Methods Manual (Academic Press, New York). Variant sequences may also be identified by analysis of existing databases of sequenced genomes. In this manner, variant sequences encoding, for instance, bZIP or bZIP SIRT element can be identified and used in the methods of the present disclosure. The variant sequences will retain the biological activity.

[0145] A bZIP 5’ UTR, a bZIP uORF, or a bZIP SIRT element of the present disclosure with one or more mutations, or fragment thereof, may be found in plants or plant parts (e.g., juice, pulp, seed, fruit, flowers, nectar, embryos, pollen, ovules, leaves, stems, branches, bark, kernels, ears, cobs, husks, stalks, roots, root tips, anthers, etc.) to which one or more mutations have been introduced by the methods of the present disclosure. A mutated bZIP SIRT element can also be found in plant products, e.g., plant extract (e.g., sweetener, antioxidants, alkaloids, etc.), plant concentrate (e.g., whole plant concentrate or plant part concentrate), plant powder (e.g., formulated powder, such as formulated plant part powder (e.g., seed flour)), and plant biomass (e.g., dried biomass, such as crushed and / or powdered biomass) obtained from such plants or plant parts.

[0146] In certain instances, mutations in any bZIP gene or its 5’ UTR, uORF, or SIRT element in a plant, plant part, population of plants or plant parts, or plant product (e.g., plant protein composition) can be identified by a detection method described herein. Such detection methods may comprise use of primers for detecting a mutation in the bZIP 5’ UTR, uORF, or SIRT region. For example, the forward primer CGCTCGATCATCTTG (SEQ ID NO: 40) and the reverse primer ACAAGCCATAGTCATAG (SEQ ID NO: 41) can be used for detection of mutation in the GmbZIP123 SIRT region near the binding site of GmbZIP 123 gRNA182 or GmbZIP123 gRNA183. The forward primer GTAGACCAACCTCCT (SEQ ID NO: 42) and the reverse primer CCAGACACTAAACAAGAA (SEQ ID NO: 43) can be used for detection of mutation in the GmbZIP 125 SIRT region at or near the binding site of GmbZIP 125 gRNA206 or GmbZIP 125 gRNA207. The forward primer CCATTTTTCAGCACCA (SEQ ID NO: 44) and the reverse primer AGTACAACCAATAGAGGA (SEQ ID NO: 45) can be used for detection of mutation in the GmbZIP124 or GmbZIP126 SIRT region at or near the binding site of GmbZIP124 gRNA97 or GmbZIP 126 gRNA156. The forward primer TTAGCAAACCCCTGC (SEQ ID NO: 46) and the reverse primer CAATTACACCAAAGTGAG (SEQ ID NO: 47) can be used for detection of mutation in the GmbZIP17 or GmbZIP18 SIRT regions at or near the binding site of GmbZIP18 gRNA91 or GmbZIPl 7 gRNA73.

[0147] In certain instances, a kit comprising a set of primers can be used for detecting mutation of bZIP genes in plants, plant parts, or plant product (e.g., plant protein composition). For example, a kit comprising the forward primer CGCTCGATCATCTTG (SEQ ID NO: 40) and the reverse primer ACAAGCCATAGTCATAG (SEQ ID NO: 41) can be used for detection of mutation in the GmbZIP123 SIRT region in plants, plant parts, or plant products (e.g., plant seed, protein, oil, or sugar compositions). A kit comprising the forward primer GTAGACCAACCTCCT (SEQ ID NO: 42) and the reverse primer CCAGACACTAAACAAGAA (SEQ ID NO: 43) can be used for detection of mutation in the GmbZIP123 SIRT region in plants, plant parts, or plant products. A kit comprising the forward primer CCATTTTTCAGCACCA (SEQ ID NO: 44) and the reverse primer AGTACAACCAATAGAGGA (SEQ ID NO: 45) can be used for detection of mutation in the GmbZIP126 SIRT region in plants, plant parts, or plant products. A kit comprising the forward primer TTAGCAAACCCCTGC (SEQ ID NO: 46) and the reverse primer CAATTACACCAAAGTGAG (SEQ ID NO: 47) can be used for detection of mutation in the GmbZIP17 SIRT region in plants, plant parts, or plant products.

[0148] In some embodiments, the one or more mutations are integrated into the plant genome and the plant or the plant part is stably transformed. In other embodiments, the one or more mutations are not integrated into the plant genome and wherein the plant or the plant part is transiently transformed.

[0149] 2. Plants with Reduced bZIP SIRT Activity and Increased bZIP Activity

[0150] The plants, plant parts (e.g., seeds, leaves), or plant products (e.g., seed composition, plant protein composition) of the present disclosure (e.g., comprising one or more mutations in the upstream regulatory region of the bZIP, e.g., bZIP SIRT element) can comprise decreased bZIP SIRT activity or loss of function / reduced function of bZIP SIRT as compared to a control (e.g., wild-type) plant, plant part, or plant product. Also provided herein is a population of plants or plant parts (e.g., seeds) comprising the plants and plant parts of the present disclosure, which has decreased bZIP SIRT activity or loss of function / reduced function of bZIP SIRT compared to a control population of plants or plant parts. The term “SIRT activity” or “SIRT function” as used herein refers to the ability of the SIRT element to regulate a downstream ORF (e.g., bZIP), such as to repress a downstream ORF (e.g., bZIP) in the presence or absence of sucrose. In specific embodiments a downstream ORF is in-frame with the SIRT element. The downstream ORF can be immediately downstream of the SIRT element coding sequence or can have intervening sequences, such as an intervening gene or regulatory region between the SIRT coding sequence and the downstream ORF. In some instances, “SIRT activity” or “SIRT function” may also refer to the ability of the SIRT element to regulate level or activity of molecules downstream of bZIP, e.g., molecules involved in carbohydrate (e.g., sugar) transport and / or metabolism, e.g., sugar transporter, sucrose transporter, cell-wall invertase, asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, or trehalose-6-phosphate phosphatase (TPP6). Accordingly, “reduced function” or “loss of function” of the SIRT element may refer to reduced ability or loss of ability of the SIRT element to regulate a downstream ORF (e.g., bZIP), such as to repress a downstream ORF (e.g., bZIP) in the presence or absence of sucrose; or reduced ability or loss of ability of the SIRT element to regulate level or activity of molecules downstream of bZIP, e.g., molecules involved in carbohydrate (e.g., sugar) transport and / or metabolism as described above. A mutated upstream regulatory region of bZIP (e.g., bZIP SIRT) in a plant or plant part, or introducing mutations to the upstream regulatory region of bZIP (e.g., bZIP SIRT) in a plant or plant part may cause reduced or loss of level, activity, or function of the SIRT element. In particular, level, activity, or function of the SIRT element in a plant, plant part, and / or plant product of the present disclosure may be reduced by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80- 100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, or 70-90% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or 90-100%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, as compared to a control plant, plant part, and / or plant product. Expression level of SIRT element can be measured by any means known in the art for measuring peptide production, such as measuring mRNA levels or protein levels. Function (i.e., activity) of SIRT can be determined by measuring the resultant expression of the corresponding downstream gene. In specific embodiments, SIRT function can be determined by measuring the activity of the downstream bZIP transcription factor gene, such as bZIP 123 transcription factor or bZIP 125 transcription factor.

[0151] Plants, plant parts (e.g., seeds, leaves), or plant products (e.g., seed composition, plant protein composition) of the present disclosure (e.g., comprising one or more mutations in the upstream regulatory region of the bZIP, e.g., bZIP SIRT element) can have altered (e.g., increased) activity or function in the bZIP transcription factor, as compared to a control plant or plant part. The SIRT element regulates the function of bZIP, including repressing the bZIP function in the presence of sucrose. Accordingly, in some embodiments, regulation by sucrose of level or activity of the bZIP transcription factor downstream of the SIRT element is reduced or eliminated (e.g., percent reduction in the presence of sucrose compared to in the absence of sucrose is reduced or eliminated, or percent increase in the presence of sucrose compared to in the absence of sucrose is increased) in a plant, plant part, and / or plant product of the present disclosure compared to a control plant, plant part, and / or plant product. In particular, regulation by sucrose of the bZIP level or activity in a plant, plant part, and / or plant product may be reduced (e.g., percent reduction in the presence of sucrose compared to in the absence of sucrose is reduced or eliminated, or percent increase in the presence of sucrose compared to in the absence of sucrose is increased) by about 10- 100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, or 70-90% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or 90-100%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, e.g., by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, as compared to a control plant, plant part, and / or plant product.

[0152] In some embodiments, level or activity of the bZIP transcription factor downstream of the SIRT element may be increased in the absence or presence of sucrose in a plant, plant part, and / or plant product as compared to a control plant, plant part, and / or plant product. In particular, level or activity of the bZIP transcription factor in the plant, plant part, and / or plant product can be increased by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80- 100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100-1000%, 200-1000%, 300- 1000%, 400-1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300-900%, 400-900%, 500-900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20- 30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100-200%, 200-300%, 300- 400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900-1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more in the absence or presence of sucrose as compared to a control plant or plant part. In specific embodiments, the level or activity of the bZIP transcription factor in the plant, plant part, and / or plant product is increased to about 150-650%, e.g., 150%-300%, 180-300%, 220- 330%, or 350-650% of that in a control plant or plant part.

[0153] Level or activity of a bZIP transcription factor in a plant, plant part, and / or plant product can be determined by one or more standard methods known in the art. For example, expression of a bZIP transcription factor can be measured by any means known in the art for measuring peptide production, such as measuring mRNA levels by RT-PCR, northern blot, and serial analysis of gene expression (SAGE); or measuring protein levels by Western blot, ELISA, or dot blot analysis. Activity of the bZIP transcription factor can be determined by measuring the level or activity of the downstream molecules, such as sugar (sucrose) transporter, cell-wall invertase, asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, or trehalose-6-phosphate phosphatase (TPP6). In some embodiments, bZIP transcription factor activity can be determined by introducing into a plant or plant part a DNA construct comprising, in operable linkage: a promoter that is functional in a plant cell; a nucleic acid sequence encoding the mutated bZIP upstream regulatory region (e.g., SIRT element); a nucleic acid sequence encoding the bZIP; and a reporter gene (e.g., GFP, an HA tag), and quantifying level or activity of the reporter gene linked to the mutated bZIP upstream regulatory region. In some embodiments, function of molecules regulated by bZIP (e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, TPP6) can be determined by assays to determine respective enzymatic activity, or by carbohydrate transport assays. Details of such procedure has been provided elsewhere in the present disclosure.

[0154] It is well understood that alteration of transcription factor expression can in turn alter the expression of the target gene(s) of said transcription factor. In some embodiments, a plant, plant part, and / or plant product of the present disclosure comprises altered level or activity of a molecule downstream of, or regulated by, the bZIP transcription factor, such that the molecule comprises, for example: (i) increased baseline level or activity in the absence of sucrose; (ii) less reduction or more increase of the level or activity in the presence of sucrose relative to the absence of sucrose; and / or (iii) increase in the level or activity in the presence of sucrose; compared to the control plant, plant part, and / or plant product. Accordingly, plants, plant parts, or plant products of the present disclosure can comprise altered level or activity of a molecule downstream of, or regulated by, the bZIP transcription factor compared to a control plant, plant part, or plant product. In some embodiments, the molecule regulated by the bZIP transcription factor regulates carbohydrate transport or metabolism in the plant or plant part. For example, the molecule regulated by the bZIP is one or more of sugar transporter, sucrose transporter, cell wall invertase, asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6 ’-phosphate phosphatase (SPP), calmodulin, trehalose-6-phosphate phosphatase (TPP6), or any other protein, peptide, carbohydrate, or enzyme that plays a role in carbohydrate metabolism or carbohydrate transport in plants. In specific embodiments, the molecule regulated by a bZIP transcription factor is fatty acid, oil, or sugar (e.g., sucrose). A “sucrose transporter” expressed in a plant or plant part regulates sucrose allocation both intracellularly and at the whole plant level by using mechanisms such as facilitated transport, energy-dependent sucrose / H+antiport, and sucrose / H+symport. In specific embodiments, the sugar transporter provided herein is encoded by one or more of Glyma.lOG217900, Glyma.02G075000, Glyma.02G218600, Glyma.l6G157100, Glyma.l6G156800, Glyma.02G124100, Glyma.01G067600, or homolog of any thereof.

[0155] A “cell wall invertase” catalyzes the breakdown of sucrose into glucose and fructose in the cell wall. A cell wall invertase is commonly co-expressed with a hexose transporter, and facilitate carbohydrate (e.g., sugar) transport intracellularly and within the plant (Proels & Huckelhoven 2014 Mol. Plant Path. 15:8;858-864). Without wishing to be bound by theory, increased activity of molecules that regulate carbohydrate transport, e.g., sugar transporter, sucrose transporter, cell wall invertase, can increase carbohydrate transport into seeds or siliques, and can increase fatty acid / oil and / or sugar (e.g., sugar) content in the seeds or siliques. In specific embodiments, the cell wall invertase provided herein is encoded by one or more of Glyma.l3G349300, Glyma.l5G024600, Glyma.l4G096600, Glyma.l6G175800, or homolog of any thereof.

[0156] In some embodiments, baseline level or activity of the molecule (e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, TPP6) in the absence of sucrose, or level or activity of the molecule in the presence of sucrose is increased in a plant, plant part, and / or plant product of the present disclosure by 10-100%, 20-100%, 30-100%, 40-100%, 50- 100%, 60-100%, 70-100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100- 1000%, 200-1000%, 300-1000%, 400-1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300-900%, 400-900%, 500-900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100- 200%, 200-300%, 300-400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900- 1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more as compared to a control plant, plant part, and / or plant product in the absence or presence, respectively, of sucrose.

[0157] In some embodiments, suppression of level or activity of the molecule (e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, TPP6) induced by sucrose (i.e., in the presence of sucrose compared to in the absence of sucrose) is reduced in a plant, plant part, and / or plant product of the present disclosure by about 10-100%, 20- 100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, or 70-90% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or 90-100%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, as compared to a control plant, plant part, and / or plant product.

[0158] In some embodiments, increase in the level or activity of the molecule (e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, TPP6) in the presence of sucrose relative to in the absence of sucrose in a plant, plant part, and / or plant product of the present disclosure is increased by 10-100%, 20-100%, 30-100%, 40-100%, 50- 100%, 60-100%, 70-100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100- 1000%, 200-1000%, 300-1000%, 400-1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300-900%, 400-900%, 500-900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100- 200%, 200-300%, 300-400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900- 1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more as compared to in the absence of sucrose (i.e., baseline).

[0159] In specific embodiments, carbohydrate (e.g., sugar) transport to seeds or siliques in a plant of the present disclosure is increased by 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60- 100%, 70-100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100-1000%, 200-1000%, 300-1000%, 400-1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200- 900%, 300-900%, 400-900%, 500-900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100- 200%, 200-300%, 300-400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900- 1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more as compared to a control plant. In the embodiments provided herein, the sugar transporter can be encoded by Glyma.10G217900, Glyma.02G075000, Glyma.02G218600, Glyma.l6G157100, Glyma.16G 156800 , Glyma.02G124100, Glyma.01G067600, or homolog of any thereof, and / or the cell wall invertase can be encoded by Glyma.13G349300, Glyma.15G024600 , Glyma.14G096600, Glyma.16G 175800, or homolog of any thereof.

[0160] Level or activity of molecules regulated by bZIP (e.g., sugar transporter, sucrose transporter, cell-wall invertase, ASN, PDH, SPS, SPP, calmodulin, and / or TPP6) in a plant, plant part, and / or plant product can be determined by one or more standard methods known in the art. For example, expression level of a molecule regulated by the bZIP transcription factor can be measured by any means known in the art for measuring levels of mRNA and / or protein expression (e.g., quantitative RT-PCR, northern blot, SAGE, western blot analysis, ELISA, dot blot analysis). Activities of the bZIP downstream targets involved in carbohydrate transport (e.g., sugar transporter, sucrose transporter, cell-wall invertase) can be measured by a standard carbohydrate (e.g., sugar) transport assay, for example by labeling the plants with13CO2 and monitoring13C enrichment levels of sucrose, fructose, and glucose in plant organs of interest, e.g., leaves, seeds, and siliques. Activity of a sugar transporter can also be measured by using a biosensor that detects conformational changes of the transporter during sugar transport. Activities of the bZIP downstream targets involved in carbohydrate (e.g., sugar) transport or metabolism (e.g., cell wall invertase, ASN, PDH, SPS, SPP, TPP6) can be measured by enzymatic assays of the respective enzymes. For example, activity of a cell wall invertase can be measured for example as described in Tomlinson et al. 2004 J. Exp. Botany 55, 2291-2303, the entire content of which is incorporated by reference herein. Activity of calmodulin can be determined by a Ca2+-calmodulin binding assay or measuring activities of Ca2+-dependent phosphodiesterase or calcium-calmodulin dependent protein kinase (CaMKII) (Bossuyt & Bers 2013 J. Mol. Med. (Berl.) 91(8):907-916).

[0161] A control plant, plant part, or population of plant or plant part disclosed herein can be a plant or plant part to which one or more mutations has not been introduced by the methods provided herein. Thus, a control plant or plant part may express wild-type (WT) SIRT element. A control plant of the present disclosure may be grown under the same environmental conditions (e.g., same or similar temperature, humidity, air quality, soil quality, water quality, and / or pH conditions) as a plant with one or more mutations described herein. A plant, plant part, and / or plant product of the present disclosure may have altered (e.g., reduced) activity of the bZIP SIRT element, altered (e.g., increased) activity of the bZIP transcription factor, and / or altered (e.g., increased) activity of the downstream molecules regulated by the bZIP, as compared to a control plant, plant part, and / or plant product, when the plant or plant part of the present disclosure is grown under the same environmental conditions as the control plant or plant part.

[0162] 3. Plants with Increased Fatty Acid / Oil Content and / or Increased Sugar Content The plant, plant part (e.g., seeds, leaves), or plant product (e.g., seed composition, plant protein composition) of the present disclosure, e.g., comprising a mutation in the bZIP SIRT element that increases the bZIP transcription factor activity, can have increased fatty acid, oil, and / or sugar (e.g., sucrose) content as compared to a control (e.g., wild-type) plant, plant part, or plant product. Also provided herein is a population of plants or plant parts (e.g., seeds) comprising the plants and plant parts of the present disclosure, which has increased fatty acid, oil, and / or sugar (e.g., sucrose) content as compared to a control population.

[0163] “Fatty acid” as used herein can include any fatty acid found in a plant, plant part, or plant oil, including palmitic acid (16:0), stearic acid (18:0), oleic acid (18: 1), linoleic acid (18:2), and linolenic acid (18:3), wherein the first and second numbers represent the number of carbons and the number of double bonds, respectively, in the fatty acid chain. Palmitic, stearic and other saturated fatty acids are typically solid at room temperature, in contrast to the unsaturated fatty acids, which remain liquid. Because saturated fatty acids have no double bonds in the acyl chain, they remain stable to oxidation at elevated temperatures. Saturated fatty acids are important components in margarines and chocolate formulations, and for many food applications, increased levels of saturated fatty acids are desired.

[0164] Oleic acid has one double bond, but is still relatively stable at high temperatures, and oils with high levels of oleic acid are suitable for cooking and other processes where heating is required. Recently, increased consumption of high oleic oils has been recommended, because oleic acid appears to lower blood levels of low density lipoproteins (“LDLs”) without affecting levels of high density lipoproteins (“HDLs”). However, some limitation of oleic acid levels is desirable, because when oleic acid is degraded at high temperatures, it creates negative flavor compounds and diminishes the positive flavors created by the oxidation of linoleic acid. Neff et al., JAOCS, 77 : 1303-1313 (2000); Warner et al., J. Agric. Food Chem. 49:899-905 (2001).

[0165] Linoleic acid is a major polyunsaturated fatty acid in foods and is an essential nutrient for humans. It is a desirable component for many food applications because it is a major precursor of fried food flavor substances such as 2,4 decadienal, which make fried foods taste good. However, linoleic acid has limited stability when heated. Preferred food oils have linoleic acid levels that are 10% or greater by weight, to enhance the formation of desirable fried food flavor substances, and also are 25% or less by weight, so that the formation of off-flavors is reduced. Linoleic acid also has cholesterol-lowering properties, although dietary excess can reduce the ability of human cells to protect themselves from oxidative damage, thereby increasing the risk of cardiovascular disease. Toborek et al., Am J. Clin. J. 75: 119-125 (2002). See generally Flavor Chemistry of Lipid Foods, editors D. B. Min & T. H. Smouse, Am Oil Chem. Soc., Champaign, Ill. (1989).

[0166] Linoleic acid, having a lower melting point than oleic acid, further contributes to improved cold flow properties desirable in biodiesel and biolubricant applications. Preferred oils for most applications have linoleic acid levels of 30% or less by weight, because the oxidation of linoleic acid limits the useful storage or use-time of frying oil, food, feed, fuel and lubricant products. See generally, Physical Properties of Fats, Oils, and Emulsifiers, ed. N. Widlak, AOCS Press (1999); Erhan & Asadauskas, Lubricant Basestocks from Vegetable Oils, Industrial Crops and Products, 11 :277-282 (2000). In addition, high linoleic acid levels in cattle feed can lead to undesirably high levels of linoleic acid in the milk of dairy cattle, and therefore poor oxidative stability and flavor. Timmons et al., J. Dairy Sci. 84:2440-2449 (2001).

