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WO2015169925A1 - Promoteur de plante - Google Patents

Promoteur de plante Download PDF

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Publication number
WO2015169925A1
WO2015169925A1 PCT/EP2015/060117 EP2015060117W WO2015169925A1 WO 2015169925 A1 WO2015169925 A1 WO 2015169925A1 EP 2015060117 W EP2015060117 W EP 2015060117W WO 2015169925 A1 WO2015169925 A1 WO 2015169925A1
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WIPO (PCT)
Prior art keywords
plant
polynucleotide
promoter
interest
plant cell
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English (en)
Inventor
Verena LIEDSCHULTE
Simon GOEPFERT
Lucien Bovet
Nicolas Sierro
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Philip Morris Products SA
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Philip Morris Products SA
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Publication of WO2015169925A1 publication Critical patent/WO2015169925A1/fr
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
    • C12N15/8216—Methods for controlling, regulating or enhancing expression of transgenes in plant cells
    • C12N15/8222—Developmentally regulated expression systems, tissue, organ specific, temporal or spatial regulation
    • C12N15/823—Reproductive tissue-specific promoters
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
    • C12N15/8216—Methods for controlling, regulating or enhancing expression of transgenes in plant cells
    • C12N15/8222—Developmentally regulated expression systems, tissue, organ specific, temporal or spatial regulation
    • C12N15/823—Reproductive tissue-specific promoters
    • C12N15/8233—Female-specific, e.g. pistil, ovule
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
    • C12N15/8216—Methods for controlling, regulating or enhancing expression of transgenes in plant cells
    • C12N15/8222—Developmentally regulated expression systems, tissue, organ specific, temporal or spatial regulation
    • C12N15/823—Reproductive tissue-specific promoters
    • C12N15/8234—Seed-specific, e.g. embryo, endosperm

Definitions

  • the present invention relates generally to the field of plant molecular biology and the regulation of gene expression in plants.
  • the invention discloses nucleic acid sequences from tobacco containing a promoter region. More specifically, the present invention relates to the regulation of gene expression in plants with specificity to flowers or seeds.
  • Suitable promoters may be selected for a particular gene construct, for expression in a specific cell type, tissue, plant or environment. Promoters that are useful for plant transgene expression include those that are viral, synthetic, inducible, constitutive, temporally regulated, spatially regulated, tissue-specific, and spatio-temporally regulated. Promoters from bacteria, fungi, viruses and plants have been used to control gene expression in plant cells. It is often desirable to have tissue-specific expression of a gene of interest in a plant. Tissue-specific promoters can promote expression exclusively in one set of tissues without expression throughout the plant. Tissue-preferred promoters can drive expression at a higher level in a subset of tissues with significantly less or even no expression in the other tissues of the plant.
  • one particular area of interest is modulating the flavour of tobacco to generate a more desirable taste when tobacco is smoked.
  • modulation of threonine synthase expression or activity in tobacco plants to modulate methionine concentration and the flavour profile of tobacco is described in WO2013029800.
  • Modulation of the expression or activity of isopropylmalate synthase to alter sucrose ester composition and the flavour profile of tobacco is described in WO2013029799.
  • Modulation of the expression or activity of neoxanthin synthase to modulate the amount of beta- damascenone that is detectable in the aerosol of heated tobacco resulting in new flavour profiles in tobacco is described in WO2013064499.
  • compositions and methods for directing flower or seed specific expression in plants are provided.
  • a novel nucleic acid molecule isolated from tobacco, that drives expression of genes in a flower or seed specific manner in plants is provided.
  • an isolated or synthetic polynucleotide promoter capable of directing expression in a plant cell, wherein said nucleic acid promoter comprises a polynucleotide sequence as set forth in SEQ ID NO: 1 or having at least 60% sequence identity thereto.
  • said promoter is capable of driving flower or seed specific expression of an operably linked polynucleotide sequence of interest.
  • a polynucleotide construct comprising a polynucleotide promoter capable of directing expression in a plant cell, wherein said nucleic acid promoter comprises a polynucleotide sequence as set forth in SEQ ID NO: 1 or having at least 60% sequence identity thereto operably linked to a polynucleotide sequence of interest.
  • the polynucleotide construct comprises, in the 5' to 3' direction, the polynucleotide promoter and the polynucleotide of interest positioned downstream from said promoter and operatively associated therewith.
  • said polynucleotide of interest encodes a protein that contributes to the flavour profile of tobacco.
  • said polynucleotide construct is a vector.
  • a plant cell comprising the polynucleotide construct.
  • a method of making a plant comprising regenerating a plant from the plant cell.
  • a plant comprising the plant cell.
  • a method for expressing a polynucleotide of interest in a plant cell comprising the use of the polynucleotide construct, preferably, wherein said method comprises the steps of: (a) transforming a plant cell with the polynucleotide construct; and (b) growing the transformed plant cell under conditions suitable for the polynucleotide of interest to be expressed by the promoter.
  • a method of modulating the flavour profile of tobacco comprising the steps of: (a) transforming a tobacco plant cell with the polynucleotide construct, wherein the polynucleotide of interest encodes a protein that contributes to the flavour profile of tobacco; (b) growing a tobacco plant containing the plant cell under conditions suitable for the polynucleotide of interest to be expressed; (c) harvesting and optionally curing tobacco plant material from the tobacco plant of step (b); and (d) determining if the flavour profile of the tobacco plant material has been modulated in comparison to control tobacco plant material.
  • a method for producing a plant having modified expression of a polynucleotide sequence of interest comprising: (a) transforming a plant cell with the polynucleotide construct; (b) selecting a transformed cell; (c) generating a plant from the transformed cell; and (d) selecting a plant having modified expression of the polynucleotide sequence of interest.
  • a method of making a transformed plant comprising transforming a plant cell with the polynucleotide construct to produce a transformed plant cell, and then regenerating a plant from said transformed plant cell.
  • nucleic acid promoter or the polynucleotide construct for expressing a polynucleotide of interest in a plant cell.
