WO2000031251A1 - Regulation de l'expression genique post-recolte chez les plantes a l'aide de promoteurs de genes regules par la recolte - Google Patents

Regulation de l'expression genique post-recolte chez les plantes a l'aide de promoteurs de genes regules par la recolte Download PDF

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WO2000031251A1
WO2000031251A1 PCT/NZ1999/000195 NZ9900195W WO0031251A1 WO 2000031251 A1 WO2000031251 A1 WO 2000031251A1 NZ 9900195 W NZ9900195 W NZ 9900195W WO 0031251 A1 WO0031251 A1 WO 0031251A1
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promoter
plant
gene
harvest
expression
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Simon Allan Coupe
Somrutai Winichayakul
Richard Lawrence Moyle
Kevin Mark Davies
Graeme Arthur King
Kevin Farnden
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University Of Otago
New Zealand Institute For Crop & Food Research Limited
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Priority to AU14183/00A priority Critical patent/AU1418300A/en
Publication of WO2000031251A1 publication Critical patent/WO2000031251A1/fr

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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • 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)
    • C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
    • C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
    • C12N15/8257—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits for the production of primary gene products, e.g. pharmaceutical products, interferon
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    • 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
    • C12N15/8237—Externally regulated expression systems
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    • 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
    • C12N15/8237—Externally regulated expression systems
    • C12N15/8238—Externally regulated expression systems chemically inducible, e.g. tetracycline
    • 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)
    • C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
    • C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
    • C12N15/8243—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine
    • 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)
    • C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
    • C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
    • C12N15/8243—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine
    • C12N15/8249—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine involving ethylene biosynthesis, senescence or fruit development, e.g. modified tomato ripening, cut flower shelf-life