[0167] Linolenic acid is also an important component of the human diet. It is used to synthesize the co-3 family of long-chain fatty acids and the prostaglandins derived therefrom. However, its double bonds are highly susceptible to oxidation, so that oils with high levels of linolenic acid deteriorate rapidly on exposure to air, especially at high temperatures. Partial hydrogenation of such oils is often necessary before they can be used in food products to retard the formation of off-flavors and rancidity when the oil is heated, but hydrogenation creates unhealthy trans fatty acids which can contribute to cardiovascular disease.

[0168] Sugar content can include the content of total sugar, or specific sugar such as sucrose, glucose, fructose, galactose, maltose, and / or lactose.

[0169] A control plant, plant part, or a population of plants or plant parts can be a plant or plant part to which one or more mutations has not be introduced by the methods provided herein, i.e., having a native bZIP SIRT element or a native uORF of the bZIP transcription factor gene; reference plants, plant parts, or population; or commodity plants, plant parts, or population). A control plant of the present disclosure may be grown under the same environmental conditions (e.g., same or similar temperature, humidity, air quality, soil quality, water quality, and / or pH conditions) as a plant with one or more mutations described herein. A plant, plant part, and / or plant product of the present disclosure may have increased levels of fatty acid and / or oil content as compared to a control plant, plant part, and / or plant product, when the plant or plant part of the present disclosure is grown under the same environmental conditions as the control plant or plant part. A reference (e.g., control) sample of soybean plant or seed, e.g., a commodity soybean or seed, can have oil content of from about 8% to about 28%, while influenced by both genotype and environmental factors (Clemente & Cahoon 2009, Plant Physiol. 151 : 1030-1040). In some embodiments, total fatty acid, oil, or sugar (e.g., sucrose) content can be increased by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80-100%, 20- 90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100-1000%, 200-1000%, 300-1000%, 400- 1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300-900%, 400-900%, 500- 900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20-30%, 30-40%, 40- 50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100-200%, 200-300%, 300-400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900-1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more in the plants or plant parts of the present disclosure as compared to a control plant or plant part. In some embodiments, total fatty acid, oil, and / or sugar (e.g., sucrose) content, as expressed by % dry weight, in the plant, plant part, or population of plant or plant parts provided herein is greater than that in control plant, plant part, or population, and the difference (by subtraction) is about 0.25- 10%, 0.5-10%, 0.75-10%, 1.0-10%, 1.5-10%, 2-10%, 2.5-10%, 3-10%, 3.5-10%, 4-10%, 4.5-10%, 5-10%, 6-10%, 7-10%, 8-10%, 9-10%, or more than 10% (e.g., by about 0.25-0.5%, 0.5-0.75%, 0.75-1.0%, 1.0-1.5%, 1.5-2.0%, 2.0-2.5%, 2.5-3.0%, 3.0-3.5%, 3.5-4.0%, 4.0-4.5%, 4.5-5.0%, 5- 6%, 6-7%, 7-8%, or 8-9%, 9-10%, or more than 10%), by about 0.25%, 0.5%, 0.75%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5%, 6%, 7%, 8%, 9%, 10%, or more, or at least 0.25%, 0.5%, 0.75%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5%, 6%, 7%, 8%, 9%, 10%, or more fatty acid, oil, and / or sugar content. In specific embodiments, total oil content as expressed by % dry weight in the plant or plant part or population of plants or plant parts provided herein is greater than that in the control plant, plant part, or population, and the difference (by subtraction) is about 0.5-2.5%, 0.8-1.8%, or 1-1.6%, e.g., about 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%. 1.1%, 1.2%, 1.3%, 1.4%, 1.5%, 1.6%, 1.7%, 1.8%, 1.9%, 2.0%, 2.1%, 2.2%, 2.3%, 2.4%, or 2.5%, e.g., 1.3% or 1.4% dry weight. Sucrose content as expressed by % dry weight in the plant or plant part or population of plants or plant parts provided herein can be greater than that in the control plant, plant part, or population, and the difference (by subtraction) can be about 0.1-1.5%, 0.1-1%, 0.4- 0.6%, e.g., about 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.1%, 1.2%, 1.3%, 1.4%, 1.5%, e.g., 0.5% dry weight.

[0170] In specific embodiments, provided herein are seeds or a population of seeds having seed fatty acid / oil and / or sugar (e.g., sucrose) content greater than control seeds or a control population of seeds (e.g., control seeds or population having a native bZIP SIRT element, reference seeds or population, commodity seeds or population). The seeds can be legume seeds, e.g., pea seeds or soybean seeds. The oil content of typical soybean cultivars average at approximately 20%, ranging from approximately 8% to approximately 28%, while influenced by both genotype and environmental factors (Clemente & Cahoon 2009, Plant Physiol. 151 : 1030-1040). In contrast, the soybean seeds or a population of soybean seeds provided herein can have seed fatty acid / oil content of at least 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more by dry weight. In specific embodiments, a reference sample of soybean plant or seed has oil content of about 20-21%, which can be increased to about 21-22% or more in the plant or plant part comprising the one or more mutations. A reference sample of soybean plant or seed can have sucrose content of about 7-8%, which can be increased to about 8-9% or more in the plant or plant part comprising the one or more mutations.

[0171] Fatty acid / oil or sugar (e.g., sucrose) content in plants, plant parts, or a population of plants or plant parts can be measured to determine if the mutations introduced to the SIRT element alters seed fatty acid / oil or sugar (e.g., sucrose) content. Amount or levels of oil, total fatty acids, and specific fatty acids can be measured by any methods for measuring oil or fatty acid amount or levels in a plant sample, including near-infrared reflectance (NIR) spectroscopy, gas chromatography-mass spectrometry (GC-MS) optionally with certain modifications (e.g., with or without initial lipid extraction, with or without isotope labeling of analytes), or nuclear magnetic resonance (NMR). Fatty acid composition (e.g., percentage of specific fatty acids normalized to total fatty acids) can be calculated based on the amount or concentration of total fatty acids and specific fatty acids in the sample. Amount or level of sucrose, total sugar, glucose, fructose, galactose, maltose, and / or lactose in seeds can be determined by any methods for measuring total or specific sugar content in a plant sample, including near-infrared reflectance (NIR) spectroscopy, solid-phase extraction (SPE), solid-phase micro-extraction (SPME), high performance liquid chromatography (HPLC), gas chromatography mass spectrometry (GCMS), and / or enzymatic assay.

[0172] In specific embodiments, the plant, plant part, or a population of plants or plant parts of the present disclosure has the trait of increased fatty acid, oil, and / or sugar content as compared to a control plant, plant part, population of plants or plant parts, or plant product, without a significant decrease in yield. In some embodiments, a reduction in yield in the plant, plant part, or population of plants or plant parts of the present disclosure, having increased fatty acid, oil, and / or sugar content, is no more than about 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, or about 5.0%, 6%, 7%, 8%, 9%, or 10%, e.g., no more than about 0-5%, 0.5-4.5%, 0.5-4%, 1-5%, 1-4%, 2- 5%, 2-4%, 0.5-10%, 0.5-8%, 1-10%, 2-10%, 3-10%, 4-10%, 5-10%, 6-10%, 7-10%, or 8-10% reduction in yield as compared to a control plant, plant part, or population of plants or plant parts. Yield can be measured and expressed by any means known in the art. In specific embodiments, yield is measured by seed weight or volume of seeds, fruits, leaves, or whole plants harvested from a given harvest area.

[0173] Amount or level of sucrose, total sugar, glucose, fructose, galactose, maltose, and / or lactose in seeds can be determined by any methods for measuring total or specific sugar content in a plant sample, including near-infrared reflectance (NIR) spectroscopy, solid-phase extraction (SPE), solid-phase micro-extraction (SPME), high performance liquid chromatography (HPLC), gas chromatography mass spectrometry (GCMS), and / or enzymatic assay.

[0174] In specific embodiments, provided herein are seeds and a population of seeds with increased bZIP transcription factor activity provided herein, having an increased fatty acid, oil, and / or sugar content as compared to control seeds or a population of seeds.

[0175] B. Plant Parts and Plant Products

[0176] The present disclosure provides plant parts and plant products obtained from the plant of the present disclosure. A “plant product”, as used herein, refers to any composition derived from the plant or plant part, including any oil products, sugar products, fiber products, protein products (such as protein concentrate, protein isolate, flake, or other protein product), seed hulls, meal, or flour, for a food, feed, aqua, or industrial product, plant extract (e.g., sweetener, antioxidants, alkaloids, etc.), plant concentrate (e.g., whole plant concentrate or plant part concentrate), plant powder (e.g., formulated powder, such as formulated plant part powder (e.g., seed flour)), plant biomass (e.g., dried biomass, such as crushed and / or powdered biomass), grains, plant protein composition, plant oil composition, and food and beverage products containing plant compositions (e.g., plant parts, plant extract, plant concentrate, plant powder, plant protein, plant oil, and plant biomass) described herein. Plant parts and plant products provided herein can be intended for human or animal consumption. In specific embodiments, provided herein are a seed composition and an oil composition produced from the plants or plant parts of the present disclosure.

[0177] Plants, seed compositions, oil compositions, and sugar compositions provided herein can be suitable for use in a variety of food and beverage products, such as plant milk, plant nut butter, plant meal, plant flour, cooking oil, salad oil, shortening, lecithin, animal feed, shakes, health drinks, alternative meat products (e.g., meatless burger patties, meatless sausages), alternative egg products (e.g., eggless mayo), non-dairy products (e.g., non-dairy whipped toppings, non-dairy milk, non-dairy creamer, non-dairy milk shakes, non-diary ice cream), energy bars, infant formula, baby foods, cereals, baked goods, edamame, tofu, and tempeh. Plants, seed compositions, and oil compositions provided herein can be suitable for use in a variety of industrial materials, such as fuel (e.g., biodiesel), soaps, candles, cosmetic products, paints, and protective coatings.

[0178] Plant parts (e.g., seeds) and plant products (e.g., plant biomass, seed compositions, protein compositions, food and / or beverage products) as disclosed herein can be meant for consumption by agricultural animals or for use as feed in an agriculture or aquaculture system. In specific embodiments, plant parts and plant products include animal feed (e.g., roughages - forage, hay, silage; concentrates - cereal grains, soybean cake) intended for consumption by bovine, porcine, poultry, lambs, goats, or any other agricultural animal. In some embodiments, plant parts and plant products include aquaculture feed for any type of fish or aquatic animal in a farmed or wild environment including, without limitation, trout, carp, catfish, salmon, tilapia, crab, lobster, shrimp, oysters, clams, mussels, and scallops.

[0179] Seeds of the present disclosure include a representative sample of seeds, from a plant of the present disclosure. A plant or plant part of the present disclosure can be a crop plant, a forage plant, or part of a crop plant or forage plant.

[0180] As provided herein, plant products (e.g., seed compositions, oil compositions, sugar compositions, and plant-based food / beverage products) of the present disclosure can be produced from plants or plant parts that contain a mutation that decreases bZIP transcription factor activity, e.g., one or more insertions, substitutions, or deletions at least partially in the SIRT element of at least one native bZIP transcription factor gene (e.g., bZIP 123, ZIP 125, bZIP 124, bZIP 126, bZIP 18, bZIP17y, reduced bZIP SIRT activity; increased bZIP transcription factor activity; increased activity or level of one or more molecules regulated by the bZIP transcription factor; increased carbohydrate transport and / or metabolism; and / or increased oil / fatty acid and / or sugar content compared to a control plant or plant part. Accordingly, the plant products (e.g., seed compositions, oil compositions, sugar composition, and plant-based food / beverage products) provided herein can contain a mutation that decreases bZIP transcription factor activity, e.g., one or more insertions, substitutions, or deletions at least partially in the SIRT element of at least one native bZIP transcription factor gene (e.g., bZIP 123, ZIP 125, bZIP 124, bZIP 126, bZIP 18, bZIP 17), reduced bZIP SIRT activity; increased bZIP transcription factor activity; increased activity or level of one or more molecules regulated by the bZIP transcription factor; increased carbohydrate transport and / or metabolism; and / or increased oil / fatty acid and / or sugar content relative to a control plant product prepared from a control (e.g., wild-type, commodity, reference) plant or plant part.

[0181] In specific embodiments, the plant products (e.g., plant seed compositions, oil compositions, sugar compositions) provided herein are obtained from a soybean plant (Glycine max) that contains a mutation that decreases bZIP transcription factor (e.g., bZIP123 transcription factor, bZIP125 transcription factor, bZIP 124 transcription factor, bZIP126, transcription factor bZIP18 transcription factor, bZIP17 transcription factor) activity, e.g., one or more insertions, substitutions, or deletions at least partially in the SIRT element of at least one native bZIP transcription factor gene (e.g., bZIP 123, ZIP 125, bZIP124, bZIP126, bZIP18, bZIP 17) or homolog thereof.

[0182] IV. Increasing Fatty Acid, Oil, and / or Sugar Content in Plants

[0183] Provided herein are methods for altering (e.g., increasing) function of a basic region / leucine zipper motif (bZIP) transcription factor gene or a homolog thereof in a plant or plant part. In some aspects, the methods comprise introducing a genetic mutation that increases the bZIP transcription factor activity into said plant or plant part. The genetic mutation that alters (e.g., decreases) the bZIP transcription factor activity in the plants and plant parts provided herein can comprise one or more insertions, substitutions, or deletions in at least one native bZIP transcription factor gene or homolog thereof, or in a regulatory region of at least one native bZIP transcription factor gene or homolog thereof. The genetic mutation that alters (e.g., increases) the bZIP transcription factor activity can be located in at least one native bZIP transcription factor gene or homolog thereof; in a regulatory region of the native bZIP transcription factor gene or homolog thereof; a coding region, a non-coding region, or a regulatory region of any other gene; or at any other site in the genome of the plant or plant part.

[0184] In specific embodiments, the methods comprise introducing a genetic mutation comprising one or more insertions, substitutions, or deletions in an upstream open reading frame (uORF) of at least one native bZIP transcription factor gene or homolog thereof, wherein the uORF comprises a sucrose induced repression of translation (SIRT) element. The mutation can alter level or activity of the SIRT element, increase level or activity of said bZIP transcription factor gene or homolog thereof, increase level or activity of a bZIP transcription factor encoded by the bZIP transcription factor gene or homolog thereof, increase carbohydrate (e.g., sugar) transport and / or metabolism in the plant or plant part, and / or increase fatty acid / oil and / or sugar (e.g., sucrose) content in the plant or plant part relative to a control plant or plant part. The method can further comprise introducing the genetic mutation that increases the bZIP transcription factor activity into a plant cell, and regenerating the plant or plant part from said plant cell. A control plant or plant part can be a plant or plant part to which a mutation provided herein has not been introduced, e.g., by methods of the present disclosure. Thus, a control plant or plant part (e.g., seeds, leaves) may express a native (e.g., wild-type) bZIP SIRT element endogenously. A control plant of the present disclosure may be grown under the same environmental conditions (e.g., same or similar temperature, humidity, air quality, soil quality, water quality, and / or pH conditions) as a plant to which the mutation is introduced according to the methods provided herein.

[0185] Also provided herein are plants, plant parts (e.g., seeds, leaves), a population of plants or plant parts, or plant product (e.g., seed composition, plant protein compositions) produced according to the methods of the present disclosure. Such plants, plant parts, a population of plants or plant parts, or plant products may have the mutation that decreases bZIP SIRT level or activity, increases bZIP transcription factor level or activity, increases level or activity of one or more target molecules regulated by the bZIP transcription factor and regulating carbohydrate transport or metabolism in the plant or plant part, increases carbohydrate transport and / or metabolism, and / or increased fatty acid / oil and / or sugar (e.g., sucrose) content, as compared to a control plant, plant part, population of plants or plant parts, when the plant, plant part, or population of plants or plant parts of the present disclosure is grown under the same environmental conditions as the control plant or plant part. In the population of plants or plant parts, having altered bZIP transcription factor level or activity relative to a control population, not all individual plants or plant parts need to have altered (e.g., reduced) bZIP transcription factor level or activity, genetic mutation that cause altered (e.g., reduced) bZIP transcription factor level or activity, or phenotypes caused by the altered (e.g., reduced) activity of the bZIP transcription factor (e.g., bZIP123 transcription factor, bZIP 125 transcription factor, bZIP 124 transcription factor, bZIP 126 transcription factor, bZIP 18 transcription factor, bZIP17 transcription factor). In specific embodiments at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more plants within a given plant population have a mutation that alters the bZIP transcription factor level or activity (e.g., a mutation in the bZIP SIRT element).

[0186] Level or activity of SIRT, bZIP, and molecules regulated by bZIP can be altered as provided below.

[0187] A. Altering bZIP Transcription Factor Level or Activity

[0188] Provided herein are compositions and methods for altering (e.g., increasing) a bZIP transcription factor (e.g., bZIP123 transcription factor, bZIP125 transcription factor) in a plant, plant part, or a population of plants or plant parts compared to a control plant, plant part, or plant population by introducing a genetic mutation that alters bZIP transcription factor activity in a plant or plant part. The genetic mutation that alters (e.g., decreases) the bZIP transcription factor activity in the plants and plant parts provided herein can comprise one or more insertions, substitutions, or deletions in at least one native bZIP transcription factor gene or homolog thereof, or in a regulatory region of at least one native bZIP transcription factor gene or homolog thereof. The genetic mutation that alters (e.g., increases) the bZIP transcription factor activity can be located in at least one native bZIP transcription factor gene or homolog thereof; in a regulatory region of the native bZIP transcription factor gene or homolog thereof; a coding region, a non-coding region, or a regulatory region of any other gene; or at any other site in the genome of the plant or plant part.

[0189] The method can further comprise introducing the genetic mutation that increases the bZIP transcription factor activity into a plant cell, and regenerating the plant or plant part from said plant cell.

[0190] 1. Introducing Mutation in Regulatory Region of bZIP Transcription Factor

[0191] The methods described herein can comprise introducing a mutation that increases bZIP transcription factor activity, e.g., one or more mutations (e.g., insertions, substitutions, or deletions) in a regulatory region, e.g., a 5’ UTR of a bZIP transcription factor gene or a homolog thereof. The 5’ UTR can comprise an uORF, and the uORF can comprise a SIRT element.

[0192] In some aspects, the methods include introducing one or more mutations in the uORF of bZIP coding sequence of the plant or plant part. For example, the method can include introducing one or more mutations in the SIRT element of the uORF of a bZIP coding sequence. In specific embodiments, the methods include introducing one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) insertions, substitutions, or deletions into an upstream regulatory region of the bZIP transcription factor gene (e.g., the bZIP SIRT element) in the genome of cells or parts of the plants. For example, the methods can include introducing a mutation, e.g., one or more insertions, substitutions, or deletions into a gene encoding the SIRT peptide, i.e., the SIRT coding sequence. In some embodiments, the methods include introducing the mutation to locate at least partially in a 5’ end region of said SIRT element.

[0193] The methods can include introducing into a plant or a plant part a mutation that comprises one or more insertions, substitutions, or deletions in a nucleotide region corresponding to the 5’ UTR of the bZIP transcription factor gene. For example, the mutation to be introduced can comprise one or more insertions, substitutions, or deletions of about 1-20, 2-20, 3-20, 4-20, 5-20, 6- 20, 7-20, 8-20, 9-20, 10-20, or 11-20 (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) of bZIP 5’ UTR in the genome of a plant cell or plant part.

[0194] The methods can include introducing into a plant or a plant part a mutation that comprises one or more insertions, substitutions, or deletions in a nucleotide region corresponding to the uORF of the bZIP transcription factor gene. For example, the plants or plant parts may comprise one or more insertions, substitutions, or deletions of about 1-20, 2-20, 3-20, 4-20, 5-20, 6-20, 7-20, 8-20, 9-20, 10-20, or 11-20 (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) nucleotides of bZIP uORF in the genome of a plant cell or plant part. In some embodiments, the plants or plant parts of the present disclosure may comprise one or more insertions, substitutions, or deletions in a nucleotide region corresponding to the SIRT element of the bZIP transcription factor gene. For example, the plants or plant parts may comprise one or more insertions, substitutions, or deletions of about 1-20, 2-20, 3-20, 4-20, 5-20, 6-20, 7-20, 8-20, 9-20, 10-20, or 11-20 (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) nucleotides of bZIP SIRT element in the genome of a plant cell or plant part.