  • said polynucleotide of interest encodes a protein that contributes to the flavour profile of tobacco.
  • Figure 1 shows the relative expression levels of NND3 and related functional CYP82E genes in different plant tissues. Bars indicate mean ⁇ SD of three biological replicates taken from three greenhouse grown mature N. tabacum var. TN90 plants.
  • Figure 2 shows relative NND3 expression levels in different parts of flowers.
  • Figure 3 shows relative expression levels of NND3 and related functional CYP82E genes in N. tabacum var. Stella leaves at different curing time points. Samples are taken from pools of several leaves. Two pools are analyzed as biological replicates - (a) replicate 1 and (b) replicate 2. Bars indicate mean ⁇ SD of three technical replicates.
  • isolated refers to any entity that is taken from its natural milieu, but the term does not connote any degree of purification.
  • An "expression vector” is a nucleic acid vehicle that comprises a combination of nucleic acid components for enabling the expression of nucleic acid. Suitable expression vectors include episomes capable of extra-chromosomal replication such as circular, double-stranded nucleic acid plasmids; linearized double-stranded nucleic acid plasmids; and other functionally equivalent expression vectors of any origin.
  • An expression vector comprises at least a promoter positioned upstream and operably-linked to a nucleic acid, nucleic acid constructs or nucleic acid conjugate.
  • construct refers to a double-stranded, recombinant nucleic acid fragment comprising one or more polynucleotides.
  • the construct comprises a "template strand” base- paired with a complementary "sense or coding strand.”
  • a given construct can be inserted into a vector in two possible orientations, either in the same (or sense) orientation or in the reverse (or anti-sense) orientation with respect to the orientation of a promoter positioned within a vector - such as an expression vector.
  • a “vector” refers to a nucleic acid vehicle that comprises a combination of nucleic acid components for enabling the transport of nucleic acid, nucleic acid constructs and nucleic acid conjugates and the like.
  • Suitable vectors include episomes capable of extra- chromosomal replication such as circular, double-stranded nucleic acid plasmids; linearized double-stranded nucleic acid plasmids; and other vectors of any origin.
  • a "promoter” is an untranslated DNA sequence typically upstream of a coding region that contains the binding site for RNA polymerase and initiates transcription of the DNA.
  • the promoter region may include other elements - such as promoter regulatory sequences - that act as regulators of gene expression.
  • Promoter regulatory sequences can contain proximal and/or distal upstream elements, the latter elements often being referred to as enhancers.
  • An enhancer is a DNA sequence that can stimulate promoter activity and may be an innate element of the promoter or a heterologous element inserted to enhance the level or tissue specificity of the promoter. It is generally capable of operating in both orientations and is capable of functioning even when moved either upstream or downstream from the promoter.
  • “Operably-linked” refers to the association of polynucleotide sequences on a single nucleic acid fragment or construct so that the function of one polynucleotide sequence is affected by the other polynucleotide sequence.
  • a promoter is operably-linked with a coding sequence when it is capable of affecting the expression of that coding sequence.
  • the coding sequence is therefore under the transcriptional control of the promoter.
  • the terms "homology, identity or similarity” refer to the degree of sequence similarity between two polypeptides or between two nucleic acid molecules compared by sequence alignment.
  • the degree of homology between two discrete nucleic acid sequences being compared is a function of the number of identical, or matching, nucleotides at comparable positions.
  • the percent identity may be determined by visual inspection and mathematical calculation. Alternatively, the percent identity of two nucleic acid sequences may be determined by comparing sequence information using a computer program such as - ClustalW, BLAST, FASTA or Smith-Waterman.
  • a “variant” means a substantially similar sequence.
  • a variant can have a similar function or substantially similar function as a wild-type sequence.
  • a similar function is at least about 50%, 60%, 70%, 80% or 90% of wild-type promoter activity under the same conditions.
  • a substantially similar function is at least about 90%, 95%, 96%, 97%, 98% or 99% of wild-type promoter function under the same conditions.
  • plant refers to any plant or plant part at any stage of its life cycle or development, and its progenies.
  • the plant is a "tobacco plant”, which refers to a plant belonging to the genus Nicotiana. Preferred species of tobacco plant are described herein.
  • Plant parts include plant cells, plant protoplasts, plant cell tissue cultures from which a whole plant can be regenerated, plant calli, plant clumps and plant cells that are intact in plants or parts of plants such as embryos, pollen, anthers, ovules, seeds, leaves, flowers, stems, branches, fruit, roots, root tips and the like. Progeny, variants and mutants of regenerated plants are also included within the scope of the disclosure, provided that they comprise the introduced polynucleotides described herein.
  • a "plant cell” refers to a structural and physiological unit of a plant.
  • the plant cell may be in the form of a protoplast without a cell wall, an isolated single cell or a cultured cell, or as a part of higher organized unit such as but not limited to, plant tissue, a plant organ, or a whole plant.
  • plant material refers to any solid, liquid or gaseous composition, or a combination thereof, obtainable from a plant, including biomass, leaves, stems, roots, flowers or flower parts, fruits, pollen, egg cells, zygotes, seeds, cuttings, secretions, extracts, cell or tissue cultures, or any other parts or products of a plant.
  • the plant material comprises or consists of biomass, stem, seed or leaves.
  • the plant material comprises or consists of leaves.
  • variable refers to a population of plants that share constant characteristics which separate them from other plants of the same species. While possessing one or more distinctive traits, a variety is further characterized by a very small overall variation between individuals within that variety. A variety is often sold commercially.
  • modulating may refer to reducing, inhibiting, increasing or otherwise affecting, for example, polynucleotide expression or transcriptional activity.
  • reduce refers to a reduction of from about 10% to about 99%, or a reduction of at least 10%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 75%, at least 80%, at least 90%, at least 95%, at least 98%, at least 99%, or at least 100% or more of a quantity or an activity - such as polynucleotide expression or transcriptional activity.