Definitions

  • the invention relates to the use of an asparagines synthetase (AS) promoter in the control of postharvest gene expression in plants.
  • AS asparagines synthetase
  • the invention in particular relates to the isolation and characterisation of the asparagus AS promoter and its use in postharvest gene expression.
  • a gene promoter can be defined as the complete sequence of nucleic acids immediately preceding the ATG start sequence of the gene of interest, and which contains regulatory elements controlling the endogenous expression of the gene.
  • Gene promoters control gene transcription through the binding or removal of transcription factors to positive and negative regulatory elements within the promoter sequence.
  • the interaction of the regulatory elements with the transcription factors denote the level of control - when, where and how much - of gene expression.
  • the biochemical or physical environment of the plant cell is translated, via a signal transduction cascade, to the active form of a transcription factor within the nucleus which then binds to, or is removed from, a regulatory element within the promoter. This connects a situation experienced by the cell to a response in the nucleus with the outcomes being a change in gene expression, changed protein synthesis and finally a response in the cell effected by the protein.
  • Gene promoters used in plasmid constructs for introducing a new gene into a transgenic plant can be of several types - constitutive, temporal or spatial.
  • constitutive promoters e.g., 35S from cauliflower mosaic virus (Gardner et al. , 1 981 )
  • Temporal promoters induce gene expression at certain times, often in response to specific conditions e.g., Cu inducible promoters (Mett et al., 1 996) .
  • Spatially induced promoters direct expression in specific plant organs e.g., the tuber-specific patatin type I promoter (Rocha-Sosa et al., 1 989) .
  • a constitutive promoter used to down-regulate the expression of a gene that is involved in deleterious processes after harvest the gene is down-regulated in all parts of the plant during growth and development as well as after harvest.
  • the expression of such genes may be necessary for the normal functioning of the plant prior to harvest and if this is the case the use of a constitutive gene promoter is inappropriate.
  • the introduction of a new quality parameter after harvest, such as a new colour or the ability to resist postharvest diseases may interfere with normal plant development.
  • the use of a gene promoter that can restrict expression to only after harvest would minimize the impact of new gene expression on plant growth and development.
  • AS in spear tips begins 3 h after harvest (Davies and King, 1 993) and follows a rapid depletion of soluble sugars, particularly sucrose, that begins within 2 h of harvest.
  • AS transcripts are not detectable in broccoli florets at harvest but increase within 2 h of harvest, and continue to increase up to 24 h.
  • AS transcripts are not detectable at harvest, but increase as excised leaves turn yellow (Downs and Somerfield, 1 997) .
  • Glutamine-dependant AS catalyses the transfer of the amide group of glutamine to aspartate, producing asparagine and glutamate.
  • the transamidation reaction catalysed by AS facilitates the storage of nitrogen in the soluble, stable and carbon efficient transportable form of asparagine.
  • the reaction catalysed by AS is particularly important during conditions of carbohydrate stress because asparagine has a lower C:N ratio than glutamine, making it the preferred nitrogen storage or transport molecule when carbon is limiting. For highly perishable vegetables, carbon
  • hexokinase acts as one sensor of carbohydrate levels within plant cells (Graham et al., 1 994; Jang and Sheen, 1 994; Davies et al., 1 996) but the steps beyond the perception of the signal - including the interaction of down-stream transcription factors with regulatory elements on the gene promoter is, as yet, unclear (Jang et al., 1 997) .
  • recent work has uncovered the nature of the sugar-responsive regulatory elements within two gene promoters.
  • the promoter sequences for cucumber isocitrate lyase and malate synthase have been analysed in detail for information on the elements conferring a response to sugar-starved up-regulation.
  • One of the sequence elements shared between cucumber isocitrate lyase and malate synthase promoters contains a core motif of CCCA which is involved in the sugar-induced down- regulation of malate synthase (Graham et al., 1 994; Reynolds and Smith, 1 995; Sarah et al., 1 996) .
  • the object of the present invention is to provide the use of an AS promoter for senescence- and/or harvest-regulated gene expression in plants.
  • the object is to provide a promoter isolated from asparagus for use in harvest-regulated gene expression in plants or to at least provide a useful choice.
  • the invention provides an isolated plant gene promoter which is regulated by harvest and/or senescence of a plant a plant part.
  • the promoter is an asparagine synthetase (AS) promoter and is preferably isolated from asparagus, more preferably Asparagus officinalis.
  • AS asparagine synthetase
  • the invention provides an asparagus AS promoter selected from the group comprising: (a) the DNA sequence depicted in Figure 1 ;
  • the invention also provides an isolated asparagus AS promoter.
  • AS promoter is isolated from Asparagus officinalis.
  • the invention also provides a suitable transformation vector or plasmid including the
  • the invention also provides the use of an AS promoter to control gene expression of a gene in a plant or plant part and preferably to control gene expression particularly in plant or a plant part after harvest and/or during senescence.
  • the invention also provides the use of the promoter to down-regulate gene expression in harvested plants or a plant part.
  • the invention also provides the use of the promoter in the identification of an element involved in harvest and senescence regulation.
  • the invention also provides the use of the promoter in the control of gene expression in response to changes in sugar concentration in a plant, or plant culture.