[0195] In specific embodiments, the methods include introducing a mutation to locate in an uORF of a bZIP transcription factor gene or homolog thereof that: (i) comprises a nucleic acid sequence having at least 80% (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to any one of SEQ ID NOs: 1-6, wherein said nucleic acid sequence encodes a polypeptide that retains bZIP transcription factor activity; (ii) comprises the nucleic acid sequence of any one of SEQ ID NOs: 1-6; (iii) encodes a polypeptide comprising an amino acid sequence having at least 80% (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to an amino acid sequence of any one of SEQ ID NOs: 7-12, wherein said polypeptide retains bZIP transcription factor activity; or (iv) encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 7-12. For example, the mutation can be located in an uORF of a gene encoding bZIP 123 transcription factor, bZIP 125 transcription factor, bZIP 124 transcription factor, bZIP 126 transcription factor, bZIP 18 transcription factor, bZIP 17 transcription factor, e.g., GmbZIP123 (Glyma.06G010200), GmbZIP 125 (Glyma.04G010300), GmbZIP 124 (Glyma.12G040600), GmbZIP 126 (Glyma.11G114800), GmbZIP 18 (Glyma.14G071400), or GmbZIP 17 (Glyma.17G253200).

[0196] In some embodiments, the methods include introducing a mutation to locate at least partially in a SIRT element in the uORF of the bZIP transcription factor gene. The SIRT element can (i) comprise a nucleic acid sequence having at least 90% (e.g., 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to any one of SEQ ID NOs: 13-18, wherein said nucleic acid sequence encodes a polypeptide that retains SIRT activity; (ii) comprise the nucleic acid sequence of any one of SEQ ID NOs: 13-18; (iii) encode a polypeptide comprising an amino acid sequence having at least 90% (e.g., 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to an amino acid sequence of any one of SEQ ID NOs: 19- 24, wherein said polypeptide retains SIRT activity; or (iv) encode a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 19-24, before the mutation is introduced. The 5’ UTR of the bZIP that contains the uORF can (v) comprise a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 48-53; or (vi) comprise the nucleic acid sequence of any one of SEQ ID NOs: 48-53, before the mutation is introduced. In specific embodiments, the methods include introducing a deletion of one or more nucleotides of: SEQ ID NO: 13 in the SIRT element in the uORF of the Glycine max bZIP123 transcription factor gene; SEQ ID NO: 14 in the SIRT element in the uORF of the Glycine max bZIP125 transcription factor gene; SEQ ID NO: 15 in the SIRT element in the uORF of the Glycine max bZIP124 transcription factor gene; SEQ ID NO: 16 in the SIRT element in the uORF of the Glycine max bZIP126 transcription factor gene; SEQ ID NO: 17 in the SIRT element in the uORF of the Glycine max bZIP18 transcription factor gene; or SEQ ID NO: 18 in the SIRT element in the uORF of the Glycine max bZIP17 transcription factor gene into the plant or plant part.

[0197] The mutation to be introduced can comprise a deletion of 7-25 nucleotides located at least partially in said SIRT element. A mutation can be introduced at least partially in the SIRT element of two or more copies of a bZIP transcription factor gene. For example, the plant or plant part can contain a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 30-34 and 54-56, a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 35-39 and 57-58, and / or a mutated 5’ UTR of the Glycine max bZIP17 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 59-62. In specific embodiments, the plant or plant part contains (i) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 32, or (ii) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NO: 54 and a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 57. In some embodiments, the one or more mutations (e.g., insertions, substitutions, or deletions) introduced into a plant or plant part comprise an out-of-frame mutation of the SIRT element. In some embodiments, the one or more insertions, substitutions, or deletions comprise an in-frame mutation, nonsense mutation, or missense mutation of the SIRT element.

[0198] A plant or plant part described herein can comprise 1-6, 2-4, 3-4, 2-5, or 3-5 (e.g., 1, 2, 3, 4, 5, or 6) copies of the bZIP transcription factor and / or the bZIP SIRT element. In particular, a plant or plant part described herein can comprise at least 2 genes encoding a bZIP transcription factor and / or a bZIP SIRT peptide, such as 2 genes that have less than 100% (e.g., less than 100%, 99%, 98%, 97%, 96%, 95%, 94%, 93%, 92%, 91%, 90%, 89%, 88%, 87%, 86%, or 85%) sequence identity to each other. In some embodiments, a plant or a plant part comprises two or more of a bZIP123 transcription factor, a bZIP125 transcription factor, a bZIP 124 transcription factor, a bZIP126 transcription factor, a bZIP18 transcription factor, and a bZIP17 transcription factor, each of which comprise a SIRT element in its uORF that has less than 100% identity to each other.

[0199] The methods can include introducing a mutation into the variants and fragments of sequences (e.g., bZIP transcription factor, , bZIP uORF, bZIP SIRT) of the present disclosure. Such sequences include sequences that are orthologs of the disclosed sequences.

[0200] Variant sequences can be isolated by PCR. Variant sequences may also be identified by analysis of existing databases of sequenced genomes. In this manner, variant sequences encoding, for instance, bZIP or bZIP SIRT element can be identified and used in the methods of the present disclosure. The variant sequences will retain the biological activity.

[0201] In certain instances, mutations introduced into any bZIP genes or their 5’ UTR, uORF, or SIRT element in a plant, plant part, or plant product (e.g., plant protein composition) according to the methods provided herein can be identified by a detection method described herein. Such detection methods may comprise use of primers for detecting mutation in the bZIP 5’ UTR, uORF, or SIRT region. For example, the forward primer CGCTCGATCATCTTG (SEQ ID NO: 40) and the reverse primer ACAAGCCATAGTCAT”G (S’Q)ID NO: 41) can be used for detection of mutation in the GmbZIP123 SIRT region near the binding site of GmbZIP 123 gRNA182 or GmbZIP 123 gRNA183. The forward primer GTAGACCAACCTCCT (SEQ ID NO: 42) and the reverse primer CC AG AC AC TA AC A AG A A (SEQ ID NO: 43) can be used for detection of mutation in the GmbZIP123 SIRT region at or near the binding site of GmbZIP 125 gRNA206 or GmbZIP 125 gRNA207. The forward primer CCATTTTTCAGCACCA (SEQ ID NO: 44) and the reverse primer AGTACAACCAATAGAGGA (SEQ ID NO: 45) can be used for detection of mutation in the GmbZIP124 or GmbZIP126 SIRT region at or near the binding site of GmbZIP124 gRNA97 or GmbZIP 126 gRNA156. The forward primer TTAGCAAACCCCTGC (SEQ ID NO: 46) and the reverse primer CAATTACACCAAAGTGAG (SEQ ID NO: 47) can be used for detection of mutation in the GmbZIP 18 or GmbZIP 17 SIRT region at or near the binding site of GmbZIP 18 gRNA91 or GmbZIP 17 gRNA73.

[0202] In some embodiments, the one or more mutations are integrated into the plant genome and the plant or the plant part is stably transformed. In other embodiments, the one or more mutations are not integrated into the plant genome and wherein the plant or the plant part is transiently transformed.

[0203] The insertion, substitution, or deletion that is introduced at least partially into the regulatory region, e.g., uORF, e.g., SIRT element of a bZIP transcription factor gene can comprise insertion, substitution, or deletion of one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43,

[0204] 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69,

[0205] 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95,

[0206] 96, 97, 98, 99, 100, or more) nucleotides. The substitute can be a cisgenic substitute, a transgenic substitute, or both.

[0207] 2. Decreasing bZIP SIRT Activity or Increasing bZIP Transcription Factor Activity

[0208] The methods of the present disclosure can decrease bZIP SIRT activity or increase bZIP transcription factor activity of a in plants, plant parts (e.g., seeds, leaves), a population of plants or plant parts, or plant products (e.g., seed composition, plant protein composition) compared to a control (e.g., wild-type) plant, plant part, population of plants or plant parts, or plant product. In particular, the methods can include introducing a mutation into the upstream regulatory region of bZIP (e.g., bZIP uORF, e.g., bZIP SIRT) in a plant or plant part, and this may cause reduced or loss of level, activity, or function of the SIRT element. In particular, the methods can reduce level, activity, or function of the SIRT element in a plant, plant part, and / or plant product of the present disclosure by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80- 100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, or 70-90% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or 90-100%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, e.g., by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, as compared to a control plant, plant part, and / or plant product. Expression level of SIRT element can be measured by any means known in the art for measuring peptide production, such as measuring mRNA levels or protein levels. Function (i.e., activity) of SIRT can be determined by measuring the resultant expression of the corresponding downstream gene. In specific embodiments, SIRT function can be determined by measuring the activity of the downstream bZIP transcription factor gene, such as bZIP 123 transcription factor, bZIP 125 transcription factor, bZIP 124 transcription factor, bZIP 126 transcription factor, bZIP 18 transcription factor, or bZIP 17 transcription factor.

[0209] The methods can increase activity or function in the bZIP transcription factor, as compared to a control plant, plant part, and / or plant product. The SIRT element regulates the function of bZIP, including repressing the bZIP function in the presence of sucrose. Accordingly, in some embodiments, the methods can reduce or eliminate regulation by sucrose of level or activity of the bZIP transcription factor downstream of the SIRT element (e.g., percent reduction in the presence of sucrose compared to in the absence of sucrose is reduced or eliminated, or percent increase in the presence of sucrose compared to in the absence of sucrose is increased) in the plant, plant part, or plant product as compared to a control plant, plant part, or plant product. In particular, the methods can reduce regulation by sucrose of the bZIP level or activity in a plant, plant part, and / or plant product by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80- 100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, or 70-90% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or 90-100%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, e.g., by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, as compared to a control plant, plant part, and / or plant product.

[0210] In some embodiments, the methods can increase level or activity of the bZIP transcription factor downstream of the SIRT element in the absence or presence of sucrose in the plant, plant part, or plant product as compared to a control plant or plant part. In particular, the methods can increase level or activity of the bZIP transcription factor in the plant, plant part, and / or plant product by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80- 100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100-1000%, 200-1000%, 300- 1000%, 400-1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300-900%, 400-900%, 500-900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20- 30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100-200%, 200-300%, 300- 400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900-1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more in the absence or presence of sucrose as compared to a control plant or plant part. In specific embodiments, the methods can increase level or activity of the bZIP transcription factor in the plant, plant part, and / or plant product about 150-650%, e.g., 150%-300%, 180-300%, 220- 330%, or 350-650% of that in a control plant or plant part.

[0211] Level or activity of a bZIP transcription factor in a plant, plant part, and / or plant product can be determined by one or more standard methods known in the art. For example, expression of a bZIP transcription factor can be measured by any means known in the art for measuring peptide production, such as measuring mRNA levels by RT-PCR, northern blot, and serial analysis of gene expression (SAGE); or measuring protein levels by Western blot, ELISA, or dot blot analysis. Activity of the bZIP transcription factor can be determined by measuring the level or activity of the downstream molecules, such as sugar transporter, sucrose transporter, cell wall invertase, asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, or trehalose-6-phosphate phosphatase (TPP6). The sugar transporter can be encoded by Glyma.lOG217900, Glyma.02G075000, Glyma.02G218600, Glyma.l6G157100, Glyma.l6G156800, Glyma.02G124100, Glyma.01G067600, or homolog of any thereof. The cell wall invertase can be encoded by Glyma.l3G349300, Glyma.l5G024600, Glyma.l4G096600, Glyma.l6G175800, or homolog of any thereof. bZIP transcription factor activity can be determined by introducing into a plant or plant part a DNA construct comprising, in operable linkage: a promoter that is functional in a plant cell; a nucleic acid sequence encoding the mutated bZIP upstream regulatory region (e.g., SIRT element); a nucleic acid sequence encoding the bZIP; and a reporter gene (e.g., GFP, an HA tag), and quantifying level or activity of the reporter gene linked to the mutated bZIP upstream regulatory region. In some embodiments, function of molecules regulated by bZIP (e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, TPP6) can be determined by assays to determine respective enzymatic activity, or by carbohydrate transport assays. Details of such procedure has been provided elsewhere in the present disclosure.

[0212] It is well understood that alteration of transcription factor expression can in turn alter the expression of the target gene(s) of said transcription factor. In some embodiments, the methods can alter level or activity of a molecule downstream of, or regulated by, the bZIP transcription factor, such that the molecule comprises, for example: (i) increased baseline level or activity in the absence of sucrose; (ii) less reduction or more increase of the level or activity in the presence of sucrose relative to the absence of sucrose; and / or (iii) increase in the level or activity in the presence of sucrose; compared to the control plant, plant part, and / or plant product. Accordingly, the methods can alter level or activity of a molecule downstream of, or regulated by, the bZIP transcription factor compared to a control plant, plant part, or plant product. In some embodiments, the molecule regulated by the bZIP transcription factor regulates carbohydrate transport or metabolism. For example, the molecule regulated by the bZIP transcription factor can be one that regulate carbohydrate transport or metabolism. For example, the molecule regulated by the bZIP transcription factor can be one or more of sugar transporter, sucrose transporter, cell wall invertase, asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6 ’-phosphate phosphatase (SPP), calmodulin, trehalose-6-phosphate phosphatase (TPP6), or any other protein, peptide, carbohydrate, or enzyme that plays a role in carbohydrate metabolism or carbohydrate transport in plants. The sugar transporter can be encoded by Glyma.lOG217900, Glyma.02G075000, Glyma.02G218600, Glyma.l6G157100, Glyma.l6G156800, Glyma.02G124100, Glyma.01G067600, or homolog of any thereof. The cell wall invertase can be encoded by Glyma.l3G349300, Glyma.l5G024600, Glyma.l4G096600, Glyma.l6G175800, or homolog of any thereof. In specific embodiments, the molecule regulated by a bZIP transcription factor is fatty acid, oil, or sugar (e.g., sucrose).

[0213] In some embodiments, the methods can increase baseline level or activity of the molecule (e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, TPP6) in the absence of sucrose, or level or activity of the molecule in the presence of sucrose, in a plant, plant part, and / or plant product of the present disclosure by 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100-1000%, 200-1000%, 300-1000%, 400-1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300-900%, 400-900%, 500-900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90- 100%, 100-200%, 200-300%, 300-400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900-1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more as compared to a control plant, plant part, and / or plant product.

[0214] In some embodiments, the methods can reduce suppression of level or activity of the molecule (e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, TPP6) induced by sucrose (i.e., in the presence of sucrose compared to in the absence of sucrose) in a plant, plant part, and / or plant product by about 10-100%, 20-100%, 30-100%, 40- 100%, 50-100%, 60-100%, 70-100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, or 70-90% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, or 90-100%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, or by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%, as compared to a control plant, plant part, and / or plant product.

[0215] In some embodiments, the methods can increase level or activity of the molecule (e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, TPP6) in the presence of sucrose relative to the absence of sucrose in a plant, plant part, and / or plant product by 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80-100%, 20- 90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100-1000%, 200-1000%, 300-1000%, 400- 1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300-900%, 400-900%, 500- 900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20-30%, 30-40%, 40- 50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100-200%, 200-300%, 300-400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900-1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more as compared to in the absence of sucrose (i.e., baseline).

[0216] In some embodiments, the methods can increase carbohydrate (e.g., sugar) transport to seeds and siliques in a plant by 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70- 100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100-1000%, 200-1000%, 300-1000%, 400-1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300- 900%, 400-900%, 500-900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100-200%, 200-300%, 300-400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900-1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more as compared to a control plant.

[0217] In the embodiments provided herein, the sugar transporter can be encoded by Glyma.10G217900, Glyma.02G075000, Glyma.02G218600, Glyma.l6G157100, Glyma.16G 156800 , Glyma.02G124100, Glyma.01G067600, or homolog of any thereof. The cell wall invertase can be encoded by Glyma.13G349300, Glyma.15G024600 , Glyma.14G096600, Glyma.16G 175800, or homolog of any thereof.

[0218] Level or activity of molecules regulated by bZIP (e.g., sugar transporter, sucrose transporter, cell-wall invertase, ASN, PDH, SPS, SPP, calmodulin, and / or TPP6) in a plant, plant part, and / or plant product can be determined by one or more standard methods known in the art. For example, expression level of a molecule regulated by the bZIP transcription factor can be measured by any means known in the art for measuring peptide production, such as measuring levels of mRNA and / or protein expression of bZIP downstream targets (e.g., quantitative RT-PCR, northern blot, SAGE, western blot analysis, ELISA, dot blot analysis). Activities of the bZIP downstream targets involved in carbohydrate transport (e.g., sugar transporter, sucrose transporter, cell-wall invertase) can be measured by a standard carbohydrate (e.g., sugar) transport assay, for example by labeling the plants with13CO2 and monitoring13C enrichment levels of sucrose, fructose, and glucose in plant organs of interest, e.g., leaves, seeds, and siliques. Activity of a sugar transporter can also be measured by using a biosensor that detects conformational changes of the transporter during sugar transport. Activities of the bZIP downstream targets involved in carbohydrate (e.g., sugar) transport or metabolism (e.g., cell wall invertase, ASN, PDH, SPS, SPP, TPP6) can be measured by enzymatic assays of the respective enzymes. For example, activity of a cell wall invertase can be measured for example as described in Tomlinson et al. 2004 J. Exp. Botany 55, 2291-2303, the entire content of which is incorporated by reference herein. Activity of calmodulin can be determined by a Ca2+-calmodulin binding assay or measuring activities of Camdependent phosphodiesterase or calcium-calmodulin dependent protein kinase (CaMKII) (Bossuyt & Bers 2013 J. Mol. Med. (Berl.) 91(8):907-916).

[0219] A control plant, plant part, or population of plant or plant part disclosed herein can be a plant or plant part to which one or more mutations has not been introduced by the methods provided herein. Thus, a control plant or plant part may express wild-type (WT) SIRT element. A control plant of the present disclosure may be grown under the same environmental conditions (e.g., same or similar temperature, humidity, air quality, soil quality, water quality, and / or pH conditions) as a plant with one or more mutations described herein. A plant, plant part, and / or plant product of the present disclosure may have altered (e.g., reduced) activity of the bZIP SIRT element, altered (e.g., increased) activity of the bZIP transcription factor, and / or altered (e.g., increased) activity of the downstream molecules regulated by the bZIP, as compared to a control plant, plant part, and / or plant product, when the plant or plant part of the present disclosure is grown under the same environmental conditions as the control plant or plant part.

[0220] B. Introducing Mutation That Decreases bZIP Transcription Factor Activity

[0221] Introducing one or more mutations into the plant genome that decreases bZIP transcription factor activity, e.g., into a SIRT element of at least one bZIP transcription factor gene, and modulating the level or activity of the bZIP transcription factor in a plant or plant part may be achieved in any method of creating a change in a nucleic acid in a plant. For example, one or more mutations can be introduced into the plant genome, e.g., into a bZIP SIRT element through the use of precise genome-editing technologies to modulate the expression of the endogenous or transgenic sequence. In this manner, a nucleic acid sequence can be inserted, substituted, or deleted proximal to or within a native plant sequence corresponding to the bZIP SIRT element through the use of methods available in the art. Such methods include, but are not limited to, use of a nuclease designed against the plant target genomic sequence of interest (D’Halluin et al 2013 Plant Biotechnol J 11 : 933-941), such as the Type II CRISPR system, the Type V CRISPR system, the CRISPR-Cas9 system, the CRISPR-Casl2a (Cpfl) system, the transcription activator-like effector nuclease (TALEN) system, the zinc finger nuclease (ZFN) system, and other technologies for precise editing of genomes [Feng et al. 2013 Cell Research 23: 1229-1232, Podevin et al. 2013 Trends Biotechnology 31 : 375-383, Wei et al. 2013 J Gen Genomics 40:281-289, Zhang et al (2013) WO 2013 / 026740, Zetsche et al. 2015 Cell 163:759-771]; Natronobacterium gregoryi Argona le-m dx&i DNA insertion (Gao et al. 2016 Nat Biotechnol doi: 10.1038 / nbt.3547); Cre- lox site-specific recombination (Dale et al. 1995 Plant 77:649-659; Lyznik, et al. 2007 Transgenic Plant J 1 :1-9; FLP-FRT recombination (Li et al. 2009 Plant Physiol 151 : 1087-1095); Bxbl- mediated integration (Yau et al. 2011 Plant 7701: 147-166); zinc-finger mediated integration (Wright et al. 2005 Plant J 44:693-705); Cai et al. 2009 Plant Mol Biol 69:699-709); and homologous recombination (Lieberman-Lazarovich and Levy 2011 Methods Mol Biol 701 : 51-65; Puchta 2002 Plant Mol Biol 48: 173-182). Reagents and compositions that can be used for introducing one or more mutations into plants or plant parts according to the methods of the present disclosure are herein described.

[0222] 1. Editing reagent

[0223] Inserting, substituting, or deleting one or more nucleotides at a precise location of interest in the genome of a plant or plant part, e.g., at the SIRT element of a bZIP transcription factor, may be achieved by introducing into the plant or plant part a system (e.g., a gene editing system), reagents (e.g., editing reagents), or a construct for introducing mutations at the target site of interest in a genome of a plant cell. A “gene editing system”, “editing system”, “gene editing reagent”, and “editing reagent” as used herein, refer to a set of one or more molecules or a construct comprising or encoding the one or more molecules for introducing one or more mutations in the genome. An exemplary gene editing system or editing reagents comprise a nuclease and / or a guide RNA. Also disclosed herein is a construct (e.g., a DNA construct, a recombinant DNA construct) for introducing one or more mutations in plants or plant parts. A construct can comprise an editing system or polynucleotides encoding editing reagents (e.g., nuclease, guide RNA, base editor) each operably linked to a promoter.