  • inhibitor refers to a reduction of from about 98% to about 100%, or a reduction of at least 98%, at least 99%, but particularly of 100%, of a quantity or an activity - such as polynucleotide expression or transcriptional activity
  • increase refers to an increase of from about 5% to about 99%, or an increase of at least 5%, at least 10%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 75%, at least 80%, at least 90%, at least 95%, at least 98%, at least 99%, or at least 100% or more of a quantity or an activity - such as polynucleotide expression or transcriptional activity.
  • control in the context of a control plant cell means a plant cell in which the expression of a polynucleotide has not been modified (for example, increased or reduced) and so it can provide a comparison with a plant cell in which the expression of the polynucleotide has been modified.
  • the control plant may comprise an empty vector.
  • the control plant cell may correspond to a wild-type plant cell.
  • the control plant cell can be the same genotype as the starting material for the genetic alteration that resulted in the subject plant cell. In all such cases, the subject plant cell and the control plant cell are cultured and harvested using the same protocols for comparative purposes. Changes in levels, ratios, activity, or distribution of the polynucleotides or genes can be measured by comparing a subject plant cell to the control plant cell, where the subject plant cell and the control plant cell have been cultured and/or harvested using the same protocols.
  • a promoter polynucleotide in one embodiment, there is provided a promoter polynucleotide.
  • the promoter polynucleotide can be an isolated or an artificial or a synthetic promoter polynucleotide.
  • the promoter polynucleotide can comprise, consist or consist essentially of a polynucleotide sequence having at least 60% sequence identity to any of the sequences described herein, including any of polynucleotides shown in the sequence listing. The sequence of the full- length complement, the reverse full-length complement, and the reverse sequence thereof are also disclosed.
  • the isolated promoter polynucleotide comprises, consists or consists essentially of a sequence having at least 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 75%, 80%, 85%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95% 96%, 97%, 98%, 99% or 100% sequence identity thereto.
  • an isolated promoter polynucleotide comprising, consisting or consisting essentially of a polynucleotide sequence having at least 60% sequence identity to SEQ ID NO:1.
  • the isolated promoter polynucleotide comprises, consists or consist essentially of a sequence having at least about 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 75%, 80%, 85%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95% 96%, 97%, 98%, 99%, 99.1 %, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or 100% sequence identity to SEQ ID NO:1.
  • a promoter polynucleotide comprising, consisting or consisting essentially of a polynucleotide with substantial homology (that is, sequence similarity) or substantial identity to SEQ ID NO:1 .
  • a promoter polynucleotide variant that has at least about 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 75%, 80%, 85%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95% 96%, 97%, 98%, 99%, 99.1 %, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% sequence identity to the sequence of SEQ ID NO:1 .
  • fragments of SEQ ID NO:1 with substantial homology (that is, sequence similarity) or substantial identity thereto that have at least about 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 75%, 80%, 85%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95% 96%, 97%, 98%, 99%, 99.1 %, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or 100% sequence identity to the corresponding fragments of SEQ ID NO:1.
  • a promoter polynucleotide comprising a sufficient or substantial degree of identity or similarity to SEQ ID NO:1.
  • Fragments of SEQ ID NO:1 that function as a promoter are also disclosed.
  • the fragments include those that can be assembled within recombinant constructs.
  • Fragments of the polynucleotide sequence may range from at least about 100 nucleotides up to the full-length polynucleotide.
  • the polynucleotides described herein include a polymer of nucleotides comprising or consisting of deoxyribonucleic acid (DNA). Although the polynucleotide sequences described herein are shown as DNA sequences, the sequences include corresponding RNA sequences, and their complementary (for example, completely complementary) DNA or RNA sequences, including the reverse complements thereof.
  • polynucleotides will generally contain phosphodiester bonds, although in some cases, polynucleotide analogues are included that may have alternate backbones, comprising, for example, phosphoramidate, phosphorothioate, phosphorodithioate, or O-methylphophoroamidite linkages; and peptide polynucleotide backbones and linkages.
  • Other analogue polynucleotides include those with positive backbones; non-ionic backbones, and non-ribose backbones. Modifications of the ribose-phosphate backbone may be done for a variety of reasons, for example, to increase the stability and half-life of such molecules in physiological environments or as probes on a biochip.
  • Mixtures of naturally occurring polynucleotides and analogues can be made; alternatively, mixtures of different polynucleotide analogues, and mixtures of naturally occurring polynucleotides and analogues may be made.
  • a variety of polynucleotide analogues are known, including, for example, phosphoramidate, phosphorothioate, phosphorodithioate, O-methylphophoroamidite linkages and peptide polynucleotide backbones and linkages.
  • Other analogue polynucleotides include those with positive backbones, non-ionic backbones and non-ribose backbones.
  • Polynucleotides containing one or more carbocyclic sugars are also included.
  • Other analogues include peptide polynucleotides which are peptide polynucleotide analogues.
  • These backbones are substantially non-ionic under neutral conditions, in contrast to the highly charged phosphodiester backbone of naturally occurring polynucleotides. This may result in advantages.
  • the peptide polynucleotide backbone may exhibit improved hybridization kinetics.
  • Peptide polynucleotides have larger changes in the melting temperature for mismatched versus perfectly matched base pairs. DNA and RNA typically exhibit a 2-4 °C drop in melting temperature for an internal mismatch.
  • the drop is closer to 7-9 °C.
  • hybridization of the bases attached to these backbones is relatively insensitive to salt concentration.
  • peptide polynucleotides may not be degraded or degraded to a lesser extent by cellular enzymes, and thus may be more stable.
  • polynucleotides that hybridize under stringent conditions, typically moderately stringent conditions, and commonly highly stringent conditions, to the polynucleotide promoter described herein.
  • stringent conditions typically moderately stringent conditions, and commonly highly stringent conditions
  • the basic parameters affecting the choice of hybridization conditions and guidance for devising suitable conditions are described in Sambrook, J., E. F. Fritsch, and T. Maniatis (1989, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.) and can be readily determined by those having ordinary skill in the art based on, for example, the length or base composition of the polynucleotide.