  • the invention describes the use of a gene promoter to control the expression of a gene related to postharvest quality in harvested horticultural and agricultural products.
  • the invention also relates to the use of the promoter to control the expression of new genes that allow the synthesis of new or novel substances or metabolites in plants or a plant part after excision.
  • the promoter described here is from Asparagus officinalis and directs the expression of the endogenous asparagine synthetase (AS) gene in asparagus spears in a harvest- responsive fashion.
  • AS asparagine synthetase
  • the isolated promoter is negatively affected by sugar levels, and is not responsive to light.
  • This promoter can be used to direct the expression of introduced genes in either the sense or antisense orientation to improve the postharvest quality of horticultural products and to enhance the keeping quality of stored forage crops.
  • the invention also provides a transgenic plant containing the promoter according to the invention.
  • Figure 1 shows the promoter sequence of the asparagus AS gene.
  • Figure 2 is a schematic diagram of the asparagus AS gene.
  • Figure 3 is a photograph of transformed arabidopsis leaves histochemically stained for GUS.
  • A AS-1 leaf after a 24 h postharvest treatment;
  • B AS-1 leaf at 0 h;
  • C C,
  • Figure 4 is a bar graph of GUS activity in transformed arabidopsis leaves using fluorometric detection.
  • Figure 5 is a graph of GUS activity in transformed arabidopsis leaves during growth in light/dark conditions, with or without 58 mM sucrose in the growing medium.
  • Figure 6 shows the transient analysis of GUS expression under the control of a 2 kb
  • Figure 7 is a northern blot of mRNA from 3 cm asparagus tips isolated from spears stored in air, or in the controlled atmospheres indicated for up to 6 days after harvest.
  • the northern blot was probed with pTIP27 (a cDNA clone encoding asparagus asparagine synthetase) .
  • the invention relates to the use of a gene promoter to control heterologous gene expression in harvested plants or plant parts.
  • the gene promoter originates from asparagus and in this plant controls expression of AS in response to harvest. Given the features of the promoter, discovered through experiments relating to the harvest- induced expression of the asparagus AS gene, and the direct analysis of the promoter
  • this gene promoter can be used to control the expression of desirable genes in harvested (excised) plants or plant parts. This promoter can also be used to down-regulate gene expression in harvested plants or plant parts, using antisense or other down-regulatory techniques.
  • the promoter outlined here controls gene expression in plants after they are harvested (excised) .
  • the promoter is light-independent and can be repressed by high levels of sucrose.
  • the promoter elements controlling sugar regulation appear to be separate from elements responsive to harvest, but may be invoked as conditions ensue in harvested plants or plant parts.
  • Harvest-regulated gene promoters can be used for (but are not limited to) the following:
  • genes introduced for the purpose of enhancing the postharvest quality of horticultural or agricultural crops including fruit, vegetables, flowers, cereal and forage crops
  • Plant material Field grown Asparagus officinalis L. cv. Limbras 10 plants provided all spear material for DNA isolation.
  • Arabidopsis thaliana (L.) Heyn. (ecotype Columbia) was used in transformation experiments. Seed was surface-sterilised and germinated on half-strength Murashige and Skoog (MS) media (Murashige and Skoog, 1 962). After the plants had reached 3 to 4 true leaves they were potted up into 5 cm pots covered with muslin and allowed to grow to maturity (approx. 4 weeks) in either a glasshouse or growth cabinet with a 1 6 h day photoperiod and a 1 5°C night/25°C minimum day temperature.
  • MS Murashige and Skoog
  • Genomic asparagus DNA was partially digested with EcoR ⁇ restriction endonuclease (Boehringer Mannheim), ligated in EMBL4 phage arms (Stratagene) and packaged into phage particles using Gigapack II Gold packaging extracts (Stratagene) according to the manufacturers instructions.
  • the library was titred at 1 .2 x 10 6 pfu.
  • the library was amplified and 8.5 x 10 4 pfu plated onto each of six 9 cm 2 LB plates. Duplicate plaque lifts from each plate using Hybond N + nylon membrane (Amersham) were fixed on 0.4 M NaOH-soaked filter paper.
  • the membranes were prehybridised, hybridised to radiolabeled pTIP27 probe (asparagus cDNA clone encoding AS, Davies and King, 1 993), and washed according to the methods described for Southern blot analysis above. Seven of the 27 positive signals identified on the primary screen were screened a further three times to obtain clonal populations of phage. DNA was isolated from each of the seven phage clones according to the methods of Sambrook et al. ( 1 989) . Restriction mapping and Southern analysis revealed six of the phage clones contained the 1 6 kb EcoRI fragment of interest and one phage clone contained the 5.5 kb EcoRI fragment of interest identified by genomic Southern blot analysis.
  • the 5.5 kb and 1 6 kb phage inserts were gel-purified from the EMBL4 phage arms using a DNA purification kit (BioRad) .
  • the 5.5 kb EcoRI fragment was subcloned into phosphatase-treated, EcoRI-digested pUC1 9 plasmid to produce plasmid pR200.
  • the 1 6 kb EcoRI fragment was subcloned into phosphatase-treated, EcoRI- digested pUC1 9 plasmid to produce plasmid pR1 00.
  • Nested deletion of pR700 resulted in the isolation of subclones providing sequence for the second strand of the AS promoter.
  • All plasmids were prepared for sequencing using the Wizard DNA purification system kits (Promega) or QIAprep spin plasmid kits (Qiagen). All sequencing was performed using an automated DNA sequencer (Applied Biosystems), at the Otago Centre for Gene Research, University of Otago, Dunedin, New Zealand.