[0224] As used herein, the terms “nuclease” and “endonuclease” are used interchangeably to refer to naturally-occurring or engineered enzymes, which cleave a phosphodiester bond within a polynucleotide chain. The cleavage could be a single strand cleavage or a double strand cleavage. In certain embodiments, the nuclease lacks cleavage activity and is referred to as nuclease dead. Nucleases that can be used in precise genome-editing technologies to modulate the expression of the native sequence (e.g., the uORF, e.g., the SIRT element of a bZIP transcription factor gene) include, but are not limited to, meganucleases designed against the plant genomic sequence of interest (D’Halluin et al (2013) Plant Biotechnol 711 : 933-941); Cas9 endonuclease; Casl2a (Cpfl) endonuclease; ortholog of Cas 12a endonuclease; Cmsl endonuclease; transcription activator-like effector nucleases (TALENs); zinc finger nucleases (ZFNs); and a deactivated CRISPR nuclease (e.g., a deactivated Cas9, Casl2a, or Cmsl endonuclease) fused to a transcriptional regulatory element (Piatek et al. (2015) Plant Biotechnol J 13:578-589). In some embodiments, the editing system or the editing reagents comprise a zinc finger nuclease (ZFN), a transcription activator-like effector nuclease (TALEN), and / or a clustered regularly interspaced short palindromic repeats (CRISPR) nuclease. In some embodiments, the editing reagents comprise a CRISPR nuclease. In some embodiments, the CRISPR nuclease is a Casl2a nuclease, herein used interchangeably with a Cpfl nuclease, e.g., a McCpfl nuclease. In some embodiments, the CRISPR nuclease is a Casl2a nuclease ortholog, e.g., Lb5Casl2a, CmaCasl2a, BsCasl2a, BoCasl2a, MICas 12a, Mb2Casl2a, TsCasl2a, and MAD7 endonucleases.

[0225] A nuclease system can introduce insertion, substitution, or deletion of genetic elements at a predefined genomic locus by causing a double-strand break at said predefined genomic locus and, optionally, providing an appropriate DNA template for insertion. This strategy is well-understood and has been demonstrated previously to insert a transgene at a predefined location in the cotton genome (D’Halluin et al. 2013 Plant Biotechnol. J. 11: 933-941). For example, a Casl2a (Cpfl) endonuclease coupled with a guide RNA (gRNA) designed against the genomic sequence of interest (i.e., uORF, e.g., SIRT element of a bZIP transcription factor gene) can be used (i.e., a CRISPR-Casl2a system). Alternatively, a Cas9 endonuclease coupled with a gRNA designed against the genomic sequence of interest (a CRISPR-Cas9 system), or a Cmsl endonuclease coupled with a gRNA designed against the genomic sequence of interest (a CRISPR-Cmsl) can be used. Other nuclease systems for use with the methods of the present invention include the CRISPR systems (e.g., Type I, Type II, Type III, Type IV, and / or Type V CRISPR systems (Makarova et al 2020 Nat Rev Microbiol 18:67-83)) with their corresponding gRNA(s), the TALEN system, the ZFN system, the meganuclease system, and the like. Alternatively, a deactivated CRISPR nuclease (e.g., a deactivated Cas9, Cas 12a, or Cmsl endonuclease) fused to a transcriptional regulatory element can be targeted to the regulatory region (e.g., upstream regulatory region) of at least one bZIP transcription factor gene, thereby modulating the transcription of the bZIP transcription factor gene (Piatek et al. 2015 Plant Biotechnol J 13:578-589). Site-specific introduction of mutations of plant cells by biolistic introduction of a ribonucleoprotein comprising a nuclease and suitable guide RNA has been demonstrated (Svitashev et al. 2016 Nat Commun doi: 10.1038 / ncommsl3274), and is herein incorporated by reference. For example, a CRISPR system comprises a CRISPR nuclease (e.g., CRISPR-associated (Cas) endonuclease or variant or ortholog thereof, such as Casl2a or Cas 12a ortholog) and a guide RNA. A CRISPR nuclease associates with a guide RNA that directs nucleic acid cleavage by the associated endonuclease by hybridizing to a recognition site in a polynucleotide. The guide RNA directs the nuclease to the target site and the endonuclease cleaves DNA at the target site. The guide RNA comprises a direct repeat and a guide sequence, which is complementary to the target recognition site. In certain embodiments, the CRISPR system further comprises a tracrRNA (trans-activating CRISPR RNA) that is complementary (fully or partially) to the direct repeat sequence present on the guide RNA. The CRISPR-Casl2a system may comprise at least one guide RNA (gRNA) operatively arranged with the ortholog endonuclease for genomic editing of a target DNA binding the gRNA. The system may comprise a CRISPR-Casl2a expression system encoding the Casl2a ortholog nucleases and crRNAs (CRISPR RNAs) for forming gRNAs that are coactive with the Casl2a nucleases. A “TALEN” nuclease is an endonuclease comprising a DNA-binding domain comprising a plurality of TAL domain repeats fused to a nuclease domain or an active portion thereof from an endonuclease or exonuclease, including but not limited to a restriction endonuclease, homing endonuclease, and yeast HO endonuclease. A “zinc finger nuclease” or “ZFN” refers to a chimeric protein comprising a zinc finger DNA-binding domain fused to a nuclease domain from an endonuclease or exonuclease, including but not limited to a restriction endonuclease, homing endonuclease, and yeast HO endonuclease.

[0226] The editing system, editing reagents, or construct described herein can comprise one or more guide RNAs (gRNAs), or gRNA cassette, to drive mutations at the locus of the SIRT element of a bZIP transcription factor gene. “Guide RNA” as used herein refers to a RNA molecule that function as guides for RNA- or DNA-targeting enzymes, e.g., nucleases. In some instances, a gRNA can comprise a targeting region (i.e., spacer) that is complementary to a targeted sequence as well as another region that allows the gRNA to form a complex with a nuclease (e.g., a CRISPR nuclease) of interest.

[0227] For example, the editing system, the editing reagent, or the construct of the present disclosure may contain a gRNA cassette, comprising one or more gRNAs or encoding one or more gRNAs, to drive one or more deletion (e.g., deletion of 10 or more nucleotides) in the uORF, e.g., the SIRT element of one or both alleles of a bZIP transcription factor gene, e.g., a Glycine max bZIP 123 transcription factor gene, a Glycine max bZIP 125 transcription factor gene, a Glycine max bZIP 124 transcription factor gene, a Glycine max bZIP 126 transcription factor gene, a Glycine max bZIP 18 transcription factor gene, a Glycine max bZIP 17 transcription factor gene. The one or more gRNAs can be designed to specifically target a regulatory region (e.g., uORF, SIRT element) of a bZIP transcription factor gene. In some embodiments, the one or more gRNAs are specific to a bZIP SIRT element, e.g., GmbZIP123 SIRT element, GmbZIP 125 SIRT element. For example, the gRNA can be specific to a nucleic acid sequence having at least 75% (75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to the nucleic acid sequence of any one of SEQ ID NOs: 13-18. The gRNA can be specific to the nucleic acid sequence of any one of SEQ ID NOs: 13-18 and / or can drive a deletion at least partially in the SIRT element of the Glycine max bZIP 123 transcription factor gene, Glycine max bZIP125 transcription factor gene, or active homolog thereof. In particular instances, the gRNA can facilitate binding of an RNA guided nuclease that cleaves a region of the bZIP uORF, e.g., bZIP SIRT element, and cause non-homologous end joining or homology-directed repair to introduce a mutation at the cleavage site.

[0228] The methods provided herein can comprise introducing into the plant, plant part, or plant cell two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more) gRNAs specific to a nucleic acid sequence having at least 75% (75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to the nucleic acid sequence of any one of SEQ ID NOs: 13-18. The two or more gRNA can be specific to the nucleic acid sequence of any one of SEQ ID NOs: 13-18 and / or can drive one or more deletions at least partially in the 5’ regulatory region (e.g., uORF, SIRT element) of the bZIP transcription factor gene, e.g., bZIP123 transcription factor gene, bZIP125 transcription factor gene, bZIP 124 transcription factor, bZIP 126 transcription factor, bZIP 18 transcription factor, bZIP 17 transcription factor or active homolog thereof in the plant, plant part, or plant cell. In some instances, introducing two or more gRNAs along with other editing reagents (e.g., nuclease) into the plant, plant part, or plant cell increases sequence diversity of mutations (e.g., insertions, substitutions, deletions) generated at or near the target site, as compared to introducing one gRNA.

[0229] The targeting region (i.e. spacer) of a gRNA that binds to the region of the SIRT element of a bZIP transcription factor gene for use in the method described herein can be about 100-300 nucleotides long, with the targeting region therein about 10-40 nucleotides long (e.g., 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, or 40 nucleotides long). For example, the targeting region of a gRNA for use in the method described herein may be about 24 nucleotides in length. In some embodiments, the targeting region of a gRNA is encoded by a nucleic acid sequence comprising a nucleic acid sequence having at least 75% (e.g., 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) sequence identity to the nucleic acid sequence of any one of SEQ ID NOs: 25-29. In particular instances, the targeting region of a gRNA for use in the method described herein is encoded by a nucleic acid sequence comprising the nucleic acid sequence of any one of SEQ ID NOs: 25-29. The methods provided herein can comprise introducing into the plant, plant part, or plant cell one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more) gRNAs, at least one of which comprising a nucleic acid sequence encoded by a nucleic acid sequence that shares at least 80% sequence identity with the nucleic acid sequence of any one of SEQ ID NOs: 25-29 or a nucleic acid sequence of any one of SEQ ID NOs: 25-29. For example, the methods provided herein can comprise introducing into the plant, plant part, or plant cell at least one of GmbZIP123 gRNA182, GmbZIP123 gRNA183, GmbZIP125 gRNA207, GmbZIP125 gRNA206, GmbZIP124 gRNA97, GmbZIP126 gRNA156, GmbZIP18 gRNA91, and GmbZIP17 gRNA73. The methods provided herein can comprise introducing into the plant, plant part, or plant cell two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more) gRNAs.

[0230] The one or more gRNAs provided herein can direct a nuclease to a specific target site at a region (e.g., SIRT element) of the uORF of a Glycine max bZIP123 transcription factor gene, a Glycine max bZIP125 transcription factor gene, a Glycine max bZIP124 transcription factor gene, a Glycine max bZIP126 transcription factor gene, a Glycine max bZIP18 transcription factor gene, or a Glycine max bZIP17 transcription factor gene, and introduce into the plant, plant part, or plant cell a deletion of one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) nucleotides in the SIRT element or a region including the SIRT element of the a Glycine max bZIP123 transcription factor gene, Glycine max bZIP125 transcription factor gene, a Glycine max bZIP124 transcription factor gene, a Glycine max bZIP126 transcription factor gene, a Glycine max bZIP18 transcription factor gene, or a Glycine max bZIP17 transcription factor gene.

[0231] In some embodiments, a gene editing efficiency of the one or more gRNAs is 0.3% or greater (e.g., 0.5%, 1%, 1.5%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100%). In specific embodiments, the methods do not introduce mutations into at least one allele of the bZIP SIRT element. In some embodiments, the methods introduce mutations into all alleles of the bZIP SIRT element.

[0232] Editing system or editing reagents can also include base editing components. For example, cytosine base editing (CBE) reagents, which change a C-G base pair to a T-A base pair, comprise a single guide RNA, a nuclease (e.g., dCas9, CAS9 nickase), a cytidine deaminase (e.g., APOB EC 1), and a uracil DNA glycosylase inhibitor (UGI). Adenine base editing (ABE) reagents, which change an A-T base pair to a G-C base pair comprise a deaminase, (TadA), a nuclease (e.g., dCas or Cas nickase), and a guide RNA.

[0233] The gene editing system (e.g., CRISPR-Casl2a system), editing reagents, or a construct of the present disclosure can comprise at least one CRISPR RNA (crRNA) regulatory element operably linked to at least one nucleotide sequence encoding a crRNA for producing gRNA for targeting a target sequence, and at least one regulatory element, which may be the same as or different from the crRNA regulatory element, operably linked to a nucleotide sequence encoding the endonuclease, for generation of a CRISPR editing structure (e.g., CRISPR-Casl2a editing structure) by which the gRNA targets the target sequence and the CRISPR endonuclease cleaves a target DNA to alter gene expression in the cell, and wherein the CRISPR-associated nuclease, and the gRNA, do not naturally occur together. In such system, the at least one crRNA regulatory element may comprise one or more than one RNA polymerase II (Pol II) promoter, or alternatively, a single transcript unit (STU) regulatory element, or one or more of ZmUbi, OsU6, OsU3, and U6 promoters.

[0234] The methods described herein, comprising introducing into such plant a non-naturally occurring heterologous CRISPR-Casl2a genomic editing system of a type as variously described herein, can cause the editing reagents to introduce mutations in at least one bZIP SIRT element. The gene editing system (e.g., the CRISPR-Casl2a system) can target PAM sites such as TTN, TTV, TTTV, NTTV, TATV, TATG, TATA, YTTN, GTTA, and / or GTTC.

[0235] Such methods of introducing mutations into plants, plant parts, or plant cells may be carried out at moderate temperatures, e.g., below 25 °C. and above temperature producing freezing or frost damage of the plant. The methods provided herein may be performed on a wide variety of plants. In particular embodiments, the methods provided herein can be carried out to introduce mutations into the Glycine max plant at the SIRT element of one or more bZIP transcription factor gene.

[0236] Methods disclosed herein are not limited to certain techniques of mutagenesis. Any method of creating a change in a nucleic acid of a plant can be used in conjunction with the disclosed invention, including the use of chemical mutagens (e.g. methanesulfonate, sodium azide, aminopurine, etc.), genome / gene editing techniques (e.g. CRISPR-like technologies, TALENs, zinc finger nucleases, and meganucleases), ionizing radiation (e.g. ultraviolet and / or gamma rays) temperature alterations, long-term seed storage, tissue culture conditions, targeting induced local lesions in a genome, sequence-targeted and / or random recombinases, etc. It is anticipated that new methods of creating a mutation in a nucleic acid of a plant will be developed and yet fall within the scope of the claimed invention when used with the teachings described herein. Any editing system or editing reagents for use in any genome-editing methods including those described herein can be expressed in a plant or plant part.

[0237] 2. Promoter

[0238] As used herein, “promoter” refers to a regulatory region of DNA that is capable of driving expression of a sequence in a plant or plant cell. A number of promoters may be used in the practice of the disclosure, e.g., to express editing reagents in plants, plant parts, or plant cells. The promoter may have a constitutive expression profile. Constitutive promoters include the CaMV 35S promoter (Odell et al. (1985) Nature 313:810-812); rice actin (McElroy et al. (1990) Plant Cell 2: 163-171); ubiquitin (Christensen et al. (1989) Plant Mol. Biol. 12:619-632 and Christensen et al. (1992) Plant Mol. Biol. 18:675-689); pEMU (Last et al. (1991) Theor. Appl. Genet. 81 :581-588); MAS (Velten et al. (1984) EMBO J. 3:2723-2730); ALS promoter (U.S. Patent No. 5,659,026), and the like.

[0239] Alternatively, promoters for use in the methods of the present disclosure can be tissuepreferred promoters. Tissue-preferred promoters include Yamamoto et al. (1997) Plant J. 12(2):255-265; Kawamata et al. (1997) Plant Cell Physiol. 38(7):792-803; Hansen et al. (1997) Mol. Gen Genet. 254(3):337-343; Russell et al. (1997) Transgenic Res. 6(2): 157-168; Rinehart et al. (1996) Plant Physiol. 112(3): 1331-1341; Van Camp et al. (1996) Plant Physiol. 112(2):525- 535; Canevascini et al. (1996) Plant Physiol. 112(2):513-524; Yamamoto et al. (1994) Plant Cell Physiol. 35(5):773-778; Lam (1994) Results Probl. Cell Differ. 20: 181-196; Orozco et al. (1993) Plant Mol Biol. 23(6): 1129-1138; Matsuoka et al. (1993) Proc Natl. Acad. Sci. USA 90(20):9586- 9590; and Guevara-Garcia et al. (1993) Plant J. 4(3):495-505. Leaf-preferred promoters are also known in the art. See, for example, Yamamoto et al. (1997) Plant J. 12(2):255-265; Kwon et al. (1994) Plant Physiol. 105:357-67; Yamamoto et al. (1994) Plant Cell Physiol. 35(5):773-778; Gotor et al. (1993) Plant J. 3:509-18; Orozco et al. (1993) Plant Mol. Biol. 23(6): 1129-1138; and Matsuoka et al. (1993) roc. Natl. Acad. Sci. USA 90(20):9586-9590.

[0240] Alternatively, promoters for use in the methods of the present disclosure can be developmentally-regulated promoters. Such promoters may show a peak in expression at a particular developmental stage. Such promoters have been described in the art, e.g., US Patent No. 10,407,670; Gan and Amasino (1995) Science 270: 1986-1988; Rinehart et al. (1996) Plant Physiol 112: 1331-1341; Gray-Mitsumune et al. (1999) Plant Mol Biol 39: 657-669; Beaudoin and Rothstein (1997) Plant Mol Biol 33: 835-846; Genschik et al. (1994) Gene 148: 195-202, and the like.

[0241] Alternatively, promoters for use in the methods of the present disclosure can be promoters that are induced following the application of a particular biotic and / or abiotic stress. Such promoters have been described in the art, e.g., Yi et al. (2010) Planta 232: 743-754; Yamaguchi- Shinozaki and Shinozaki (1993) Mol Gen Genet 236: 331-340; U.S. Patent No. 7,674,952; Rerksiri et al. (2013) Sci World J 2013: Article ID 397401; Khurana et al. (2013) PloS One 8: e54418; Tao et al. (2015) Plant Mol Biol Rep 33: 200-208, and the like. Alternatively, promoters for use in the methods of the present disclosure can be cellpreferred promoters. Such promoters may preferentially drive the expression of a downstream gene in a particular cell type such as a mesophyll or a bundle sheath cell. Such cell-preferred promoters have been described in the art, e.g., Viret et al. (1994) Proc Natl Acad USA 91 : 8577-8581; U.S. Patent No. 8,455,718; U.S. Patent No. 7,642,347; Sattarzadeh et al. (2010) Plant Biotechnol J 8: 112-125; Engelmann et al. (2008) Plant Physiol 146: 1773-1785; Matsuoka et al. (1994) Plant J 6 : 311-319, and the like.

[0242] It is recognized that a specific, non-constitutive expression profile may provide an improved plant phenotype relative to constitutive expression of a gene or genes of interest. For instance, many plant genes are regulated by light conditions, the application of particular stresses, the circadian cycle, or the stage of a plant’s development. These expression profiles may be important for the function of the gene or gene product in planta. One strategy that may be used to provide a desired expression profile is the use of synthetic promoters containing cv.s-regulatory elements that drive the desired expression levels at the desired time and place in the plant. Cis-regulatory elements that can be used to alter gene expression in planta have been described in the scientific literature (Vandepoele et al. (2009) Plant Physiol 150: 535-546; Rushton et al. (2002) Plant Cell 14: 749-762). G'.s-regulatory elements may also be used to alter promoter expression profiles, as described in Venter (2007) Trends Plant Sci 12: 118-124.

[0243] 3. Transfer DNA

[0244] Nucleic acid molecules comprising transfer DNA (T-DNA) sequences can be used in the practice of the disclosure, e.g., to express editing reagents in plants, plant parts, or plant cells. For example, a construct of the present disclosure may contain T-DNA of tumor-inducing (Ti) plasmid of Agrobacterium tumefaciens. Alternatively, a recombinant DNA construct of the present disclosure may contain T-DNA of tumor-inducing (Ti) plasmid of Agrobacterium rhizogenes. The vir genes of the Ti plasmid may help in transfer of T-DNA of a recombinant DNA construct into nuclear DNA genome of a host plant. For example, Ti plasmid of Agrobacterium tumefaciens may help in transfer of T-DNA of a recombinant DNA construct of the present disclosure into nuclear DNA genome of a host plant, thus enabling the transfer of a gRNA of the present disclosure into nuclear DNA genome of a host plant (e.g., a pea plant).

[0245] 4. Regulatory signal

[0246] Construct described herein may contain regulatory signals, including, but not limited to, transcriptional initiation sites, operators, activators, enhancers, other regulatory elements, ribosomal binding sites, an initiation codon, termination signals, and the like. See, for example, U.S. Pat. Nos. 5,039,523 and 4,853,331; EPO 0480762A2; Sambrook et al. (1992) Molecular Cloning: A Laboratory Manual, ed. Maniatis et al. (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.), hereinafter “Sambrook 11”; Davis et al., eds. (1980) Advanced Bacterial Genetics (Cold Spring Harbor Laboratory Press), Cold Spring Harbor, N. Y., and the references cited therein.