  • One way of achieving moderately stringent conditions involves the use of a prewashing solution containing 5x Standard Sodium Citrate, 0.5% Sodium Dodecyl Sulphate, 1 .0 mM Ethylenediaminetetraacetic acid (pH 8.0), hybridization buffer of about 50% formamide, 6x Standard Sodium Citrate, and a hybridization temperature of about 55 °C (or other similar hybridization solutions, such as one containing about 50% formamide, with a hybridization temperature of about 42 °C), and washing conditions of about 60 °C, in 0.5x Standard Sodium Citrate, 0.1 % Sodium Dodecyl Sulphate.
  • highly stringent conditions are defined as hybridization conditions as above, but with washing at approximately 68 °C, 0.2 x Standard Sodium Citrate, 0.1 % Sodium Dodecyl Sulphate.
  • SSPE (1 x SSPE is 0.15M sodium chloride, 10 mM sodium phosphate, and 1.25 mM Ethylenediaminetetraacetic acid, pH 7.4) can be substituted for Standard Sodium Citrate (1 x Standard Sodium Citrate is 0.15M sodium chloride and 15 mM sodium citrate) in the hybridization and wash buffers; washes are performed for 15 minutes after hybridization is complete.
  • wash temperature and wash salt concentration can be adjusted as necessary to achieve a desired degree of stringency by applying the basic principles that govern hybridization reactions and duplex stability, as known to those skilled in the art and described further below (see, for example, Sambrook, J., E. F. Fritsch, and T. Maniatis (1989, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.).
  • the hybrid length is assumed to be that of the hybridising polynucleotide.
  • the hybrid length can be determined by aligning the sequences of the polynucleotides and identifying the region(s) of optimal sequence complementarity.
  • a linear DNA molecule has two possible orientations: the 5'-to-3' direction and the 3'-to-5' direction.
  • the reference sequence is positioned in the 5'-to-3' direction
  • a second sequence is positioned in the 5'-to-3' direction within the same polynucleotide molecule/strand
  • the reference sequence and the second sequence are orientated in the same direction, or have the same orientation.
  • the promoter sequence as described herein and a polynucleotide (gene) of interest under the regulation of the given promoter are positioned in the same orientation.
  • the reference sequence and the second sequence are orientated in anti-sense direction, or have anti-sense orientation.
  • Two sequences having anti-sense orientations with respect to each other can be alternatively described as having the same orientation, if the reference sequence (5'-to-3' direction) and the reverse complementary sequence of the reference sequence (reference sequence positioned in the 5'-to-3') are positioned within the same polynucleotide molecule/strand. The sequences set forth herein are shown in the 5'-to-3' direction.
  • the promoter described herein can be used to express of one or more polynucleotides of interest in a host cell.
  • the promoter described herein can be used to express of one or more polynucleotides of interest in a plant cell - such as a tobacco plant cell.
  • a construct (which can be, for example, a vector or an expression vector or a plasmid and the like) that is compatible with the cell to be transformed can be prepared which comprises one or more polynucleotides of interest together with the promoter described herein positioned upstream to express (suitably, overexpress) the polynucleotide(s) in the cell.
  • the coding sequence of the polynucleotide(s) of interest can be cloned between the promoter described herein and an optional transcriptional terminator whereby the coding sequence is operatively linked to the promoter and the transcriptional terminator is operatively linked to the coding sequence.
  • the construct can optionally include a selectable marker coding sequence. Examples of visible markers include, but are not limited to, .beta. -glucuronidase (GUS), Chloramphenicol Acetyl Transferase (CAT), Luciferase (LUC) and proteins with fluorescent properties - such as Green Fluorescent Protein (GFP) from Aequora victoria.
  • the promoter can drive expression of the polynucleotide(s) of interest to produce the encoded protein in the cell.
  • the polypeptide(s) of interest encoded by the recombinant polynucleotide(s) of interest can be a native polypeptide, or can be heterologous to the cell.
  • a construct carrying one or more polynucleotides of interest is generated to express a polynucleotide(s) of interest in a plant cell.
  • the construct carries the promoter described herein upstream of the polynucleotide(s) driving its expression in the plant cell.
  • the construct can optionally carry an antibiotic resistance gene to confer selection of the transformed cell.
  • one or more targeting sequences may be employed to target the polypeptide of interest to an intracellular compartment within cells or to the extracellular environment.
  • a DNA sequence encoding a transit or signal peptide sequence may be operably linked to a sequence encoding a desired polypeptide of interest such that, when translated, the transit or signal peptide can transport the polypeptide of interest to a particular intracellular or extracellular destination, respectively, and can then be post- translationally removed.
  • Transit or signal peptides act by facilitating the transport of proteins through intracellular membranes, e.g., vacuole, vesicle, plastid and mitochondrial membranes, whereas signal peptides direct proteins through the extracellular membrane.
  • the transit or signal peptide can direct a desired protein to a particular organelle - such as a plastid, rather than to the cytoplasm.
  • the polynucleotide of interest may be identified in genomic DNA sequences - such as genomic DNA sequences of plants - for which genome sequence information is available to the public or isolated from polynucleotide libraries.
  • the polynucleotides may be artificial or synthetic polynucleotides. Such artificial or synthetic polynucleotides can be synthesised using known state-of-the-art techniques.
  • the polynucleotides may be synthesised using automated oligonucleotide synthesizers (for example, the Beckman DNA OLIGO 1000M synthesiser) so as to obtain polynucleotide fragments of desired length. A multitude of these polynucleotide fragments may then be linked using known DNA manipulation techniques.
  • a recombinant polynucleotide construct for use in the present disclosure can comprise one or more polynucleotides of interest encoding one or more polypeptides of interest, operably linked to the promoter described herein for expressing the polynucleotides in a plant cell.
  • Plant cells in which protein expression occurs can be non-naturally occurring plant cells, transgenic plant cells, man-made plant cells, genetically engineered plant cells or mutant plant cells. Plants in which protein expression occurs can be non-naturally occurring plants, transgenic plants, man-made plants, genetically engineered plants or mutant plants. Expression may occur in certain cells or in certain parts of a plant. Expression may occur in certain tissues of a plant. Expression may occur in specific tissues of a plant. Expression may occur in a specific tissue of a plant.