  • An existing promoter-GUS reporter gene expression plasmid was altered by removing the existing promoter sequence and replacing it with approximately 2 kb of AS promoter from the 5.5 kb genomic clone isolated earlier. Two bases around the ATG translation start site of the asparagus AS gene were changed by site-directed mutagenesis using the Sculptor in vitro mutagenesis kit (Amersham) to form an ⁇ /col site.
  • the AS promoter/GUS/CaMV poly-A expression cassette was cloned into the binary plasmid pART-27 (Gleave et al., 1 992) for Agrobacterium-med ated transformation of arabidopsis.
  • This plasmid contains the nptll gene (conferring kanamycin resistance) as a selectable marker.
  • This plasmid was called pAS-GUS.
  • the AS promoter/GUS/CaMV poly-A expression cassette was subcloned into the direct transformation plasmid pRT99 (Topfer et al., 1 988) to produce a plasmid pASP3 used for transient expression analysis in asparagus callus cultures.
  • a second plasmid, pRT99-GUS was constructed containing a CaMV 35S promoter/GUS/CaMV poly-A expression cassette and was used as a positive transformation control.
  • the transformation of arabidopsis with an AS promoter-GUS construct was achieved using the method of Bechtold et al. ( 1 993) with minor modifications.
  • the LBA4404 strain of Agrobacterium tumefaciens (Life Technologies), containing the binary vector pAS-GUS was used for transformation of arabidopsis.
  • LBA4404 Agrobacterium were grown with streptomycin (50 mg L "1 ) and kanamycin (100 mg L "1 ) f or 1 6 h at 28 °C in LB medium (final density of 2.0 AU at 600 nm) . After centrifugation, the bacterial pellet was resuspended in the infiltration medium as specified by Bechtold et al.
  • a particle accelerating gun with the modification of using a Nalgene desiccator as the vacuum chamber instead of a steel box, was used to transform the asparagus calli.
  • DNA-coated gold slurry (2 ⁇ l) of was loaded into the centre of a sterile filter unit (Millipore) which was screwed into the luer-lok fitting inside the top of the particle gun desiccator chamber.
  • a media tub containing the piece of asparagus calli to be transformed was placed on the stage at the bottom of the particle gun chamber directly under the filter unit.
  • a vacuum of 95 kPa 1 4 psi was applied to the chamber of the particle gun.
  • the prechamber was loaded with 500 kPa (60 psi) helium which was subsequently fired through the particle gun by detonating the timer set to 50 milliseconds.
  • the vacuum on the particle gun chamber was slowly released and the transformed asparagus calli removed and incubated for 48 h under near dark lighting at 22°C and 100% humidity.
  • the 5.5 kb and 1 6 kb EcoRI fragments identified by genomic Southern blot analysis were isolated by screening an EMBL4 library constructed with EcoRI partially digested asparagus genomic DNA using radiolabeled pTIP27 (asparagus cDNA encoding AS) as a probe.
  • the 5.5 kb EcoRI fragment and 2 kb of the 1 6 kb EcoRI fragment were sequenced.
  • the full length AS gene was contained within the 5.5 kb EcoRI fragment and the 1 6 kb EcoRI fragment (Moyle et al., 1 996).
  • the 5.5 kb EcoRI fragment also contained approximately 3 kb of AS promoter sequence.
  • Leaves from the eight plants putatively expressing the GUS gene under the control of the asparagus AS promoter had low levels of GUS activity at 0 h which dramatically increased after a 24 h postharvest treatment. This increased activity had a wide range of variability from an 8-fold increase (AS-3) to over a 100- fold increase (AS-5) with an average increase of 47-fold.
  • Plasmid pASP3 containing a 2 kb AS promoter fragment-GUS reporter gene construct and the positive control plasmid pRT99-GUS containing a CaMV 35S promoter-GUS construct were analysed for sugar-regulated gene expression using a particle gun- mediated transformation system. Calli transformed with the positive control CaMV 35S-GUS construct produced blue zones corresponding to GUS activity regardless of the presence or absence of sucrose ( Figure 6). Calli starved of sucrose for 48 h and transformed with the 2 kb AS promoter-GUS construct produced blue zones corresponding to GUS activity when assayed in the absence of sucrose.
  • AS gene expression in harvested spears in response to controlled atmosphere storage AS expression in air-stored and all CA-treated asparagus was identical both in timing and intensity ( Figure 7).
  • signals other than sucrose levels are involved in the harvest induction of the asparagus AS promoter and the promoter can control expression independently of sucrose.
  • Pea AS gene expression is up-regulated in attached leaves during dark treatments and repressed during light treatments (Tsai, 1 991 ; Tsai and Coruzzi, 1 991 ), and regulatory elements involved in the light response have been identified (Ngai et al., 1 997) .
  • the pea AS1 promoter has also been reported as being repressed by sucrose (Ngai et al., 1 997, in reference to Ngai and Coruzzi, unpublished data), and contains a CCCA core motif (Neuhaus et al., 1 997) that in the cucumber isocitrate lyase and malate synthase promoters is involved in the response element controlling sugar down-regulation (Graham et al., 1 994; Reynolds and Smith, 1 995; Sarah et al., 1 996).
  • Analysis of the asparagus AS promoter revealed a CCCA motif within a fragment at - 1 289 and - 1 299 bp ( Figure 1 ). This motif may be involved in sugar regulation as with the promoters for isocitrate lyase and malate synthase.
  • the invention provides an asparagus AS promoter and its use in the control of postharvest gene expression in plants or plant parts.
  • the use of the isolated promoter will assist in controlling spoilage after harvest in crops.
  • the promoter may also be useful in the response to sucrose levels in a plant/plant parts after harvest.
  • Plant Cell 6 1 665-1 679.
  • Postharvest biology and technology an overview. In: Postharvest Technology of Horticultural Crops. Ed. Kader AA. Second edition. University of California, Division of Agriculture and Natural Resources.
  • Lam H-M Peng S, Coruzzi GM. 1 994. Metabolic regulation of the gene encoding glutamine-dependent asparagine synthetase in Arabidopsis tha/iana. Plant Physiol. 1 06: 1 347-1 357.