[0247] 5. Reporter genes / selectable marker genes

[0248] Reporter genes or selectable marker genes may be included in the expression cassettes of the present invention. Examples of suitable reporter genes known in the art can be found in, for example, Jefferson, et al., (1991) in Plant Molecular Biology Manual, ed. Gelvin, et al., (Kluwer Academic Publishers), pp. 1-33; DeWet, et al., (1987) Mol. Cell. Biol. 7:725-737; Goff, et al., (199Q) EMBO J. 9:2517-2522; Kain, et al., (1995) Bio Techniques 19:650-655 and Chiu, et al., (1996) Current Biology 6:325-330, herein incorporated by reference in their entirety.

[0249] Selectable marker genes for selection of transformed cells or tissues can include genes that confer antibiotic resistance or resistance to herbicides. Examples of suitable selectable marker genes include, but are not limited to, genes encoding resistance to chloramphenicol (Herrera Estrella, et al., (1983) EMBO J. 2:987-992); methotrexate (Herrera Estrella, et al., (1983) Nature 303:209-213; Meijer, et al., (1991) Plant Mol. Biol. 16:807-820); hygromycin (Waldron, et al., (1985) Plant Mol. Biol. 5: 103-108 and Zhijian, et al., (1995) Plant Science 108:219-227); streptomycin (Jones, et al., (1987) Mol. Gen. Genet. 210:86-91); spectinomycin (Bretagne-Sagnard, et al. , (1996) Transgenic Res. 5: 131-137); bleomycin (Hille, et al. , (1990) Plant Mol. Biol. 7 : 171- 176) ; sulfonamide (Guerineau, et al., (1990) Plant Mol. Biol. 15: 127-36); bromoxynil (Stalker, et al., (1988) Science 242:419-423); glyphosate (Shaw, et al., (1986) Science 233:478-481 and US Patent Application Serial Numbers 10 / 004,357 and 10 / 427,692); phosphinothricin (DeBlock, et al., (1987) EMBO J. 6:2513-2518), herein incorporated by reference in their entirety.

[0250] Selectable marker genes include genes encoding antibiotic resistance, such as those encoding neomycin phosphotransferase II (NEO), spectinomycin / streptinomycin resistance (SpcR, AAD), and hygromycin phosphotransferase (HPT or HGR) as well as genes conferring resistance to herbicidal compounds. Herbicide resistance genes generally code for a modified target protein insensitive to the herbicide or for an enzyme that degrades or detoxifies the herbicide in the plant before it can act. For example, resistance to glyphosate has been obtained by using genes coding for mutant target enzymes, 5 -enolpyruvylshikimate-3 -phosphate synthase (EPSPS). Genes and mutants for EPSPS are well known, and further described below. Resistance to glufosinate ammonium, bromoxynil, and 2,4-dichlorophenoxyacetate (2,4-D) have been obtained by using bacterial genes encoding PAT or DSM-2, a nitrilase, an AAD-1, or an AAD-12, each of which are examples of proteins that detoxify their respective herbicides. Herbicides can inhibit the growing point or meristem, including imidazolinone or sulfonylurea, and genes for resistance / tolerance of acetohydroxyacid synthase (AHAS) and acetolactate synthase (ALS) for these herbicides are well known. Glyphosate resistance genes include mutant 5 -enolpyruvylshikimate-3 -phosphate synthase (EPSPs) and dgt-28 genes (via the introduction of recombinant nucleic acids and / or various forms of in vivo mutagenesis of native EPSPs genes), aroA genes and glyphosate acetyl transferase (GAT) genes, respectively). Resistance genes for other phosphono compounds include bar and pat genes from Streptomyces species, including Streptomyces hygroscopicus and Streptomyces viridichromogenes, and pyridinoxy or phenoxy proprionic acids and cyclohexones (ACCase inhibitor-encoding genes). Exemplary genes conferring resistance to cyclohexanediones and / or aryloxyphenoxypropanoic acid (including haloxyfop, diclofop, fenoxyprop, fluazifop, quizalofop) include genes of acetyl coenzyme A carboxylase (ACCase); Accl-Sl, Accl-S2 and Accl-S3. Herbicides can also inhibit photosynthesis, including triazine (psbA and ls+ genes) or benzonitrile (nitrilase gene). Further, such selectable markers can include positive selection markers such as phosphomannose isomerase (PMI) enzyme.

[0251] Selectable marker genes can further include, but are not limited to genes encoding: 2,4-D; SpcR; neomycin phosphotransferase II; cyanamide hydratase; aspartate kinase; dihydrodipicolinate synthase; tryptophan decarboxylase; dihydrodipicolinate synthase and desensitized aspartate kinase; bar gene; tryptophan decarboxylase; neomycin phosphotransferase (NEO); hygromycin phosphotransferase (HPT or HYG); dihydrofolate reductase (DHFR); phosphinothricin acetyltransferase; 2,2-dichloropropionic acid dehalogenase; acetohydroxyacid synthase; 5- enolpyruvyl-shikimate-phosphate synthase (aroA); haloarylnitrilase; acetyl-coenzyme A carboxylase; dihydropteroate synthase (sul I); and 32 kD photosystem II polypeptide (psbA). Selectable marker genes can further include genes encoding resistance to: chloramphenicol; methotrexate; hygromycin; spectinomycin; bromoxynil; glyphosate; and phosphinothricin.

[0252] Other selectable marker genes that could be employed on the expression constructs disclosed herein include, but are not limited to, GUS (beta-glucuronidase; Jefferson, (1987) Plant Mol. Biol. Rep. 5:387), GFP (green fluorescence protein; Chalfie, et al., (1994) Science 263:802), luciferase (Riggs, et al., (1987) Nucleic Acids Res. 15(19):8115 and Luehrsen, et al., (1992) Methods Enzymol. 216:397-414), red fluorescent protein (DsRFP, RFP, etc), beta-galactosidase, and the maize genes encoding for anthocyanin production (Ludwig, et al., (1990) Science 247:449), and the like (See Sambrook, et al., Molecular Cloning: A Laboratory Manual, Third Edition, Cold Spring Harbor Press, N.Y., 2001), herein incorporated by reference in their entirety. The above list of selectable marker genes is not meant to be limiting. Any reporter or selectable marker gene are encompassed by the present disclosure. 6. Terminator

[0253] A transcription terminator may also be included in the expression cassettes of the present invention. Plant terminators are known in the art and include those available from the Ti-plasmid of A. tumefaciens, such as the octopine synthase and nopaline synthase termination regions. See also Guerineau et al. (1991) Mol. Gen. Genet. 262: 141-144; Proudfoot (1991) Cell 64:671-674; Sanfacon et al. (1991) Genes Dev. 5: 141-149; Mogen et al. (1990) Plant Cell 2: 1261-1272; Munroe et al. (1990) Gene 91 : 151-158; Ballas et al. (1989) Nucleic Acids Res . 17:7891-7903; and Joshi et al. (1987) Nucleic Acids Res. 15:9627-9639.

[0254] 7. Vector

[0255] Disclosed herein are vectors containing constructs (e.g., recombinant DNA constructs encoding editing reagents) of the present disclosure. As used herein, “vector” refers to a nucleotide molecule (e.g., a plasmid, cosmid), bacterial phage, or virus for introducing a nucleotide construct, for example, a recombinant DNA construct, into a host cell. Cloning vectors typically contain one or a small number of restriction endonuclease recognition sites at which foreign DNA sequences can be inserted in a determinable fashion without loss of essential biological function of the vector, as well as a marker gene that is suitable for use in the identification and selection of cells transformed with the cloning vector. Marker genes typically include genes that provide tetracycline resistance, hygromycin resistance or ampicillin resistance. In some embodiments, provided herein are expression cassettes located on a vector comprising gRNA sequence specific to a bZIP uORF or a bZIP SIRT element.

[0256] In some embodiments, a vector is a plasmid containing a recombinant DNA construct of the present disclosure. For example, the present disclosure may provide a plasmid containing a recombinant DNA construct that comprises a gRNA to drive mutations at the locus of the uORF or SIRT element of one or more bZIP transcription factor gene copies.

[0257] In some embodiments, a vector is a recombinant virus containing a recombinant DNA construct of the present disclosure. For example, the present disclosure may provide a recombinant virus containing a recombinant DNA construct that comprises a gRNA, wherein the gRNA can drive mutations at the locus of the uORF or SIRT element of one or more bZIP transcription factor genes. A recombinant virus described herein can be a recombinant lentivirus, a recombinant retrovirus, a recombinant cucumber mosaic virus (CMV), a recombinant tobacco mosaic virus (TMV), a recombinant cauliflower mosaic virus (CaMV), a recombinant odontoglossum ringspot virus (ORSV), a recombinant tomato mosaic virus (ToMV), a recombinant bamboo mosaic virus (BaMV), a recombinant cowpea mosaic virus (CPMV), a recombinant potato virus X (PVX), a recombinant Bean yellow dwarf virus (BeYDV), or a recombinant turnip vein-clearing virus (TVCV).

[0258] 8. Cells

[0259] Also provided herein are cells comprising the reagent (e.g., editing reagent, e.g., nuclease, gRNA), the system (e.g., gene editing system), the construct (e.g., expression cassette), and / or the vector of the present disclosure for introducing mutations into the uORF, e.g., a SIRT element of at least one bZIP transcription factor gene. The cell can be a plant cell, a bacterial cell, or a fungal cell. The cell can be a bacterium, e.g., an Agrobacterium lumefaciens. containing the gRNA targeting the uORF, e.g., the SIRT element of a bZIP transcription factor gene and driving mutations at the target site of interest. The cells of the present disclosure may be grown, or have been grown, in a cell culture.

[0260] C. Increasing Fatty Acid / Oil and / or Sugar Content in Plants

[0261] The methods of the present disclosure, by introducing a mutation that increases bZIP transcription factor activity into plants, plant parts, or plant cells and / or regenerating plants from transformed cells, can increase fatty acid, oil, and / or sugar content in the plants, plant parts (e.g., seeds, leaves), a population of plants or plant parts, or plant products (e.g., seed composition, oil composition, sugar composition) as compared to a control (e.g., wild-type) plant, plant part, population of plants or plant parts, or plant product. “Fatty acid” as used herein can include any fatty acid found in a plant, plant part, or plant oil, including palmitic acid (16:0), stearic acid (18:0), oleic acid (18: 1), linoleic acid (18:2), and linolenic acid (18:3).

[0262] A control plant, plant part, or a population of plants or plant parts can be a plant or plant part to which one or more mutations has not be introduced by the methods provided herein, i.e., having a native bZIP SIRT element or a native uORF of the bZIP transcription factor gene; reference plants, plant parts, or population; or commodity plants, plant parts, or population). A control plant of the present disclosure may be grown under the same environmental conditions (e.g., same or similar temperature, humidity, air quality, soil quality, water quality, and / or pH conditions) as a plant with one or more mutations described herein. A plant, plant part, and / or plant product of the present disclosure may have increased levels of fatty acid, oil, and / or sugar (e.g., sucrose) content as compared to a control plant, plant part, and / or plant product, when the plant or plant part of the present disclosure is grown under the same environmental conditions as the control plant or plant part. A reference (e.g., control) sample of soybean plant or seed, e.g., a commodity soybean or seed, can have oil content of from about 8% to about 28%, while influenced by both genotype and environmental factors (Clemente & Cahoon 2009, Plant Physiol. 151 : 1030-1040).

[0263] In some embodiments, the methods can increase total fatty acid, oil, and / or sugar (e.g., sucrose) content by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100%, 70-100%, 80-100%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 100-1000%, 200-1000%, 300- 1000%, 400-1000%, 500-1000%, 600-1000%, 700-1000%, 800-1000%, 200-900%, 300-900%, 400-900%, 500-900%, 600-900%, 700-900%, or more than 1000% (e.g., by about 10-20%, 20- 30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100-200%, 200-300%, 300- 400%, 400-500%, 500-600%, 600-700%, 700-800%, 800-900%, 900-1000%, or more than 1000%), e.g., by about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more, or at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 200%, 300%, 400%, 500%, 600%, 700%, 800%, 900%, 1000%, or more in the plants or plant parts of the present disclosure as compared to a control plant or plant part. In some embodiments, the methods can increase total fatty acid, oil, and / or sugar (e.g., sucrose) content, as expressed by % dry weight, in the plant, plant part, or population of plant or plant parts provided herein relative to that in control plant, plant part, or population, and the difference (by subtraction) is about 0.25-10%, 0.5-10%, 0.75-10%, 1.0-10%, 1.5-10%, 2-10%, 2.5- 10%, 3-10%, 3.5-10%, 4-10%, 4.5-10%, 5-10%, 6-10%, 7-10%, 8-10%, 9-10%, or more than 10% (e.g., by about 0.25-0.5%, 0.5-0.75%, 0.75-1.0%, 1.0-1.5%, 1.5-2.0%, 2.0-2.5%, 2.5-3.0%, 3.0- 3.5%, 3.5-4.0%, 4.0-4.5%, 4.5-5.0%, 5-6%, 6-7%, 7-8%, or 8-9%, 9-10%, or more than 10%), by about 0.25%, 0.5%, 0.75%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5%, 6%, 7%, 8%, 9%, 10%, or more, or at least 0.25%, 0.5%, 0.75%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5%, 6%, 7%, 8%, 9%, 10%, or more fatty acid / oil and / or sugar content. In specific embodiments, total oil content as expressed by % dry weight in the plant or plant part or population of plants or plant parts provided herein is greater than that in the control plant, plant part, or population, and the difference (by subtraction) is about 0.5-2.5%, 0.8-1.8%, or 1-1.6%, e.g., about 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%. 1.1%, 1.2%, 1.3%, 1.4%, 1.5%, 1.6%, 1.7%, 1.8%, 1.9%, 2.0%, 2.1%, 2.2%, 2.3%, 2.4%, or 2.5%, e.g., 1.3% or 1.4% dry weight. Sucrose content as expressed by % dry weight in the plant or plant part or population of plants or plant parts provided herein can be greater than that in the control plant, plant part, or population, and the difference (by subtraction) can be about 0.1-1.5%, 0.1-1%, 0.4-0.6%, e.g., about 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.1%, 1.2%, 1.3%, 1.4%, 1.5%, e.g., 0.5% dry weight.

[0264] In specific embodiments, the methods can increase seed fatty acid / oil and / or sugar (e.g., sucrose) content in seeds or a population of seeds relative to control seeds or a control population of seeds (e.g., control seeds or population having a native bZIP SIRT element, reference seeds or population, commodity seeds or population). The seeds can be legume seeds, e.g., pea seeds or soybean seeds. The oil content of typical soybean cultivars average at approximately 20%, ranging from approximately 8% to approximately 28%, while influenced by both genotype and environmental factors (Clemente & Cahoon 2009, Plant Physiol. 151 : 1030-1040). In contrast, the methods can increase seed fatty acid / oil content in the soybean seeds or population of seeds to at least 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more by dry weight. In specific embodiments, a reference sample of soybean plant or seed has oil content of about 20-21%, which can be increased to about 21-22% or more in the plant or plant part comprising the one or more mutations. A reference sample of soybean plant or seed can have sucrose content of about 7-8%, which can be increased to about 8-9% or more in the plant or plant part comprising the one or more mutations.

[0265] Fatty acid / oil content in plants, plant parts, or a population of plants or plant parts can be measured to determine if the mutations introduced to the SIRT element alters seed fatty acid / oil content. Amount or levels of oil, total fatty acids, and specific fatty acids can be measured by any methods for measuring oil or fatty acid amount or levels in a plant sample, including NIR, GC-MS optionally with certain modifications (e.g., with or without initial lipid extraction, with or without isotope labeling of analytes), or NMR. Fatty acid composition (e.g., percentage of specific fatty acids normalized to total fatty acids) can be calculated based on the amount or concentration of total fatty acids and specific fatty acids in the sample. Amount or level of sucrose, total sugar, glucose, fructose, galactose, maltose, and / or lactose in seeds can be measured by any methods for measuring total or specific sugar content in a plant sample, including NIR, SPE, SPME, HPLC, GCMS, and / or enzymatic assay.

[0266] In specific embodiments, the methods can increase fatty acid, oil, and / or sugar content in the plant, plant part, or plant product as compared to a control plant, plant part, population of plants or plant parts, or plant product, without a significant decrease in yield. In some embodiments, the methods can produce a reduction in yield in the plant, plant part, or population of plants or plant parts of the present disclosure, having increased fatty acid, oil, and / or sugar content, that is no more than about 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, or about 5.0%, 6%, 7%, 8%, 9%, or 10%, e.g., no more than about 0-5%, 0.5-4.5%, 0.5-4%, 1-5%, 1-4%, 2-5%, 2-4%, 0.5-10%, 0.5-8%, 1-10%, 2-10%, 3-10%, 4-10%, 5-10%, 6-10%, 7-10%, or 8-10% reduction in yield as compared to a control plant, plant part, or population of plants or plant parts. Yield can be measured and expressed by any means known in the art. In specific embodiments, yield is measured by seed weight or volume of seeds, fruits, leaves, or whole plants harvested from a given harvest area. In specific embodiments, provided herein are methods of increasing bZIP transcription factor activity and fatty acid, oil, and / or sugar (e.g., sucrose) content in seeds or a population of seeds as compared to control seeds or a population of seeds.

[0267] D. Plants, plant parts, population, and plant products produced by present methods

[0268] The present disclosure provides plants, plant parts, a population of plants or plant parts, and plant products produced according to the methods provided herein. Such plants, plant parts, population of plants or plant parts, and plant products (e.g., seed compositions, oil compositions, sugar composition, and plant-based food / beverage products) can contain a mutation that decreases bZIP transcription factor activity, e.g., one or more insertions, substitutions, or deletions at least partially in the SIRT element of at least one native bZIP transcription factor gene (e.g., bZIP123, ZIP125, bZIP124, bZIP126, bZIP18, bZIP17); reduced bZIP SIRT activity; increased bZIP transcription factor activity; increased activity or level of one or more molecules regulated by the bZIP transcription factor (e.g., molecules that regulate carbohydrate transport or metabolism, e.g., sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, TPP6); increased carbohydrate (e.g., sugar) transport to seeds or siliques; and / or increased oil / fatty acid and / or sugar content compared to a control (e.g., wild-type, commodity, reference) plant, plant part, population, or a control plant product produced therefrom. In specific embodiments, the plants, plant parts, a population of plants or plant parts, or plant products (e.g., plant seed compositions, oil compositions, sugar compositions) produced by the methods provided herein are a soybean (Glycine max) plant, plant part, plant population, or a plant product produced therefrom that contains a mutation that decreases bZIP transcription factor (e.g., bZIP123 transcription factor, bZIP125 transcription factor, bZIP124 transcription factor, bZIP 126 transcription factor, bZIP 18 transcription factor, bZIP 17 transcription factor) activity, e.g., one or more insertions, substitutions, or deletions at least partially in the SIRT element of at least one native bZIP transcription factor gene (e.g., bZIP123, ZIP125, bZIP124, bZIP 126, bZIP 18, bZIP 17) or homolog thereof.

[0269] A “plant part” produced according to the methods described herein can include any part of a plant, including seeds (e.g., a representative sample of seeds), plant cells, embryos, pollen, ovules, leaves, flowers, branches, fruit, kernels, ears, cobs, husks, stalks, roots, root tips, anthers, plant protoplasts, plant cell tissue cultures from which plants can be regenerated, plant calli, plant clumps, juice, pulp, nectar, stems, branches, and bark. A “plant product”, as used herein, refers to any composition derived from the plant or plant part, including any oil products, sugar products, fiber products, protein products (such as protein concentrate, protein isolate, flake, or other protein product), seed hulls, meal, or flour, for a food, feed, aqua, or industrial product, plant extract (e.g., sweetener, antioxidants, alkaloids, etc.), plant concentrate (e.g., whole plant concentrate or plant part concentrate), plant powder (e.g., formulated powder, such as formulated plant part powder (e.g., seed flour)), plant biomass (e.g., dried biomass, such as crushed and / or powdered biomass), grains, plant protein composition, plant oil composition, plant sugar composition, and food and beverage products containing plant compositions (e.g., plant parts, plant extract, plant concentrate, plant powder, plant protein, plant oil, and plant biomass) described herein. Plant parts and plant products produced according to the methods provided herein can be intended for human or animal consumption. In specific embodiments, provided herein are a seed composition, an oil composition, and a sugar composition produced from the plants or plant parts of the present disclosure.

[0270] Also provided herein are food and / or beverage products obtained from the plants, plant parts, or plant compositions (e.g., seed composition, plant protein compositions) produced according to the methods of the present disclosure. Plants, seed compositions, oil compositions, and sugar compositions provided herein can be suitable for use in a variety of food and beverage products, such as plant milk, plant nut butter, plant meal, plant flour, cooking oil, salad oil, shortening, lecithin, animal feed, shakes, health drinks, alternative meat products (e.g., meatless burger patties, meatless sausages), alternative egg products (e.g., eggless mayo), non-dairy products (e.g., non-dairy whipped toppings, non-dairy milk, non-dairy creamer, non-dairy milk shakes, nondiary ice cream), energy bars, infant formula, baby foods, cereals, baked goods, edamame, tofu, and tempeh. Plants, seed compositions, oil compositions, and sugar compositions provided herein can be suitable for use in a variety of industrial materials, such as fuel (e.g., biodiesel), soaps, candles, cosmetic products, paints, and protective coatings.