  • the promoter of the present invention is capable of expressing polynucleotides in flowers or seeds.
  • the non-naturally occurring, transgenic, man-made, genetically engineered or mutant plant cell or plant can comprise a genome that has been altered by the integration (suitably, the stable integration) of recombinant DNA therein.
  • Recombinant DNA includes DNA which has been genetically engineered and constructed outside of a cell and includes DNA containing naturally occurring DNA or cDNA or synthetic DNA.
  • a transgenic, man-made, genetically engineered or mutant plant cell can include a plant or plant cell regenerated from an originally-transformed plant or plant cell and progeny transgenic plants or plant cells from later generations or crosses of a transformed plant or plant cell.
  • a plant cell can be transformed by having the recombinant polynucleotide or a construct comprising same integrated into its genome to become stably transformed.
  • the plant cell can be stably transformed.
  • Stably transformed cells typically retain the introduced polynucleotide(s) with each cell division.
  • a plant cell can be transiently transformed such that the recombinant polynucleotide or a construct comprising same is not integrated into its genome.
  • Transiently transformed cells typically lose all or some portion of the introduced recombinant polynucleotide or construct comprising same with each cell division such that the introduced recombinant polynucleotide cannot be detected in daughter cells after a sufficient number of cell divisions.
  • the plant cell is stably transformed.
  • the constructs, vectors and the like may be introduced into a plant genome by a variety of conventional techniques. For example, A. tumefaciens mediated transformation, electroporation, protoplast fusion, injection in reproductive organs, injection in immature embryos; microinjection of plant cell protoplasts; use of ballistic methods - such as DNA coated particle bombardment.
  • the choice of technique will depend on the plant type to be transformed. For example, dicot plants and some monocots and gymnospermae may be transformed using Agrobacterium Ti plasmid technology.
  • the constructs, vectors and the like may be combined with appropriate T-DNA flanking regions and introduced into the conventional A. tumefaciens host vector. The virulence factor of the A.
  • Tissues such as leaf tissues, dissociated cells, protoplasts, seeds, embryos, meristemic regions, cotyledons, hypocotyledons and others can be transformed.
  • Vectors containing the recombinant polynucleotide constructs are also provided.
  • Suitable vector backbones include, for example, those routinely used in the art such as plasmids, viruses, artificial chromosomes, bacterial artificial chromosomes, yeast artificial chromosomes, or bacteriophage artificial chromosomes.
  • Suitable expression vectors include, without limitation, plasmids and viral vectors derived from, for example, bacteriophage, baculoviruses, and retroviruses. Numerous vectors and expression systems are commercially available.
  • the vectors can include, for example, origins of replication, scaffold attachment regions or markers.
  • a marker gene can confer a selectable phenotype on a plant cell.
  • a marker can confer biocide resistance, such as resistance to an antibiotic (for example, kanamycin, G418, bleomycin, or hygromycin), or an herbicide (for example, glyphosate, chlorsulfuron or phosphinothricin).
  • an expression vector can include a tag sequence designed to facilitate manipulation or detection (for example, purification or localization) of the expressed polypeptide.
  • Tag sequences such as luciferase, beta-glucuronidase, green fluorescent protein, glutathione S-transferase, polyhistidine, c- myc or hemagglutinin sequences typically are expressed as a fusion with the encoded polypeptide.
  • tags can be inserted anywhere within the polypeptide, including at either the carboxyl or amino terminus.
  • TMV Tobacco Mosaic Virus
  • MCMV Maize Chlorotic Mottle Virus
  • AMV Alfalfa Mosaic Virus
  • leader sequences known in the art include but are not limited to: picornavirus leaders, for example, EMCV leader (Encephalomyocarditis 5' noncoding region), potyvirus leaders, for example, TEV leader (Tobacco Etch Virus), MDMV leader (Maize Dwarf Mosaic Virus), human immunoglobulin heavy-chain binding protein (BiP) leader, untranslated leader from the coat protein mRNA of alfalfa mosaic virus (AMV RNA 4) and tobacco mosaic virus leader (TMV).
  • picornavirus leaders for example, EMCV leader (Encephalomyocarditis 5' noncoding region)
  • potyvirus leaders for example, TEV leader (Tobacco Etch Virus), MDMV leader (Maize Dwarf Mosaic Virus), human immunoglobulin heavy-chain binding protein (BiP) leader, untranslated leader from the coat protein mRNA of alfalfa mosaic virus (AMV RNA 4) and tobacco mosaic virus leader (TMV).
  • Plants suitable for use in the present disclosure include, but are not limited to, monocotyledonous and dicotyledonous plants and plant cell systems, including species from one of the following families: Acanthaceae, Alliaceae, Alstroemeriaceae, Amaryllidaceae, Apocynaceae, Arecaceae, Asteraceae, Berberidaceae, Bixaceae, Brassicaceae, Bromeliaceae, Cannabaceae, Caryophyllaceae, Cephalotaxaceae, Chenopodiaceae, Colchicaceae, Cucurbitaceae, Dioscoreaceae, Ephedraceae, Erythroxylaceae, Euphorbiaceae, Fabaceae, Lamiaceae, Linaceae, Lycopodiaceae, Malvaceae, Melanthiaceae, Musaceae, Myrtaceae, Nyssaceae, Papaveraceae, Pinacea
  • Suitable species may include members of the genera Abelmoschus, Abies, Acer, Agrostis, Allium, Alstroemeria, Ananas, Andrographis, Andropogon, Artemisia, Arundo, Atropa, Berberis, Beta, Bixa, Brassica, Calendula, Camellia, Camptotheca, Cannabis, Capsicum, Carthamus, Catharanthus, Cephalotaxus, Chrysanthemum, Cinchona, Citrullus, Coffea, Colchicum, Coleus, Cucumis, Cucurbita, Cynodon, Datura, Dianthus, Digitalis, Dioscorea, Elaeis, Ephedra, Erianthus, Erythroxylum, Eucalyptus, Festuca, Fragaria, Galanthus, Glycine, Gossypium, Helianthus, Hevea, Hordeum, Hyoscyamus, Jatropha, Lactuca, Linum, Lolium, Lup
  • Suitable species may include Panicum spp., Sorghum spp., Miscanthus spp., Saccharum spp., Erianthus spp., Populus spp., Andropogon gerardii (big bluestem), Pennisetum purpureum (elephant grass), Phalaris arundinacea (reed canarygrass), Cynodon dactylon (bermudagrass), Festuca arundinacea (tall fescue), Spartina pectinata (prairie cord-grass), Medicago sativa (alfalfa), Arundo donax (giant reed), Secale cereale (rye), Salix spp.