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Abstract

L'invention concerne le promoteur du gène de l'asparagine synthétase (AS) issue de l'asperge. Ce promoteur est utilisé pour régir l'expression post-récolte des gènes dans les plantes ou dans des parties de plantes.
PCT/NZ1999/000195 1998-11-26 1999-11-25 Regulation de l'expression genique post-recolte chez les plantes a l'aide de promoteurs de genes regules par la recolte WO2000031251A1 (fr)

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2006056176A3 (fr) * 2004-11-26 2006-08-03 Kws Saat Ag Promoteur induit au stockage
US7329798B2 (en) 2002-06-28 2008-02-12 University Of Guelph Harvest-inducible regulatory elements and methods of using same
US7388091B2 (en) 2002-06-28 2008-06-17 University Of Guelph Harvest-inducible genes from alfalfa (Medicago sativa) and methods of use thereof

Citations (1)

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Cited By (6)

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Publication number Priority date Publication date Assignee Title
US7329798B2 (en) 2002-06-28 2008-02-12 University Of Guelph Harvest-inducible regulatory elements and methods of using same
US7388091B2 (en) 2002-06-28 2008-06-17 University Of Guelph Harvest-inducible genes from alfalfa (Medicago sativa) and methods of use thereof
WO2006056176A3 (fr) * 2004-11-26 2006-08-03 Kws Saat Ag Promoteur induit au stockage
DE102004057291C5 (de) * 2004-11-26 2010-08-26 Südzucker AG Mannheim/Ochsenfurt Lagerungsinduzierte Promotoren
EP2333077A1 (fr) * 2004-11-26 2011-06-15 KWS Saat AG Promoteur induite au stockage
US8093457B2 (en) 2004-11-26 2012-01-10 Kws Saat Ag Storage-induced promoter

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