[0271] Plant parts (e.g., seeds) and plant products (e.g., plant biomass, seed compositions, protein compositions, food and / or beverage products) produced by the methods provided herein can be meant for consumption by agricultural animals or for use as feed in an agriculture or aquaculture system. In specific embodiments, plant parts and plant products produced according to the methods provided herein include animal feed (e.g., roughages - forage, hay, silage; concentrates - cereal grains, soybean cake) intended for consumption by bovine, porcine, poultry, lambs, goats, or any other agricultural animal. In some embodiments, plant parts and plant products produced according to the methods include aquaculture feed for any type of fish or aquatic animal in a farmed or wild environment including, without limitation, trout, carp, catfish, salmon, tilapia, crab, lobster, shrimp, oysters, clams, mussels, and scallops.

[0272] Seeds of the present disclosure include a representative sample of seeds, from a plant produced by the methods provided herein. A plant or plant part of the present disclosure can be a crop plant, a forage plant, or part of a crop plant or forage plant. E. Transformation of Plants

[0273] Provided herein are methods for transforming plants or plant parts by introducing into the plants or plant parts one or more mutations (e.g., insertions, substitutions, and / or deletions) to the uORF (e.g., a SIRT element) of at least one bZIP transcription factor gene. The methods can comprise introducing a system (e.g., a gene editing system), reagents (e.g., editing reagents), or a construct for introducing mutations at the target site of interest. The methods can also comprise introducing a construct containing a transgene (e.g., encoding a mutated bZIP SIRT element, a mutated bZIP uORF comprising a SIRT element, or functional fragments thereof) into the plant or plant part.

[0274] The term “transform” or “transformation” as used herein refers to any method used to introduce genetic mutations (e.g., insertions, substitutions, or deletions in the genome), polypeptides, or polynucleotides into plant cells. For purpose of the present disclosure, the transformation can be “stable transformation”, wherein the one or more mutations (e.g., in the uORF of a bZIP transcription factor, e.g., the SIRT element of the bZIP transcription factor) or the transformation constructs (e.g., a construct comprising a nucleic acid molecule encoding a gRNA and / or a nuclease for use in the methods of the present invention, or a construct comprising are introduced into a host (e.g., a host plant, plant part, plant cell, etc.), integrate into the genome of the host, and are capable of being inherited by the progeny thereof; or “transient transformation”, wherein the one or more mutations (e.g., in the uORF of a bZIP transcription factor, e.g., the SIRT element of the bZIP transcription factor) or the transformation constructs (e.g., a construct comprising a gRNA and / or a gene encoding a nuclease for use in the methods of the present invention, or a construct comprising a transgene (e.g., encoding a mutated bZIP SIRT element, a mutated bZIP uORF comprising a SIRT element, or functional fragments thereof) and expressed temporarily. The methods disclosed herein can also be used for insertion of heterologous genes and / or modification of native plant gene expression to achieve desirable plant traits, e.g., increased fatty acid / oil and / or sugar content.

[0275] Any mutation or any polynucleotide of interest (e.g., editing reagents, e.g., a nuclease and a guide RNA; a polynucleotide encoding a mutated bZIP SIRT element, a mutated bZIP uORF comprising a SIRT element, or functional fragment thereof) can be introduced into a plant cell, organelle, or plant embryo by a variety of means of transformation, including microinjection (Crossway et al. (1986) Biotechniques 4:320-334), electroporation (Riggs el al. (1986) Proc. Natl. Acad. Sci. USA 83:5602-5606, Agrobacleritim-m dx&i transformation (U.S. Patent No. 5,563,055 and U.S. Patent No. 5,981,840), direct gene transfer (Paszkowski et al. (198 A) EMB O J. 3:2717- 2722), and ballistic particle acceleration [see, for example, U.S. Patent Nos. 4,945,050; U.S. Patent No. 5,879,918; U.S. Patent No. 5,886,244; and, 5,932,782; Tomes et al. (1995) in Plant Cell, Tissue, and Organ Culture: Fundamental Methods, ed. Gamborg and Phillips (Springer-Verlag, Berlin); McCabe et al. (1988) Biotechnology 6:923-926]; and Lecl transformation (WO 00 / 28058). Also see Weissinger et al. (1988) Ann. Rev. Genet. 22:421-477; Sanford et al. (1987) Particulate Science and Technology 5:27-37 (onion); Christou et al. (1988) Plant Physiol. 87:671-674 (soybean); McCabe et al. (1988) Bio / Technology 6:923-926 (soybean); Finer and McMullen (1991) In Vitro Cell Dev. Biol. 27P: 175-182 (soybean); Singh et al. (1998) Theor. Appl. Genet. 96:319-324 (soybean); Datta et al. (1990) Biotechnology 8:736-740 (rice); Klein et al. (1988) Proc. Natl. Acad. Sci. USA 85:4305-4309 (maize); Klein et al. (1988) Biotechnology 6:559-563 (maize); U.S. Patent Nos. 5,240,855; 5,322,783; and, 5,324,646; Klein et al. (1988) Plant Physiol. 91 :440-444 (maize); Fromm et al. (1990) Biotechnology 8:833-839 (maize); Hooykaas-Van Slogteren et al. (1984) Nature (London) 311 :763-764; U.S. Patent No. 5,736,369 (cereals); Bytebier et al. (1981) Proc. Natl. Acad. Sci. USA 84:5345-5349 (Liliaceae),' De Wet et al. (1985) in The Experimental Manipulation of Ovule Tissues, ed. Chapman et al. (Longman, New York), pp. 197-209 (pollen); Kaeppler et al. (1990) Plant Cell Reports 9:415-418 and Kaeppler et al. (1992) Theor. Appl. Genet. 84:560-566 (whisker-mediated transformation); D’Halluin et al. (1992) Plant Cell 4: 1495-1505 (electroporation); Li et al. (1993) Plant Cell Reports 12:250-255 and Christou and Ford (1995) Annals of Botany 75:407-413 (rice); Osjoda et al. (1996) Nature Biotechnology 14:745-750 (maize via Agrobacterium lumefaciens)\ all of which are herein incorporated by reference.

[0276] The embodiments disclosed herein are not limited to certain methods of introducing nucleic acids into a plant, and are not limited to certain forms or structures that the introduced nucleic acids take. Any method of transforming a cell of a plant described herein with nucleic acids are incorporated into the teachings of this innovation. Agrobacterium-avA biolistic-mediated transformation remain the two predominantly employed approaches. However, one of ordinary skill in the art will realize that the use of particle bombardment (e.g. using a gene-gun), infection by other bacterial species capable of transferring DNA into plants (e.g., Ochrobactrum sp., Ensifer sp., Rhizobium sp.) or virus (e.g., Caulimoriviruses, Geminiviruses, RNA plant viruses) optionally with Agrobacterium infection, transfection, microinjection, electroporation, microprojection, electroporation, silica / carbon fibers, ultrasound mediated, PEG mediated, calcium phosphate coprecipitation, polycation DMSO technique, DEAE dextran procedure, liposome mediated and other techniques can be used to deliver nucleic acid sequences into a plant described herein. Methods disclosed herein are not limited to any size of nucleic acid sequences that are introduced, and thus one could introduce a nucleic acid comprising a single nucleotide (e.g. an insertion) into a nucleic acid of the plant and still be within the teachings described herein. Nucleic acids introduced in substantially any useful form, for example, on supernumerary chromosomes (e.g. B chromosomes), plasmids, vector constructs, additional genomic chromosomes (e.g. substitution lines), and other forms is also anticipated. It is envisioned that new methods of introducing nucleic acids into plants and new forms or structures of nucleic acids will be discovered and yet fall within the scope of the claimed invention when used with the teachings described herein.

[0277] More than one polynucleotides of interest can be introduced into the plant, plant cell, plant organelle, or plant embryo simultaneously or sequentially. For example, different editing reagents, e.g., nuclease polypeptides (or encoding nucleic acid), guide RNAs (or DNA molecules encoding the guide RNAs), donor polynucleotide(s), and / or repair templates can be introduced into the plant cell, organelle, or plant embryo simultaneously or sequentially. The amount or ratio of more than one polynucleotides of interest, or molecules encoded therein, can be adjusted by adjusting the amount or concentration of the polynucleotides and / or timing and dosage of introducing the polynucleotides into the plant or plant part. For example, the ratio of the nuclease (or encoding nucleic acid) to the guide RNA(s) (or encoding DNA) to be introduced into plants or plant parts generally will be about stoichiometric such that the two components can form an RNA-protein complex with the target DNA. In one embodiment, DNA encoding a nuclease and DNA encoding a guide RNA are delivered together within a plasmid vector.

[0278] Alteration of the level or activity of the uORF or the SIRT element of a bZIP transcription factor in plants, plant parts, or plant cells may also be achieved through the use of transposable element technologies to alter gene expression. It is well understood that transposable elements can alter the expression of nearby DNA (McGinnis et al. (1983) Cell 34:75-84). Alteration of the level or activity of the uORF or the SIRT element of a bZIP transcription factor may be achieved by inserting a transposable element into the uORF or the SIRT element of a bZIP transcription factor.

[0279] The cells that have been transformed may be grown into plants (i.e., cultured) in accordance with conventional ways. See, for example, McCormick et al. (1986) Plant Cell Reports 5:81-84. In this manner, the present invention provides transformed plants or plant parts, transformed seed (also referred to as “transgenic seed”) or transformed plant progenies having a nucleic acid modification stably incorporated into their genome.

[0280] The present invention may be used for transformation of any plant species, e.g., both monocots and dicots (including legumes). Plants or plant parts to be transformed according to the methods disclosed herein can be a legume, i.e., a plant belonging to the family Fabaceae (or Leguminosae), or a part (e.g., fruit or seed) of such a plant. When used as a dry grain, the seed of a legume is also called a pulse. Examples of legume include, without limitation, soybean (Glycine max), beans (Phaseolus spp., Cigna spp.), common bean (Phaseolus vulgaris), mung bean (Cigna radiata), cowpea (Cigna unguiculata), adzuki bean Vigna angularis), fava bean (Vicia faba , pea Pisum sativum), chickpea (Cicer arietinum), peanut (Arachis hypogaea), lentils (Lens culinaris, Lens esculenta), lupins (Lupinus spp.), white lupin (Lupinus albus), mesquite (Prosopis spp.), carob (Ceratonia sHiqua , tamarind (Tamarindus indica), alfalfa (Medicago sativa), barrel medic (Medicago truncatula), birdsfood trefoil (Lotus japonicus), licorice (Glycyrrhiza glabra), and clover Trifolium spp.). In specific embodiments, a plant or plant part to be transformed according to the methods of the present disclosure is Glycine max or a part of Glycine max. Additionally, a plant or plant part to be transformed according to the methods present disclosure can be a crop plant or part of a crop plant, including legumes. Examples of crop plants include, but are not limited to, corn (Zea mays'), Brassica sp. (e.g., B. napus, B. rapa, B.juncea), particularly those Brassica species useful as sources of seed oil, alfalfa (Medicago sativa), rice (Oryza sativa), rye (Secale cereale), sorghum (Sorghum bicolor, Sorghum vulgare), camelina (Camelina sativa), millet (e.g., pearl millet (Pennisetum glaucum), proso millet (Panicum miliaceum), foxtail millet (Setaria italica), finger millet Eleusine coracana)), sunflower (Helianthus annuus), quinoa (Chenopodium quinoa), chicory (Cichorium intybus), lettuce (Lactuca sativa), safflower (Carthamus tinctorius), wheat (Triticum aestivum), soybean (Glycine max), tobacco (Nicotiana spp., e.g., Nicotiana tabacum, Nicotiana sylvestris), potato (Solanum tuberosum), tomato (Solanum lycopersicum), peanuts (Arachis hypogaea), cotton (Gossypium barbadense, Gossypium hirsutum), sweet potato (Ipomoea batatus), cassava (Manihot esculenta), coffee (Coffea spp.), coconut (Cocos nucifera), pineapple (Ananas comosus), citrus trees (Citrus spp.), cocoa (Theobroma cacao), tea (Camellia sinensis), banana (Musa spp.), avocado (Persea americana), fig (Ficus casica), guava (Psidium guajava), mango (Mangifera indica), grapes (Vitis vinifera, Vitis riparia), olive (Olea europaea), papaya (Carica papaya), cashew (Anacardium occidentale), macadamia (Macadamia integrifolia), almond (Prunus amygdalus), sugar beets (Beta vulgaris), sugarcane (Saccharum spp.), oil palm (Elaeis guineensis), poplar (Populus spp.), pea Pisum sativum), eucalyptus (Eucalyptus spp.), oats (Avena sativa), barley (Hordeum vulgare), vegetables, ornamentals, and conifers. Additionally, a plant or plant part of the present disclosure can be an oilseed plant (e.g., canola (Brassica napus , cotton (Gossypium sp.), camelina (Camelina sativa) and sunflower (Helianthus sp.)), or other species including wheat (Triticum sp., such as Triticum aestivum L. ssp. Aestivum (common or bread wheat), other subspecies of Triticum aestivum, Triticum turgidum L. ssp. Durum (durum wheat, also known as macaroni or hard wheat), Triticum monococcum L.ssp. monococcum (cultivated einkorn or small spelt), Triticum timopheevi ssp. Timopheevi, Triticum turgigum L. ssp. Dicoccon (cultivated emmer), and other subspecies of Triticum turgidum (Feldman)), barley (Hordeum vulgare), maize Zea mays), oats (Avena sativa), or hemp (Cannabis sativa). Additionally, a plant or plant part of the present disclosure can be a forage plant or part of a forage plant. Examples of forage plants include legumes and crop plants described herein as well as grass forages including Agrostis spp., Lolium spp., Festuca spp., Poa spp., and Bromus spp.

[0281] The present disclosure provides plants and plant parts transformed according to the methods of the present disclosure. Transformed plant parts of the invention include plant cells, plant protoplasts, plant cell tissue cultures from which plants can be regenerated, plant calli, plant clumps, and plant cells that are intact in plants or parts of plants such as embryos, pollen, ovules, seeds, grains, leaves, flowers, branches, fruit, kernels, ears, cobs, husks, stalks, roots, root tips, anthers, and the like. Progeny, variants, and mutants of the regenerated plants are also included within the scope of the disclosure, provided that these parts comprise the introduced mutations, polynucleotides, or polypeptides.

[0282] F. Breeding of Plants

[0283] Also disclosed herein are methods for breeding a plant, such as a soybean plant that has been modified to contain one or more mutations in the bZIP SIRT upstream regulatory region, or to contain a polynucleotide containing, e.g., a gRNA specific to a bZIP SIRT element or a polynucleotide comprising an edited bZIP SIRT element. A plant containing the one or more mutations or the polynucleotide of the present disclosure may be regenerated from a plant cell or plant part, wherein the genome of the plant cell or plant part is genetically-modified to contain the one or more mutations or the polynucleotide of the present disclosure. Using conventional breeding techniques or self-pollination, one or more seeds may be produced from the plant that contains the one or more mutations or the polynucleotide of the present disclosure. Such a seed, and the resulting progeny plant grown from such a seed, may contain the one or more mutations or the polynucleotide of the present disclosure, and therefore may be transgenic. Progeny plants are plants having a genetic modification to contain the one or more mutations or the polynucleotide of the present disclosure, which descended from the original plant having modification to contain the one or more mutations or the polynucleotide of the present disclosure. Seeds produced using such a plant of the invention can be harvested and used to grow generations of plants having genetic modification to contain the one or more mutations or the polynucleotide of the present disclosure, e.g., progeny plants, of the invention, comprising the polynucleotide and optionally expressing a gene of agronomic interest (e.g., herbicide resistance gene). Descriptions of breeding methods that are commonly used for different crops can be found in one of several reference books, see, e.g., Allard, Principles of Plant Breeding, John Wiley & Sons, NY, U. of CA, Davis, Calif., 50-98 (1960); Simmonds, Principles of Crop Improvement, Longman, Inc., NY, 369-399 (1979); Sneep and Hendriksen, Plant breeding Perspectives, Wageningen (ed), Center for Agricultural Publishing and Documentation (1979); Fehr, Soybeans: Improvement, Production and Uses, 2ndEdition, Monograph, 16:249 (1987); Fehr, Principles of Variety Development, Theory and Technique, (Vol.

[0284] 1) and Crop Species Soybean (Vol. 2), Iowa State Univ., Macmillan Pub. Co., NY, 360-376 (1987).

[0285] V. Method of Selecting a Plant with Increased fatty acid, Oil, and / or Sugar Content

[0286] In some aspects, the present disclosure provides methods of selecting a plant with increased fatty acid, oil, and / or sugar content. The methods can comprise the steps of: (i) introducing into a plant cell gene editing reagents (e.g., a nuclease, a guide RNA), which can target an uORF, e.g., a SIRT element, of a basic region / leucine zipper motif (bZIP) transcription factor gene or homologs thereof to generate at least one mutated SIRT elements; (ii) making DNA constructs each comprising, in operable linkage: (a) a promoter that is functional in a plant cell; and (b) a nucleic acid sequence encoding one of the mutated SIRT elements; (iii) transforming plants with the DNA constructs; (iv) evaluating a function of the modified SIRT elements in the transformed plants; and (v) selecting a plant with increased bZIP transcription factor activity or increased fatty acid / oil content. The evaluating step can comprise: quantifying an expression of a reporter gene linked to the modified SIRT element; quantifying, in the absence and the presence of sucrose, expression of one or more genes regulated by the bZIP transcription factor gene; and / or quantifying total fatty acid, oil, and / or sugar (e.g., sucrose) content in the plant or plant part (e.g., seeds). In some embodiments, the one or more genes regulated by the bZIP transcription factor gene regulate carbohydrate transport or metabolism, and comprise one or more of sugar transporter (e.g., encoded by Glyma.10G217900, Glyma.02G075000, Glyma.02G218600, Glyma.l6G157100, Glyma.16G 156800 , Glyma.02G124100, Glyma.01G067600, or homolog of any thereof), sucrose transporter, cell wall invertase (e.g., encoded by Glyma.13G349300, Glyma.15G024600, Glyma.14G096600, Glyma.16G 175800, or homolog of any thereof), asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, and trehalose-6-phosphate phosphatase (TPP6).

[0287] VI. Nucleic Acid Molecules, Constructs, and Cells Comprising a Mutated Upstream Open Reading Frame of bZIP Transcription Factor Gene

[0288] A. Nucleic acid molecules

[0289] Nucleic acid molecules are provided herein comprising a polynucleotide sequence that alters (e.g., increases) bZIP transcription factor activity in a plant or plant part. The nucleic acid molecule can comprise any nucleic acid sequence that alters (e.g., increases) bZIP transcription factor activity in a plant or plant part including those described herein, e.g., an altered (e.g., mutated) nucleic acid sequence of the uORF, e.g., SIRT element, of a bZIP transcription factor gene, or an altered nucleic acid sequence of bZIP transcription factor gene or transcript. Such nucleic acid molecules may be present in, or obtained from, a plant cell, plant part, or plant of the present disclosure, or may be obtained by the methods described herein, e.g., by introducing one or more mutations into the bZIP transcription factor gene or the uORF, e.g., SIRT element thereof, or introducing editing reagents targeting a site of interest in the bZIP transcription factor gene or the uORF, e.g., SIRT element thereof. The nucleic acid molecule described herein can contain a modified regulatory region (e.g., uORF, SIRT element) of a bZIP transcription factor gene that decreases level or activity of the bZIP SIRT element, or increases level or activity of the bZIP transcription factor or a downstream gene operably linked in lieu of the bZIP transcription factor. The increased bZIP transcription factor level or activity can result in the increased level or activity of downstream molecules regulated by the bZIP transcription factor, e.g., molecules that regulate carbohydrate transport or metabolism, e.g., sugar transporter, sucrose transporter, cell wall invertase, ASN, PDH, SPS, SPP, calmodulin, and TPP6.

[0290] The nucleic acid molecule provided herein can comprise a sequence of a mutated bZIP uORF, e.g., SIRT element, containing one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) insertions, substitutions, and / or deletions that increases the bZIP transcription factor activity.

[0291] In some embodiments, the nucleic acid molecule comprises a nucleic acid molecule comprising a nucleic acid sequence of a mutated uORF of a bZIP transcription factor gene, wherein said bZIP transcription factor gene comprises any one of SEQ ID NOs: 1-6. In some embodiments, the nucleic acid molecule comprises a nucleic acid molecule comprising a nucleic acid sequence of a mutated SIRT element, wherein said nucleic acid sequence comprises any one of SEQ ID NOs: 13-18 comprising one or more insertions, substitutions, or deletions therein. For example, the nucleic acid sequence of the 5’ UTR comprising the mutated uORF or the mutated SIRT element provided herein can comprise any one of SEQ ID NOs: 30-39 and 54-62. The nucleic acid molecule can increase level or activity of an operably-linked polynucleotide of interest compared to a control nucleic acid molecule without said mutation.