  • Eucalyptus spp. (willow), Eucalyptus spp. (eucalyptus), Triticosecale (tritic wheat times rye), bamboo, Helianthus annuus (sunflower), Carthamus tinctorius (safflower), Jatropha curcas (jatropha), Ricinus communis (castor), Elaeis guineensis (palm), Linum usitatissimum (flax), Brassica juncea, Beta vulgaris (sugarbeet), Manihot esculenta (cassaya), Lycopersicon esculentum (tomato), Lactuca sativa (lettuce), Musyclise alca (banana), Solanum tuberosum (potato), Brassica oleracea (broccoli, cauliflower, Brussels sprouts), Camellia sinensis (tea), Fragaria ananassa (strawberry), Theobroma cacao (cocoa),
  • Phleum pratense timothy
  • Panicum virgatum switchgrass
  • Sorghum bicolor sorghum, sudangrass
  • Miscanthus giganteus micanthus
  • Saccharum sp Saccharum sp.
  • the disclosure can be applied to any species of the genus Nicotiana, including N. rustica and N. tabacum (for example, LA B21 , LN KY171 , Tl 1406, Basma, Galpao, Perique, Beinhart 1000-1 , and Petico).
  • Other species include N. acaulis, N. acuminata, N. africana, N. alata, N. ameghinoi, N. amplexicaulis, N. arentsii, N. attenuata, N. azambujae, N. benavidesii, N. benthamiana, N. bigelovii, N. bonariensis, N. cavicola, N. clevelandii, N.
  • cordifolia N. corymbosa, N. debneyi, N. excelsior, N. forgetiana, N. fragrans, N. glauca, N. glutinosa, N. goodspeedii, N. gossei, N. hybrid, N. ingulba, N. kawakamii, N. knightiana, N. langsdorffii, N. linearis, N. longiflora, N. maritima, N. megalosiphon, N. miersii, N. noctiflora, N. nudicaulis, N. obtusifolia, N. occidentalis, N. occidentalis subsp. hesperis, N. otophora, N.
  • paniculata N. pauciflora, N. petunioides, N. plumbaginifolia, N. quadrivalvis, N. raimondii, N. repanda, N. rosulata, N. rosulata subsp. ingulba, N. rotundifolia, N. setchellii, N. simulans, N. solanifolia, N. spegazzinii, N. Stockton ii, N. suaveolens, N. sylvestris, N. thyrsiflora, N. tomentosa, N. tomentosiformis, N. trigonophylla, N. umbratica, N. undulata, N. velutina, N. wigandioides, and N. x sanderae.
  • Nicotiana tabacum varieties include Burley type, dark type, flue-cured type, and Oriental type tobaccos.
  • varieties or cultivars are: BU 64, CC 101 , CC 200, CC 27, CC 301 , CC 400, CC 500, CC 600, CC 700, CC 800, CC 900, Coker 176, Coker 319, Coker 371 Gold, Coker 48, CD 263, DF91 1 , DT 538 LC Galpao tobacco, GL 26H, GL 350, GL 600, GL 737, GL 939, GL 973, HB 04P, HB 04P LC, HB3307PLC, Hybrid 403LC, Hybrid 404LC, Hybrid 501 LC, K 149, K 326, K 346, K 358, K394, K 399, K 730, KDH 959, KT 200, KT204LC, KY10, KY14, KY 160, KY 17, KY 171
  • a further aspect relates to a seed of a plant described herein.
  • the seed is a tobacco seed of a tobacco plant.
  • a further aspect relates to pollen or an ovule of the plant.
  • a plant as described herein which further comprises a nucleic acid conferring male sterility.
  • the regenerable cells include but are not limited to cells from leaves, pollen, embryos, cotyledons, hypocotyls, roots, root tips, anthers, flowers and a part thereof, ovules, shoots, stems, stalks, pith and capsules or callus or protoplasts derived therefrom.
  • the promoter described here may increase the level of expression of one or more polynucleotides of interest.
  • An increase in expression as compared to a control may be from about 5 % to about 100 %, or an increase of at least 10 %, at least 20 %, at least 25 %, at least 30 %, at least 40 %, at least 50 %, at least 60 %, at least 70 %, at least 75 %, at least 80 %, at least 90 %, at least 95 %, at least 98 %, or 100 % or more - such as 200% or 300% or more, which includes an increase in transcriptional activity.
  • the promoter described here may reduce the level of expression of one or more polynucleotides of interest.
  • a reduction in expression as compared to a control may be from about 5 % to about 100 %, or a reduction of at least 10 %, at least 20 %, at least 25 %, at least 30 %, at least 40 %, at least 50 %, at least 60 %, at least 70 %, at least 75 %, at least 80 %, at least 90 %, at least 95 %, at least 98 %, or 100 %, which includes a reduction in transcriptional activity.
  • Expression cassettes can be constructed with a visible marker.
  • Transient transformation methods can be used to assess promoter activity. Using transformation methods - such as microprojectile bombardment, Agrobacterium transformation or protoplast transformation, expression cassettes can be delivered to plant cells or tissues.