[0292] The nucleic acid molecule may comprise an in-frame mutation, a frameshift (out-of-frame) mutation, a missense mutation, or a nonsense mutation of the bZIP SIRT element. The nucleic acid molecule described herein can comprise a mutated SIRT element as well as the other region of the upstream regulatory region of the bZIP transcription factor gene or exon / intron region of the bZIP transcription factor gene. B. DNA constructs, vectors, and cells

[0293] The nucleic acid molecules encoding the mutated uORF (e.g., SIRT element) of a bZIP transcription factor gene, or the mutated bZIP transcription factor gene, provided herein can be assembled within a DNA construct with an operably-linked promoter and / or an operably-linked polynucleotide of interest. When transiently or stably transformed with such DNA construct, a plant, plant part, or plant cell can have increased level or activity of the operably-linked polynucleotide of interest as compared to a control plant, plant part, or plant cell (e.g., that is not transformed with the DNA construct, or that is transformed with a DNA construct comprising, in operable linkage, a promoter, a control (e.g., wild-type) bZIP uORF (e.g., SIRT element)). For example, the nucleic acid molecules described herein can be provided in expression cassettes or expression constructs along with a promoter sequence of interest, a native or heterologous promoter sequence, for expression in the plant of interest. By “heterologous promoter sequence” is intended a sequence that is not naturally operably linked with the nucleic acid molecule of interest. For instance, a 2x35s promoter, a native promoter, or a promoter (native or heterologous) comprising an exogenous or synthetic motif sequence may be operably linked to the nucleic acid sequences comprising a mutated bZIP uORF (e.g., SIRT element) and a polynucleotide of interest. The promoter, the mutated bZIP uORF (e.g., SIRT element), and the polynucleotide of interest may each be homologous, native, heterologous, or foreign to the plant host. It is recognized that the heterologous promoter may also drive expression of its homologous or native nucleic acid sequence. In this case, the transformed plant will have a change in phenotype.

[0294] Accordingly, the present disclosure provides DNA constructs comprising, in operable linkage, a uORF of a bZIP transcription factor gene that can be native (without mutation, e.g., comprising the sequence of any one of SEQ ID NOs: 13-18) or mutated (e.g., comprising the sequence of any one of SEQ ID NOs: 13-18 with one or more insertions, substitutions, or deletions therein, e.g., the 5’ UTR comprising the mutated SIRT comprising the sequence of any one of SEQ ID NOs: 30-39 and 54-62), and a polynucleotide of interest (e.g., a bZIP transcription factor gene or a reporter gene, e.g., GFP, luciferase, HA tag). The DNA construct can comprise, in operable linkage, a promoter and / or a reporter / selectable marker construct (e.g., GFP, luciferase, HA tag). Any reporter or selectable marker can be used, including the reporters and selectable markers described elsewhere in the present disclosure.

[0295] Provided herein are vectors comprising the nucleic acid molecule and / or the DNA construct of the present disclosure comprising an altered or native nucleic acid sequence of the uORF (e.g., SIRT element) of the bZIP transcription factor gene. Any vectors can be used, including the vectors described elsewhere in the present disclosure. Also provided herein are cells comprising the nucleic acid molecule, the DNA construct, and / or the vector of the present disclosure comprising an altered nucleic acid sequence of the uORF (e.g., SIRT element) of the bZIP transcription factor gene. The cell can be a plant cell, a bacterial cell, and a fungal cell. The cell can be a bacterium, e.g., an Agrobacterium lumefaciens. containing the nucleic acid molecule, the DNA construct, or the vector of the present disclosure. The cell can be a plant cell. The cells of the present disclosure may be grown, or have been grown, in a cell culture.

[0296] Also provided herein are methods for generating a plant, plant part (e.g., seed), plant cell, or a population of plants or plant parts (e.g., seeds) comprising decreased bZIP SIRT level or activity, increased bZIP transcription factor level or activity, increased level or activity of one or more molecules regulated by the bZIP transcription factor (e.g., molecules regulating carbohydrate transport or metabolism, e.g., .sugar transporter, sucrose transporter, cell-wall invertase, ASN, PDH, SPS, SPP, calmodulin, and / or TPP6), and / or increased fatty acid / oil and / or sugar content, by introducing into the plant, plant part, or plant cell the nucleic acid molecule, the DNA construct, the vector, or the cell of the present disclosure. In some embodiments, the nucleic acid molecule, DNA construct, vector, or cell is introduced into the plant by stable transformation. In other embodiments, the nucleic acid molecule, DNA construct, vector, or cell is introduced into the plant by transient transformation. The present disclosure further provides plants, plant parts (seed, juice, pulp, fruit, flowers, nectar, embryos, pollen, ovules, leaves, stems, branches, bark, kernels, ears, cobs, husks, stalks, roots, root tips, anthers, etc.), or plant products (e.g., seed compositions, plant protein, plant protein compositions, plant extract, plant concentrate, plant powder, plant biomass, and food and beverage products) generated by the methods described herein.

[0297] It will be readily apparent to those skilled in the art that other suitable modifications and adaptations of the methods of the invention described herein are obvious and may be made using suitable equivalents without departing from the scope of the invention or the embodiments disclosed herein. Having now described the invention in detail, the same will be more clearly understood by reference to the following examples, which are included for purposes of illustration only and are not intended to be limiting. Unless otherwise noted, all parts and percentages are by dry weight. EXAMPLES

[0298] EXAMPLE 1: Expression of bZIP transcription factor gene copies in wild-type soybean tissues

[0299] Transcript expression levels of two bZIPl copies in soybean, GmbZIP 123 (Glyma.06G010200), GmbZIP 125 (Glyma.04G010300), GmZIP124 (Glyma.12G040600), GmZIP 126 Glyma.11G 114800), GmZIP18 (14G071400), and GmZIP17 (Glyma.17G253200) in various tissues of two varieties of soybean were studied based on data available from a soy expression database. As shown in FIGs. 1-6, GmbZIP 123, GmbZIP 125, GmbZIP 124, GmbZIP 126, GmbZIP 18, and GmbZIP 17 were expressed throughout various tissues of soybean, with highest expression in the seed pot and seed coat. As shown in FIG. 6, GmbZIP 17 was also highly expressed in the nodule.

[0300] EXAMPLE 2: Modification of the SIRT element in the uORF of bZIP transcription factor gene and effect on expression of operably-linked gene of interest

[0301] Guide RNAs targeting the uORF of GmZIP123, GmbZIP 125, GmZIP124, GmZIP126, GmZIP18, and / or GmZIP17 at or near the SIRT element were designed according to standard methods of the art (Zetsche et al., Cell, Volume 163, Issue 3, Pages 759-771, 2015; Cui et al., Interdisciplinary Sciences: Computational Life Sciences, volume 10, pages 455-465, 2018). Optimized gRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9 and CRISPR-Casl2a have been extensively characterized (Nat Biotechnol 2016;34: 184-191, doi: 10.1038 / nbt.3437). The CRISPR-Casl2a system described herein can be employed for targeting PAM sites such as TTN, TTV, TTTV, NTTV, TATV, TATG, TATA, YTTN, GTTA, and GTTC, utilizing corresponding gRNAs.

[0302] Soybean protoplasts were transformed with constructs comprising the guide RNAs targeting a genomic site in the uORF of GmZIP 123, GmbZIP 125, GmZIP124, GmZIP126, GmZIP18, and / or GmZIP17 at or near the SIRT element and a nuclease using Agrobacterium transformation. Amplicons are produced near the target sites, and are sequenced to detect mutations. A mutated read is recorded for any sequence with more than two reads containing a deletion at the predicted cleavage site. Editing efficiency is calculated based on the percentage of mutated reads to total aligned reads using next generation sequencing (NGS).

[0303] A number of mutants having mutation at least partially in the SIRT element in the uORF of GmZIP 123, GmbZIP 125, GmZIP 124, GmZIP 126, GmZIP 18, and / or GmZIP 17 were generated by introducing into protoplasts the gene editing reagents provided herein, including one or more guide RNAs. The tested guide RNAs include GmbZIP 123 gRNA182, GmbZIP 123 gRNA183, GmbZIP 125 gRNA207, GmbZIP125 gRNA206, GmbZIP124 gRNA97, GmbZIP126 gRNA156, GmbZIP18 gRNA91, and GmbZIP17 gRNA73.

[0304] The mutants having mutation at least partially in the SIRT element in the uORF of GmZIP 123, GmbZIP 125, GmZIP124, GmZIP126, GmZIP18, and / or GmZIP17 were screened for editing efficiency and expression levels. Expression cassettes comprising GmZIP 123, GmbZIP 125, GmZIP 124, GmZIP 126, GmZIP 18, and / or GmZIP 175’ UTR comprising the mutated SIRT element, operably linked to a functional promoter and a downstream polynucleotide encoding firefly luciferase were generated. The cassettes with mutations, as well as no mutations (wild-type) were transiently transfected in soybean protoplasts. Expression levels of firefly luciferase in protoplasts were measured, as an indicator of transcription initiation activity of the bZIP 5’ UTR with the SIRT element.

[0305] As shown in FIG. 7, in soybean protoplasts transfected with constructs containing GmbZIP 123 5’ UTR including a mutated SIRT (182-1 Obp, 182-1 Ibp, 183-10bp, 183-8bp, or 183- 7bp), expression of firefly luciferase was increased to about 1.5 to about 3 times as compared to protoplasts transfected with constructs containing wild-type GmbZIP 123 5’ UTR (WT). Similarly, as shown in FIG. 8, in soybean protoplasts transfected with constructs containing GmbZIP 125 5’ UTR with a mutated SIRT element (206-10bp, 206-1 Ibp, 207-8bp, or 207-7bp), expression of firefly luciferase was increased to about 1.5 to about 3 times as compared to protoplasts transfected with constructs containing wild-type GmbZIP 125 5’ UTR (WT). As shown in FIG. 9, in soybean protoplasts that have been transfected with a construct containing GmbZIP 123 5’ UTR with a mutated SIRT element (183-16bp, 183-1 Ibp, 183-7bp, 207-10bp), expression of firefly luciferase was increased to about 1.8 to about 3 times as compared to protoplasts transfected with constructs containing wild-type GmbZIP 123 5’ UTR (WT). As shown in FIG. 10, in soybean protoplasts that have been transfected with a construct containing GmbZIP 125 5’ UTR with a mutated SIRT element (207-19bp, 207-7bp, 207-5bp, 207-1 Obp), expression of firefly luciferase was increased to about 2.2 to about 3.3 times as compared to protoplasts transfected with constructs containing wildtype GmbZIP 125 5’ UTR (WT). As shown in FIG. 11, in soybean protoplasts that have been transfected with a construct containing GmbZIP 175’ UTR with a mutated SIRT element (73-5bp, 73-7bpA, 73-7bpB, 73-25bp), expression of firefly luciferase was increased to about 3.5 to about 6.5 times as compared to protoplasts transfected with constructs containing wild-type GmbZIP175’ UTR (WT). The results indicate that a mutation (e.g., a deletion) in the SIRT element in a bZIP 5’ UTR can increase level or activity of the bZIP or an otherwise operably-linked polynucleotide of interest. Expression cassettes comprising GmZIP123, GmbZIP125, GmZIP124, GmZIP126, GmZIP 18, and / or GmZIP17 5’ UTR comprising the mutated SIRT element, operably linked to a functional promoter and other downstream polynucleotides of interest (e.g., encoding bZIP, HA- tagged bZIP, reporter (e.g., GFP)) are also generated, and analyzed similarly. Expression levels of an operably linked reporter gene (e.g., GFP, luciferase, HA-tag) are measured by a reporter assay, e.g., visualization of GFP or a luciferase assay, as described for example by Thalor et al. 2012 PLoS One 7(3): e33111. Expression levels of the polynucleotide of interest are measured by standard methods for measuring mRNA levels of a gene, including quantitative RT-PCR, northern blot, and serial analysis of gene expression (SAGE). Levels of the protein encoded by the polynucleotide of interest are also measured by standard methods for measuring protein levels, including western blot analysis, ELISA, or dot blot analysis of a plant sample using an antibody directed to the protein encoded by the polynucleotide of interest.

[0306] Mutant SIRT / bZIP constructs described above are further tested in soybean plants to study whether the SIRT mutations that remove sucrose repression of bZIP transcription factor or otherwise enhance bZIP transcription factor activity modify level or activity of targets downstream of bZIP and / or increase seed fatty acid / oil and / or sugar content. Three (3) to 4 week old soybean plants are infiltrated with the mutant SIRT / bZIP constructs (expression cassettes driven by native promoter with independent synthetic deletions in SIRT element upstream of HA-tagged bZIP) described above, to transiently express the bZIP / SIRT cassettes. At day 3 post-infiltration, sampling is conducted and plants are tested for mutations.

[0307] Levels of mRNA and / or protein expression of bZIP downstream targets, e.g., sugar transporter, sucrose transporter, cell-wall invertase, asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, and / or trehalose-6-phosphate phosphatase (TPP6) is tested in soybean leaves using standard methods (e.g., quantitative RT-PCR, northern blot, SAGE, western blot analysis, ELISA, dot blot analysis), to determine if molecules responsible for sucrose transport and / or metabolism are affected by the mutations introduced to the SIRT element. Activities of the bZIP downstream targets involved in carbohydrate transport (e.g., sugar transporter, sucrose transporter, cell-wall invertase) can be measured by a standard carbohydrate (e.g., sugar) transport assay, for example by labeling the plants with13CO2 and monitoring13C enrichment levels of sucrose, fructose, and glucose in plant organs of interest, e.g., leaves, seeds, and siliques. Activity of a sugar transporter can also be measured by using a biosensor that detects conformational changes of the transporter during sugar transport. Activities of the bZIP downstream targets involved in carbohydrate (e.g., sugar) transport or metabolism (e.g., cell wall invertase, ASN, PDH, SPS, SPP, TPP6) can be measured by enzymatic assays of the respective enzymes. For example, activity of a cell wall invertase can be measured for example as described in Tomlinson et al. 2004 J. Exp. Botany 55, 2291-2303, the entire content of which is incorporated by reference herein. Activity of calmodulin can be determined by a Ca2+-calmodulin binding assay or measuring activities of Camdependent phosphodiesterase or calcium-calmodulin dependent protein kinase (CaMKII) (Bossuyt & Bers 2013 J. Mol. Med. (Berl.) 91(8):907-916).

[0308] Seed fatty acid / oil or sugar (e.g., sucrose) content is measured to determine if the mutations introduced to the SIRT element alters seed fatty acid / oil or sugar (e.g., sucrose) content. Amount or levels of oil, total fatty acids, and specific fatty acids can be measured by any methods for measuring oil or fatty acid amount or levels in a plant sample, including NIR, GC-MS optionally with certain modifications (e.g., with or without initial lipid extraction, with or without isotope labeling of analytes), or NMR. Fatty acid composition (e.g., percentage of specific fatty acids normalized to total fatty acids) can be calculated based on the amount or concentration of total fatty acids and specific fatty acids in the sample. Amount or level of sucrose, total sugar, glucose, fructose, galactose, maltose, and / or lactose in seeds can be measured by any methods for measuring total or specific sugar content in a plant sample, including NIR, SPE, SPME, HPLC, GCMS, and / or enzymatic assay.

[0309] EXAMPLE 3: Generation of TO and T1 plants with modified SIRT upstream of bZIP

[0310] Embryonic axes of mature seeds of soybean varieties are stably transformed with constructs comprising one, two, or multiple guide RNAs targeting the uORF of GmZIP 123, GmbZIP 125, GmZIP 124, GmZIP 126, GmZIP 18, and / or GmZIP17 at or near the SIRT element and a nuclease using Agrobacterium transformation. Transformed plants are identified by selective marker (e.g., resistance to an herbicide). Amplicons are produced of the genomic regions near the targeted sites and sequenced to evaluate the presence of the mutation using a pair of primers to detect mutations introduced. Transgenic events are recorded, and the TO plants were assigned unique plant names and are subjected to molecular characterization and propagation. TO plants are self-pollinated and T1 plants are generated. Crosses are made to generate lines that are homozygous or heterozygous for the target mutation and lack the editing reagents.

[0311] Soybean plants containing GmbZIP123 5’ UTR with a mutated SIRT element (183-10 bp) as well as soybean plants containing GmbZIP123 5’ UTR with a mutated SIRT element (183-16 bp) and GmbZIP125 5’ UTR with a mutated SIRT element (207-19 bp) were generated.

[0312] EXAMPLE 4: Screening of plants with mutations

[0313] Transformed plants are screened using a variety of molecular tools to identify plants and genotypes that will result in the expected phenotype. For example, expression levels of the GmZIP 123, GmbZIP 125, GmZIP124, GmZIP126, GmZIP18, and / or GmZIP17 transcription factor gene and levels and activities of the GmZIP 123, GmbZIP 125, GmZIP 124, GmZIP 126, GmZIP 18, and / or GmZIP 17 transcription factor are measured in mutant plants (e.g., having a homozygous or heterozygous mutation in the bZIP transcription factor gene promoter). Expression levels of the GmZIP 123, GmbZIP 125, GmZIP 124, GmZIP 126, GmZIP 18, and / or GmZIP 17 transcription factor are measured by any standard methods for measuring mRNA levels of a gene, including quantitative RT-PCR, northern blot, and serial analysis of gene expression (SAGE). Expression levels of the GmZIP 123, GmbZIP 125, GmZIP 124, GmZIP 126, GmZIP 18, and / or GmZIP 17 transcription factor are measured by any standard methods for measuring protein levels, including western blot analysis, ELISA, or dot blot analysis of a protein sample obtained from the plant using an antibody directed to the bZIP transcription factor.

[0314] Activity of the GmZIP 123, GmbZIP 125, GmZIP 124, GmZIP 126, GmZIP 18, and / or GmZIP 17 transcription factor is assessed by measuring levels and / or activity of the downstream targets, i.e., molecules regulated by GmZIP 123, GmbZIP 125, GmZIP 124, GmZIP 126, GmZIP 18, and / or GmZIP 17, e.g., sugar transporter, sucrose transporter, cell-wall invertase, asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’- phosphate phosphatase (SPP), calmodulin, and trehalose-6-phosphate phosphatase (TPP6). For example, levels of mRNA and / or protein expression of bZIP downstream targets described above can be tested in soybean leaves using standard methods, e.g., quantitative RT-PCR, northern blot, SAGE, western blot, ELISA, or dot blot. Activities of the bZIP downstream targets involved in carbohydrate transport (e.g., sugar transporter, sucrose transporter, cell-wall invertase) can be measured by a standard carbohydrate (e.g., sugar) transport assay, for example by labeling the plants with13CO2 and monitoring13C enrichment levels of sucrose, fructose, and glucose in plant organs of interest, e.g., leaves, seeds, and siliques. Activity of a sugar transporter can also be measured by using a biosensor that detects conformational changes of the transporter during sugar transport. Activities of the bZIP downstream targets involved in carbohydrate (e.g., sugar) transport or metabolism (e.g., cell wall invertase, ASN, PDH, SPS, SPP, TPP6) can be measured by enzymatic assays of the respective enzymes. For example, activity of a cell wall invertase can be measured for example as described in Tomlinson et al. 2004 J. Exp. Botany 55, 2291-2303, the entire content of which is incorporated by reference herein. Activity of calmodulin can be determined by a Ca2+-calmodulin binding assay or measuring activities of Ca2+-dependent phosphodiesterase or calcium-calmodulin dependent protein kinase (CaMKII) (Bossuyt & Bers 2013 J. Mol. Med. (Berl.) 91(8):907-916). Oil and sucrose content was measured by NIR in seeds of the T1 soybean plants with mutation in GmbZIP123 5’ UTR and / or GmbZIP125 5’ UTR SIRT elements described above. As shown in FIG. 12, soybean plants containing GmbZIP123 5’ UTR with a mutated SIRT element (183-10 bp) showed a 1.3% increase in oil content as measured by NIR, and soybean plants containing GmbZIP123 5’ UTR with a mutated SIRT element (183-16 bp) and GmbZIP125 5’ UTR with a mutated SIRT element (207-19 bp) showed a 1.4% increase in oil content as measured by NIR, relative to control plants without mutation (null).

[0315] As shown in FIG. 13, soybean plants containing GmbZIP123 5’ UTR with a mutated SIRT element (183-16 bp) and a mutated GmbZIP125 5’ UTR with a mutated SIRT element (207-19 bp) showed a 0.5% increase in sucrose content as measured by NIR relative to control plants without mutation (null).

[0316] Plants with mutation and a desirable phenotype (e.g., increased level or activity of the bZIP transcription factor, increased level or activity of molecules involved in carbohydrate (e.g., sugar) transport and / or metabolism, increased oil, fatty acid, and / or sugar (e.g., sucrose) content) as compared to a control plant (e.g., without the mutation) when grown under the same environmental conditions, are selected.

[0317] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described in any way.

[0318] It is appreciated that certain features of the disclosure, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the disclosure, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable sub-combination or as suitable in any other described embodiment of the disclosure. Certain features described in the context of various embodiments are not to be considered essential features of those embodiments, unless the embodiment is inoperative without those elements.

[0319] While various aspects of the invention are described herein, it is not intended that the invention be limited by any particular aspect. On the contrary, the invention encompasses various alternatives, modifications, and equivalents, as will be appreciated by those of skill in the art. Furthermore, where feasible, any of the aspects disclosed herein may be combined with each other (e.g., the feature according to one aspect may be added to the features of another aspect or replace an equivalent feature of another aspect) or with features that are well known in the art, unless indicated otherwise by context. TABLE 4. Sequence Table

Claims

What is claimed is:

1. A plant or plant part comprising increased basic region / leucine zipper motif (bZIP) transcription factor activity compared to a control plant or plant part, wherein said plant or plant part comprises a genetic mutation that increases the bZIP transcription factor activity, wherein the mutation comprises one or more insertions, substitutions, or deletions in an upstream open reading frame (uORF) of at least one bZIP transcription factor gene or homolog thereof, wherein the uORF comprises a sucrose induced repression of translation (SIRT) element, wherein said mutation alters level or activity of the SIRT element, wherein level or activity of said bZIP transcription factor gene or homolog thereof, or a bZIP transcription factor encoded by said bZIP transcription factor gene or homolog thereof is increased compared to a control plant or plant part, and wherein carbohydrate transport in said plant or plant part is increased compared to a control plant or plant part.

2. The plant or plant part of claim 1, comprising increased fatty acid, oil, and / or sugar content in said plant or plant part compared to a control plant or plant part.

3. The plant or plant part of claim 2, comprising the oil content that is increased by about 1% or more dry weight and / or the sugar content that is increased by about 0.5% or more dry weight compared to a control plant or plant part.

4. The plant or plant part of any one of claims 1-3, wherein said mutation is located in an uORF of a bZIP transcription factor gene or homolog thereof that:(i) comprises a nucleic acid sequence having at least 80% sequence identity to any one of SEQ ID NOs: 1-6, wherein said nucleic acid sequence encodes a polypeptide that retains bZIP transcription factor activity;(ii) comprises the nucleic acid sequence of any one of SEQ ID NOs: 1-6;(iii) encodes a polypeptide comprising an amino acid sequence having at least 80% sequence identity to an amino acid sequence of any one of SEQ ID NOs: 7-12, wherein said polypeptide retains bZIP transcription factor activity; or(iv) encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs:7-12.

5. The plant or plant part of any one of claims 1-4, wherein said mutation is located at least partially in an SIRT element in said uORF, wherein said SIRT element:(i) comprises a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 13-18, wherein said nucleic acid sequence encodes a polypeptide that retains SIRT activity;(ii) comprises the nucleic acid sequence of any one of SEQ ID NOs: 13-18;(iii) encodes a polypeptide comprising an amino acid sequence having at least 90% sequence identity to an amino acid sequence of any one of SEQ ID NOs: 19-24, wherein said polypeptide retains SIRT activity; or(iv) encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 19-24; and / or wherein a 5’ untranslated region (5’ UTR) that contains the uORF:(v) comprises a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 48-53; or(vi) comprises the nucleic acid sequence of any one of SEQ ID NOs: 48-53, before the mutation is located.

6. The plant or plant part of any one of claims 1-5, comprising:(i) a deletion of one or more nucleotides of SEQ ID NO: 13 in the SIRT element in the uORF of the Glycine max bZIP123 transcription factor gene;(ii) a deletion of one or more nucleotides of SEQ ID NO: 14 in the SIRT element in the uORF of the Glycine max bZIP125 transcription factor gene;(iii) a deletion of one or more nucleotides of SEQ ID NO: 15 in the SIRT element in the uORF of the Glycine max bZIP124 transcription factor gene;(iv) a deletion of one or more nucleotides of SEQ ID NO: 16 in the SIRT element in the uORF of the Glycine max bZIP126 transcription factor gene;(v) a deletion of one or more nucleotides of SEQ ID NO: 17 in the SIRT element in the uORF of the Glycine max bZIP18 transcription factor gene; and / or(vi) a deletion of one or more nucleotides of SEQ ID NO: 18 in the SIRT element in the uORF of the Glycine max bZIP17 transcription factor gene.

7. The plant or plant part of any one of claims 1-6, wherein the mutation is located at least partially in a 5’ end region of said SIRT element and / or the mutation comprises a deletion of 7-25 nucleotides located at least partially in said SIRT element.

8. The plant or plant part of claim 6, comprising a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 30-34 and 54-56, a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 35-39 and 57-58, and / or a mutated 5’ UTR of the Glycine max bZIP17 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 59-62.

9. The plant or plant part of claim 8, comprising (i) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 32, or (ii) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NO: 54 and a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 57.

10. The plant or plant part of any one of claims 1-9, wherein said mutation comprises an out-of-frame mutation of the SIRT element.

11. The plant or plant part of any one of claims 1-9, wherein said mutation comprises a nonsense mutation of the SIRT element.

12. The plant or plant part according to any one of claims 1-11, wherein said plant or plant part comprises 2 or more genes encoding a bZIP transcription factor.

13. The plant or plant part according to claim 12, wherein said 2 or more genes have less than 100% sequence identity to one another.

14. The plant or plant part of any one of claims 1-13, wherein level or activity of one or more molecules regulated by the bZIP transcription factor is modulated, wherein the modulation comprises:(i) an increased baseline level or activity in the absence of sucrose;(ii) a decreased suppression or an increased increase of the level or activity in the presence of sucrose relative to the absence of sucrose; and / or(iii) an increased level or activity in the presence of sucrose, relative to the control plant or plant part.

15. The plant or plant part of claim 14, wherein said one or more molecules regulated by the bZIP transcription factor comprises a sugar transporter and / or a cell-wall invertase.

16. The plant or plant part of claim 15, wherein the sugar transporter is encoded by Glyma.10G217900, Glyma.02G075000, Glyma.02G218600, Glyma.l6G157100,Glyma.16G 156800 , Glyma.02G124100, Glyma.01G067600, or homolog of any thereof, and / or the cell wall invertase is encoded by Glyma.13G349300, Glyma.15G024600, Glyma.14G096600, Glyma.16G 175800, or homolog of any thereof.

17. The plant or plant part of claim 15 or 16, wherein said one or more molecule regulated by the bZIP transcription factor further comprises asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, and / or trehalose-6-phosphate phosphatase (TPP6).

18. The plant or plant part of any one of claims 1-17, wherein said plant or plant part is a legume.

19. The plant or plant part of claim 18, wherein said plant or plant part is selected from the group consisting of soybean (Glycine max), beans (Phaseolus spp.), common bean (Phaseolus vulgaris), fava bean (Vicia faba , mung bean (Vigna radiata), pea (Pisum sativum , chickpea (Cicer arietinum), peanut (Arachis hypogaea), lentils (Lens culinaris, Lens esculenta), lupins (Lupinus spp.), white lupin (Lupinus albus), mesquite (Prosopis spp.), carob (Cer atonia siliqua), tamarind (Tamarindus indica), alfalfa (Medicago sativa), barrel medic (Medicago truncatula), birdsfood trefoil (Lotus japonicus), licorice (Glycyrrhiza glabra , and clover (Trifolium spp.).

20. The plant or plant part of claim 19, wherein said plant or plant part is Glycine max.

21. The plant or plant part of any one of claims 1-17, wherein said plant or plant part is selected from the group consisting of corn (Zea mays), Brassica species, Brassica napus, Brassica rapa, Brassica juncea, rice (Oryza sativa), rye (Secale cereale), sorghum (Sorghum bicolor, Sorghum vulgare), millet, pearl millet (Pennisetum glaucum), proso millet (Panicum miliaceum), foxtail millet (Setaria italica), finger millet (Eleusine coracana), sunflower (Helianthus annuus), safflower (Carthamus tinctorius), wheat (Triticum aestivum), tobacco (Nicotiana tabacum), potato (Solanum tuberosum), peanuts (Arachis hypogaea), cotton (Gossypium barbadense, Gossypium hirsutum), sweet potato (Ipomoea batatus), cassava (Manihot esculenta), coffee (Coffea spp.), coconut (Cocos nucifera), pineapple (Ananas comosus), citrus trees (Citrus spp.), cocoa (Theobroma cacao), tea (Camellia sinensis), banana (Musa spp.), avocado (Per sea americana), fig(Ficus casica), guava (Psidium guajava), mango (Mangifera indica), olive Olea europaea), papaya (Carica papaya), cashew (Anacardium occidentals), macadamia (Macadamia integrifolia), almond (Prunus amygdalus), sugar beets (Beta vulgaris), sugarcane (Saccharum spp.), oats, barley, vegetables, ornamentals, and conifers.

22. The plant or plant part of any one of claims 1-21, wherein said plant or plant part is a seed.

23. A population of plants or plant parts comprising the plant or plant part of any one of claims 1-23, wherein the population comprises increased bZIP transcription factor activity, increased carbohydrate transport, and / or increased fatty acid, oil, and / or sugar content in seeds compared to a control population.

24. The population of plants or plant parts of claim 23, wherein said plant or plant part is a seed, and said population is a population of seeds.

25. A method for increasing basic region / leucine zipper motif (bZIP) transcription factor activity in a plant or plant part, said method comprising introducing a genetic mutation that increases the bZIP transcription factor activity into said plant or plant part, wherein the mutation comprises one or more insertions, substitutions, or deletions in an upstream open reading frame (uORF) of at least one native bZIP transcription factor gene or homolog thereof, wherein the uORF comprises a sucrose induced repression of translation (SIRT) element, wherein said mutation alters level or activity of the SIRT element, wherein said mutation increases level or activity of said bZIP transcription factor gene or homolog thereof, or a bZIP transcription factor encoded by said bZIP transcription factor gene or homolog thereof, and wherein said mutation increases carbohydrate transport in said plant or plant part.

26. The method of claim 25, further comprising introducing the genetic mutation that increases the bZIP transcription factor activity into a plant cell, and regenerating said plant or plant part from said plant cell.

27. The method of claim 25 or 26, wherein fatty acid, oil, and / or sugar content is increased in said plant or plant part compared to a control plant or plant part.

28. The method of claim 27, wherein the oil content is increased by about 1% or more dry weight and / or the sugar content is increased by about 0.5% or more dry weight in said plant or plant part compared to a control plant or plant part.

29. The method of any one of claims 25-28, wherein the mutation is introduced into an uORF of a bZIP transcription factor gene or homolog thereof that:(i) comprises a nucleic acid sequence having at least 80% sequence identity to any one of SEQ ID NOs: 1-6, wherein said nucleic acid sequence encodes a polypeptide that retains bZIP transcription factor activity;(ii) comprises the nucleic acid sequence of any one of SEQ ID NOs: 1-6;(iii) encodes a polypeptide comprising an amino acid sequence having at least 80% sequence identity to an amino acid sequence of any one of SEQ ID NOs: 7-12, wherein said polypeptide retains bZIP transcription factor activity; or(iv) encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 7-12.

30. The method of any one of claims 25-29, wherein the mutation is introduced at least partially in an SIRT element in said uORF, wherein said SIRT element:(i) comprises a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 13-18, wherein said nucleic acid sequence encodes a polypeptide that retains SIRT activity;(ii) comprises the nucleic acid sequence of any one of SEQ ID NOs: 13-18;(iii) encodes a polypeptide comprising an amino acid sequence having at least 90% sequence identity to an amino acid sequence of any one of SEQ ID NOs: 19-24, wherein said polypeptide retains SIRT activity; or(iv) encodes a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 19-24; and / or wherein a 5’ untranslated region (5’ UTR) that contains the uORF:(v) comprises a nucleic acid sequence having at least 90% sequence identity to any one of SEQ ID NOs: 48-53, wherein said nucleic acid sequence encodes a polypeptide that retains SIRT activity; or(vi) comprises the nucleic acid sequence of any one of SEQ ID NOs: 48-53, before the mutation is introduced.

31. The method of any one of claims 25-30, wherein the mutation comprises:(i) a deletion of one or more nucleotides of SEQ ID NO: 13 in the SIRT element in the uORF of the Glycine max bZIP123 transcription factor gene;(ii) a deletion of one or more nucleotides of SEQ ID NO: 14 in the SIRT element in the uORF of the Glycine max bZIP125 transcription factor gene;(iii) a deletion of one or more nucleotides of SEQ ID NO: 15 in the SIRT element in the uORF of the Glycine max bZIP124 transcription factor gene;(iv) a deletion of one or more nucleotides of SEQ ID NO: 16 in the SIRT element in the uORF of the Glycine max bZIP126 transcription factor gene;(v) a deletion of one or more nucleotides of SEQ ID NO: 17 in the SIRT element in the uORF of the Glycine max bZIP18 transcription factor gene; and / or(vi) a deletion of one or more nucleotides of SEQ ID NO: 18 in the SIRT element in the uORF of the Glycine max bZIP17 transcription factor gene.

32. The method of any one of claims 25-31, wherein the mutation is introduced at least partially in a 5’ end region of said SIRT element, and / or the mutation comprises a deletion of 7-25 nucleotides located at least partially in said SIRT element.

33. The method of claim 31, wherein the plant or plant part comprises a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 30-34 and 54-56, a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 35-39 and 57-58, and / or a mutated 5’ UTR of the Glycine max bZIP17 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NOs: 59-62, after the mutation is introduced.

34. The method of claim 33, wherein the plant or plant part comprises (i) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 32, or (ii) a mutated 5’ UTR of the Glycine max bZIP123 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of any one of SEQ ID NO: 54 and a mutated 5’ UTR of the Glycine max bZIP125 transcription factor gene comprising a mutated SIRT element and a nucleic acid sequence of SEQ ID NO: 57, after the mutation is introduced35. The method of any one of claims 25-34, wherein introducing the mutation comprises introducing an out-of-frame mutation into said SIRT element.

36. The method of any one of claims 25-35, further comprising introducing editing reagents or a nucleic acid construct encoding said editing reagents into said plant, plant part, or plant cell.

37. The method of claim 36, wherein said editing reagents comprise at least one nuclease, wherein the nuclease cleaves a target site in said uORF of said at least one native bZIP transcription factor or homolog thereof in said plant, plant part, or plant cell, and said mutation is introduced at said cleaved target site.

38. The method of claim 37, wherein the at least one nuclease comprises a CRISPR nuclease.

39. The method of claim 38, wherein the CRISPR nuclease is a Type II CRISPR system nuclease, a Type V CRISPR system nuclease, a Cas9 nuclease, a Cast 2a (Cpfl) nuclease, or a Cmsl nuclease.

40. The method of claim 39, wherein the CRISPR nuclease is a Casl2a nuclease or an ortholog thereof.

41. The method of any one of claims 36-40, wherein the editing reagents comprise one or more guide RNAs (gRNAs).

42. The method of claim 41, wherein the one or more gRNAs comprise a nucleic acid sequence complementary to a region of a genomic DNA sequence of said uORF or said SIRT element in said at least one native bZIP transcription factor or homolog thereof in said plant or plant part.

43. The method of claim 41, wherein at least one of the one or more gRNAs comprise a nucleic acid sequence encoded by:(a) a nucleic acid sequence that shares at least 80% sequence identity with the nucleic acid sequence of any one of SEQ ID NOs: 25-29; or(b) a nucleic acid sequence of SEQ ID NOs: 25-29.

44. The method of any one of claims 25-43, wherein level or activity of one or more molecules regulated by the bZIP transcription factor in the plant or plant part is modulated, wherein the modulation comprises:(i) an increased baseline level or activity in the absence of sucrose;(ii) a decreased suppression or an increased increase of the level or activity in the presence of sucrose relative to the absence of sucrose; and / or(iii) an increased level or activity in the presence of sucrose, relative to the control plant or plant part.

45. The method of claim 44, wherein said one or more molecules regulated by the bZIP transcription factor comprises a sugar transporter and / or a cell-wall invertase.

46. The method of claim 45, wherein the sugar transporter is encoded by Glyma.10G217900, Glyma.02G075000, Glyma.02G218600, Glyma.l6G157100,Glyma.16G 156800 , Glyma.02G124100, Glyma.01G067600, or homolog of any thereof, and / or the cell wall invertase is encoded by Glyma.13G349300, Glyma.15G024600, Glyma.14G096600, Glyma.16G 175800, or homolog of any thereof.

47. The method of any one of claims 44-46, wherein said one or more molecule regulated by the bZIP transcription factor further comprises asparagine synthase (ASN), proline dehydrogenase (PDH), sucrose phosphate synthase (SPS), sucrose-6’ -phosphate phosphatase (SPP), calmodulin, and / or trehalose-6-phosphate phosphatase (TPP6).

48. The method of any one of claims 25-47, wherein said plant or plant part is a legume.

49. The method of claim 48, wherein said plant or plant part is selected from the group consisting of soybean (Glycine max), beans (Phaseolus spp.), common bean (Phaseolus vulgaris), fava bean (Vicia faba , mung bean (Vigna radiata), pea (Pisum sativum , chickpea (Cicer arietinum), peanut (Arachis hypogaea), lentils (Lens culinaris, Lens esculenta), lupins (Lupinus spp.), white lupin (Lupinus albus), mesquite (Prosopis spp.), carob (Cer atonia siliqua), tamarind (Tamarindus indica), alfalfa (Medicago sativa), barrel medic (Medicago truncatula), birdsfood trefoil (Lotus japonicus), licorice (Glycyrrhiza glabra , and clover (Trifolium spp.).

50. The method of claim 49, wherein said plant or plant part is Glycine max.

51. The method of any one of claims 25-47, wherein said plant or plant part is selected from the group consisting of com (Zea mays), Brassica species, Brassica napus, Brassica rapa, Brassica juncea, rice (Oryza sativa), rye (Secale cereale), sorghum (Sorghum bicolor, Sorghum vulgare), millet, pearl millet (Pennisetum glaucum), proso millet (Panicum miliaceum), foxtail millet (Setaria italica), finger millet (Eleusine coracana), sunflower (Helianthus annuus), safflower (Carthamus tinctorius), wheat (Triticum aestivum), tobacco (Nicotiana tabacum), potato(Solanum tuberosum), peanuts (Arachis hypogaea), cotton (Gossypium barbadense, Gossypium hirsutum), sweet potato (Ipomoea batatus), cassava (Manihot esculenta), coffee (Coffea spp.), coconut (Cocos nucifera), pineapple (Ananas comosus), citrus trees (Citrus spp.), cocoa (Theobroma cacao), tea (Camellia sinensis , banana (Musa spp.), avocado (Per sea americana), fig (Ficus casica), guava (Psidium guajava), mango (Mangifera indica), olive (Olea europaea), papaya (Carica papaya), cashew (Anacardium occidentale), macadamia (Macadamia integrifolia), almond (Prunus amygdalus), sugar beets (Beta vulgaris), sugarcane (Saccharum spp.), oats, barley, vegetables, ornamentals, and conifers.

52. A plant or plant part produced by the method of any one of claims 25-51, wherein said plant or plant part comprises increased bZIP transcription factor activity compared to a control plant or plant part.

53. The plant or plant part of claim 52, comprising increased fatty acid, oil, and / or sugar content in seeds compared to a control plant or plant part.

54. The plant or plant part of claim 52 or 53, wherein said plant or plant part is a seed.

55. A population of plants or plant parts produced by the method of any one of claims 25-54, wherein the population comprises increased bZIP transcription factor activity, increased carbohydrate transport, and / or increased fatty acid, oil, and / or sugar content in seeds compared to a control population.

56. A population of plants or plant parts of claim 55, wherein said plant or plant part is a seed, and said population is a population of seeds.

57. A seed composition produced from the plant or plant part or the population of plants or plant parts of any one of claims 1-24 and 52-56.

58. An oil composition produced from the plant or plant part or the population of plants or plant parts of any one of claims 1-24 and 52-56, or the seed composition of claim 57.

59. A nucleic acid molecule comprising:(i) a nucleic acid sequence of a 5’ untranslated region (5’ UTR) of a bZIP transcription factor gene comprising a mutated sucrose induced repression of translation (SIRT) element, wherein said bZIP transcription factor gene comprises a nucleic acid sequence of any one of SEQ ID NOs: 1-6, or wherein the 5’ UTR comprises the nucleic acid sequence of any one of SEQ ID NOs: 48-53 comprising one or more insertions, substitutions, or deletions therein; or(ii) a nucleic acid sequence of a mutated SIRT element, wherein said nucleic acid sequence comprises any one of SEQ ID NOs: 13-18 comprising one or more insertions, substitutions, or deletions therein, wherein said nucleic acid molecule increases level or activity of an operably -linked polynucleotide of interest compared to a control nucleic acid molecule without said mutation.

60. The nucleic acid molecule of claim 59, wherein the nucleic acid sequence of the 5’ UTR comprising a mutated SIRT element comprises any one of SEQ ID NOs: 30-39 and 54-62.

61. A DNA construct comprising, in operable linkage:(i) a promoter that is functional in a plant cell; (ii) the nucleic acid molecule of claim 55 or 56; and(iii) a polynucleotide of interest.

62. A cell comprising the nucleic acid molecule of claim 59 or 60, or the DNA construct of claim 61.

63. The cell of claim 62, wherein the cell is a plant cell or a bacterial cell.

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