  • Reporter gene activity such as .beta. -glucuronidase activity, luciferase activity or GFP fluorescence - can be monitored after transformation over time after DNA delivery using methods well known in the art. Reporter gene activity can be monitored by enzymatic activity, by staining cells or tissue with substrate for the enzyme encoded by the reporter gene or by direct visualisation under an appropriate wavelength of light.
  • RNA levels may be measured using methods well known in the art - such as Northern blotting, competitive reverse transcriptase PCR and RNAse protection assays. These assays can be used to measure the level of expression of a promoter. Further confirmation of promoter activity can be obtained by stable transformation of the promoter in an expression cassette comprising a visible marker or polynucleotide of interest into a plant. Using various methods - such as enzymatic activity assays, RNA analysis and protein assays - promoter activity can be monitored over development, and additionally by monitoring expression in different tissues in the primary transformants and through subsequent generations of transgenic plants.
  • flavour and/or non-flavour compounds stored have been artificially manipulated.
  • such plants can be manipulated to synthesise and store flavour and non-flavour compounds at desired concentrations.
  • Flavour and non-flavour compounds synthesised in plant cells can be so synthesised from precursor compounds by the activity of enzymes.
  • the promoter of the present invention is useful for this purpose as it can be used to express one or more genes at modulated levels or in certain locations.
  • the promoter described here can be used to produce a polynucleotide either having a direct influence or encoding a polypetide having and influence on the reproductive function of the plant. Therefore, the promoter can influence the expression of a ribonucleic acid or a polypeptide which can promote sterility, induce mutations and/or DNA strand breaks or facilitate chromosomal crossings.
  • genes can be introduced and specifically expressed in seeds and flowers, which are responsible for induction of sterility in the plant.
  • Exemplary genes include the atp9 mitochondrial gene; see Hernould, et al., Proc Natl Acad Sci U S A. 1993 Mar 15;90(6):2370-4.
  • ribonucleases, antisense RNA or RNAi mediators can be encoded, which can suppress the expression of genes specifically in the seeds or flowers of the plant (see Mariani et al., 1990 Nature 347:737-741 ; AN et al., 2013 PLOS ONE vol. 8, e6816).
  • the promoter described herein can also be used to express genes in plant flowers, modifying the scent of flowers for commercial ends. For example, see Luecker et al., 2004 Plant Pysiol 134:510; El Tamer et al., 2003 J. Biotechnol 106:15; Luecker et al., 2004 Plant J. 39:135). Flower scents can be modified in ornamental species, as well as comestible species and smoking tobacco.
  • Cured plant material from the tobacco plants described herein is also provided.
  • Processes of curing green tobacco leaves are known by those having skills in the art and include without limitation air-curing, fire-curing, flue-curing and sun-curing.
  • the process of curing green tobacco leaves depends on the type of tobacco harvested. For example, Virginia flue (bright) tobacco is typically flue-cured, Burley and certain dark strains are usually air-cured, and pipe tobacco, chewing tobacco, and snuff are usually fire-cured.
  • Dried plant material from the tobacco plants described herein is also provided.
  • Processes of drying tobacco flowers or seeds are known by those having skills in the art and include without limitation freeze drying, air drying, or sun curing.
  • flower and/or seed materials which have been modified with a promoter as described herein.
  • such materials can be used for inclusion in tobacco products, for animal feeds, for the production of commercially desirable products including chemicals, pharmaceuticals and nutraceuticals, for the production of human and/or animal feedstuffs and as biological reactors.
  • flower material which has a modified scent can be included in tobacco products.
  • tobacco products including tobacco-containing aerosol forming materials comprising plant material - such as leaves, preferably cured leaves - from the plants described herein.
  • plant material - such as leaves, preferably cured leaves - from the plants described herein.
  • the tobacco products described herein can be a blended tobacco product which may further comprise unmodified tobacco.
  • Methods for producing seeds are also described comprising cultivating the plant described herein, and collecting seeds from the cultivated plants. Seeds from the plants can be conditioned and bagged in packaging material by means known in the art to form an article of manufacture. Packaging material such as paper and cloth are well known in the art. A package of seed can have a label, for example, a tag or label secured to the packaging material, a label printed on the package that describes the nature of the seeds therein.
  • tissue specificity of the newly identified NND3 promoter sequence To determine the tissue specificity of the newly identified NND3 promoter sequence, different tissues of greenhouse grown N. tabacum var. TN90 are harvested and analysed for NND3 gene expression via quantitative PCR.
  • RNA extraction is performed using the RNeasy Mini kit (Qiagen) according to the manufacturer's instructions. RNA samples are diluted in water to obtain 1 ⁇ g RNA in 10 ⁇ final volume. Then DNase digestion is performed with RQ1 RNase-Free DNase (Promega). The DNase reaction is stopped using RQ1 DNase stop solution (Promega). Immediately, the reverse transcriptase (RT) reaction is performed to convert RNA into complementary DNA (cDNA). For RT reactions, M-MLV Reverse Transcriptase, RNase H Minus, Point Mutant (Promega) was used in combination with oligo(dT)15 primers.
  • RT reverse transcriptase
  • Quantitative real-time PCR is performed using the Stratagene Mx3005P and the corresponding software. For each target, different primer pairs are designed according to the guidelines of the Mx3005P user handbook. The primer pairs are tested for primer dimer formation and their performance in a qPCR run. Their efficiency is tested using a standard curve with a five-fold dilution of cDNA. The PCR products are sequenced in order to verify that the primers specifically amplify their target sequence. As the different CYP82E genes are close in sequence, primer design is complex and the chosen primers do not always show optimal efficiency.
  • the discrepancies in efficiency mean that the qPCR experiment values are gene specific and do not represent an absolute expression value. Therefore, the comparison in expression values is valid for each gene between the various tissues but not between genes.
  • Employed primer pairs and their efficiency are listed in Table 1 .
  • ABsolute Blue QPCR SYBR Green low ROX Mix (Thermo Scientific) is used with primer concentrations of 300 nM.
  • a denaturation temperature of 95°C is employed, initially for 15 minutes and then in each cycle for 15 seconds, 15 seconds at 60°C for annealing and 25 seconds at 72°C for elongation, for 50 cycles. All samples are run in triplicates. Furthermore, biological triplicates are employed.
  • the expression of the actin9 gene (house-keeping gene) is used for all samples as normaliser. The results are shown in Figure 1 and confirm the expression of CYP82E5 and CYP82E10 in green leaves. These genes are also highly expressed in all other tissues that were tested. They show a very similar expression pattern. Whilst CYP82E4 and NND3 each show expression in flowers and very minor expression in roots, NND3 is not expressed at detectable levels in leaves whereas CYP82E4 shows expression exclusively in senescent leaves. For primer specificity verification, the PCR products have been purified and sequenced. The low signal observed for NND3 in roots contained mixed products of NND3 and CYP82E4 amplification whereas products from flower material contained NND3 amplification product only.
  • NND3 expression in flowers was further analyzed. Flowers were dissected into different tissues, RNA was extracted and NND3 expression analysis was performed (using primers NND3_F6 and NND3_R7). The results are shown in Figure 2 and demonstrate specific expression of NND3 in ovary tissue. More than 10-times reduced expression can be observed in other reproductive tissues (stigma, filaments, and anthers). However, in the leaves surrounding the reproductive organs, only very low NND3 expression was observed (100-fold and more than 1000-fold reduced expression in petals and sepals, respectively, compared to the ovaries).
  • CYP82E5 and CYP82E10 are expressed to a similar extent in all leaf samples.
  • Expression of CYP82E4 is activated in senescent leaves and increases under curing conditions.
  • NND3 is not expressed in leaves under all conditions analyzed. Surprisingly NND3 shows a flower specific expression pattern with significant expression in flowers and flower derived material (capsules: i.e. dry fruit formed from fower tissue containing seeds).
  • SEQ ID NO: 1 AAAAAATAAAAAATTTAAAGTTGAATTGTTTGGCCAAACTTTTGGAGTAAAAAAAGTGTTTT

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Abstract

La présente invention concerne un promoteur d'acide nucléique isolé ou synthétique capable de diriger l'expression dans une cellule végétale, lequel promoteur d'acide nucléique comprend une séquence telle que présentée dans SEQ ID NO: 1 ou ayant au moins 60 % d'identité de séquence avec elle, de préférence, ladite cellule végétale étant une cellule de plante de tabac.
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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN117721112A (zh) * 2023-12-19 2024-03-19 海南省海洋与渔业科学院 红树植物白骨壤的内源启动子amdrep8及其用途
US11999960B2 (en) 2017-02-07 2024-06-04 University Of Kentucky Research Foundation Method of producing tobacco plants with increased sucrose ester content

Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996026639A1 (fr) * 1995-02-28 1996-09-06 Calgene, Inc. Modification de coton a l'aide de facteurs de transcription de tissu ovarien
EP0913469A1 (fr) * 1996-12-27 1999-05-06 Japan Tobacco Inc. Genes floraux specifiques d'organe et leurs sequences de promoteur
US5907082A (en) * 1995-11-17 1999-05-25 The Regents Of The University Of California Ovule-specific gene expression
WO2005111217A2 (fr) * 2004-04-29 2005-11-24 U.S. Smokeless Tobacco Company Molecules d’acide nucleique de nicotiana et leurs utilisations
WO2007044992A2 (fr) * 2005-10-13 2007-04-19 Cornell University Polynucleotides codant des phenylpropanoides et des enzymes dans le cafeier, et processus de biosynthese de flavonoides
WO2010122110A1 (fr) * 2009-04-22 2010-10-28 Basf Plant Science Company Gmbh Promoteur spécifique des graines entières
WO2013029800A1 (fr) * 2011-09-02 2013-03-07 Philip Morris Products S.A Thréonine synthase provenant de nicotiana tabacum et procédés et utilisations associés
WO2013064499A1 (fr) * 2011-10-31 2013-05-10 Philip Morris Products S.A. Modulation de bêta-damascenone chez des plantes

Patent Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996026639A1 (fr) * 1995-02-28 1996-09-06 Calgene, Inc. Modification de coton a l'aide de facteurs de transcription de tissu ovarien
US5907082A (en) * 1995-11-17 1999-05-25 The Regents Of The University Of California Ovule-specific gene expression
EP0913469A1 (fr) * 1996-12-27 1999-05-06 Japan Tobacco Inc. Genes floraux specifiques d'organe et leurs sequences de promoteur
WO2005111217A2 (fr) * 2004-04-29 2005-11-24 U.S. Smokeless Tobacco Company Molecules d’acide nucleique de nicotiana et leurs utilisations
WO2007044992A2 (fr) * 2005-10-13 2007-04-19 Cornell University Polynucleotides codant des phenylpropanoides et des enzymes dans le cafeier, et processus de biosynthese de flavonoides
WO2010122110A1 (fr) * 2009-04-22 2010-10-28 Basf Plant Science Company Gmbh Promoteur spécifique des graines entières
WO2013029800A1 (fr) * 2011-09-02 2013-03-07 Philip Morris Products S.A Thréonine synthase provenant de nicotiana tabacum et procédés et utilisations associés
WO2013064499A1 (fr) * 2011-10-31 2013-05-10 Philip Morris Products S.A. Modulation de bêta-damascenone chez des plantes

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
DIEGO ZAVALLO ET AL: "Isolation and functional characterization of two novel seed-specific promoters from sunflower (Helianthus annuus L.)", PLANT CELL REPORTS, SPRINGER, BERLIN, DE, vol. 29, no. 3, 19 January 2010 (2010-01-19), pages 239 - 248, XP019779612, ISSN: 1432-203X, DOI: 10.1007/S00299-010-0816-X *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11999960B2 (en) 2017-02-07 2024-06-04 University Of Kentucky Research Foundation Method of producing tobacco plants with increased sucrose ester content
CN117721112A (zh) * 2023-12-19 2024-03-19 海南省海洋与渔业科学院 红树植物白骨壤的内源启动子amdrep8及其用途

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