EP1708558A2 - Proteines conferant aux plantes une resistance au stress biotique et abiotique - Google Patents

Proteines conferant aux plantes une resistance au stress biotique et abiotique

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Publication number
EP1708558A2
EP1708558A2 EP04787517A EP04787517A EP1708558A2 EP 1708558 A2 EP1708558 A2 EP 1708558A2 EP 04787517 A EP04787517 A EP 04787517A EP 04787517 A EP04787517 A EP 04787517A EP 1708558 A2 EP1708558 A2 EP 1708558A2
Authority
EP
European Patent Office
Prior art keywords
nucleic acid
acid sequences
plants
sequences seq
protein products
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP04787517A
Other languages
German (de)
English (en)
Other versions
EP1708558A4 (fr
Inventor
Villoo M. Avestha Gengraine Tech. Pvt.Ltd PATELL
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Avesthagen Ltd
Original Assignee
Avestha Gengraine Technologies Pvt Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Avestha Gengraine Technologies Pvt Ltd filed Critical Avestha Gengraine Technologies Pvt Ltd
Publication of EP1708558A2 publication Critical patent/EP1708558A2/fr
Publication of EP1708558A4 publication Critical patent/EP1708558A4/fr
Withdrawn legal-status Critical Current

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Classifications

    • 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
    • C12N15/8261—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield
    • C12N15/8271—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants

Definitions

  • the present invention relates to the isolation of a nucleic acid sequences plants, the products of which, confer resistance to various biotic and abiotic stress like wounding, pathogen infection and drought in plants.
  • AGPs are components of Gum Arabic, a gummy exudation originating from the Acacia tree and known to be produced by stress conditions such as heat, drought and wounding (Clarke et al, 1979). AGPs function in several biological process including cell-cell adhesion, pollen-stigma recognition, water retention and disease resistance in crops.
  • WO9515377 patent application provides plant arabinogalactan proteins (AGPs) and their genes.
  • AGPs were isolated from Nicotiana alata, Nicotiana plumbaginafolia, and Pyrus communis. Amino acid sequences of isolated AGP peptide fragments are presented. Isolated AGP fragments were used to synthesize oligonucleotide probes to prepare oligonucleotide primers for PCR or prepare RNA probes to screen cDNA libraries of N. alata, N. plumbaginafolia, and P. communis. cDNA clones encoding amino acid sequences of isolated AGP fragments were isolated. The invention presents for the first time an intact AGP amino acid sequence derived from a corresponding AGP gene. The instant invention further provides methods useful in obtaining AGP genes encoding an AGP peptide comprising a specific isolated hydroxyproline-rich (OAST-rich) sequence or a specific isolated hydroxyproline-poor sequence.
  • OFAST-rich isolated
  • Beta glucosidase catalyses one of the last steps in the lignin synthesis in plants. (Ellis Brian et al., 1999). The increased level of beta glucosidase significantly increases the efficiency of hydrolysis of cellulose to glucose by cellulase enzymes, thereby enhancing the production of fuel ethanol from cellulose. In spite of a significant amount of research effort, there has not been a means to produce sufficiently high levels of beta glucosidase. Such a process would be a large step forward in the production of fuel alcohol from cellulose (Theresa White & Christopher Hindle, 2000).
  • Beta glucosidase activity has been found to be 2-4 times higher as compared to the control plants in the presence of pathogens. (David Blanchard & Asim Esen, 2000). With respect to phosphate starvation stress of the eleven genes preferentially expressed in phosphate starved plant cells, beta glucosidase was one of the candidate genes (Mohd. AH Malboobi, 2000).
  • Beta glucosidase which is produced naturally in fungi is able to break down and release the aromatic substances in wine.
  • introduction of this enzyme into simple wines can convert them into more expensive products with greater appeal and commanding a higher price.
  • This gene By introducing this gene into wine yeast through genetic engineering, the scientists have succeeded in creating a wine yeast that produces the beta-glucosidase in large quantities. This technology may be utilised to produce wines with richer aroma than would normally be expected.
  • the same gene has also been implanted into tobacco plants to achieve a more aromatic plant (Dr. Oded Shoseyov et al., 2001).
  • over-expression vectors may be used to suppress coniferin beta- glucosidase enzyme activity through sense-suppression, as described in U.S. Pat. Nos. 5,034,323 and 5,283,184, both entitled "Genetic Engineering of Novel Plant Phenotypes".
  • US patent 5,997,913 describes a process for expressing extracellular .beta.-glucosidase in a filamentous fungus by expressing a fungal DNA sequence encoding enhanced, deleted or altered .beta.-glucosidase in a recombinant host microorganism is disclosed.
  • Recombinant fungal cellulase compositions containing enhanced, deleted or altered expression of ,beta.-glucosidase is also disclosed.
  • US patent 5,283,184 describes the use of recombinant DNA methods for genetically altering plants, and more particularly, to improved means for altering plant phenotypes, such as color patterns and color intensity of flowers and other plant parts.
  • Another object of the present invention is to provide transgenic plants constitutively transcribing sense or antisense mRNA strands of DNA sequences encoding plant PRPs or transcribing sense or antisense mRNA strands of DNA sequences substantially homologous to genomic or cDNA sequences encoding plant PRPs, such transgenic plants thus having an enhanced disease-resistant phenotype with respect to wild-type plants.
  • WO9220800 patent application describes methods of combating fungal disease and fungicidal compositions are provided in which a P14 protein, preferably PI 4a, PI 4b, PI 4c, P14d, P14e or P14f, or a fungicidally active analogue thereof is used as active ingredient.
  • the P14 proteins may be obtained from plant material but the proteins and analogues are preferably prepared by use of recombinant DNA technology.
  • DNA sequences coding for the P14 proteins and analogues, vectors, containing the DNA sequences and host cells transformed with the DNA sequences, as well as processes for production of the protein by culturing the transformed host cells, are provided.
  • transformed plant cells and. plants are provided having resistance to fungal disease.
  • Plant diseases caused by viral, bacterial, fungal and other pathogens are responsible for enormous economic loss.
  • the ability of a plant to stop invasion of a pathogen depends on the presence of performed barriers.
  • a distinct class of PR1 proteins, called intracellular proteins is expressed during wounding (Warner et al., 1992), osmotic stress (Iturriage et al., 1994) and pathogen colonisation (Chang & Hadwiger, 1990). IPR proteins are classified under PR1 since their function is not known but they share a low homology to them.
  • TSI-1 transcripts increased as the concentration of SA was increased and were maximal at 10 mM SA after 48 hours.
  • TSI-1 was not expressed in the control and an extremely faint signal was obtained in leaves treated with low concentration of SA.
  • high intensity signals were obtained in the fungal infected leaves after exposure for 24 hours but no signal was detected in the control lane after exposure for 2 days.
  • the TSI-1 was not expressed constitutively but induced during fungal infection (C.S. Sree Vidya et al., 1999).
  • Thionins are small, basic, cysteine rich proteins, which, may function as defence molecules against an array of plant pathogens (Florack & Stiekema, 1994; Broekaert et al., 1995). These genes appear to be expressed in response to pathogens and to be developmentally regulated. Several accumulate in reproductive tissue (Gu et al., 1992; Milligan & Gasser, 1995; Meyer et al., 1996).
  • Thionin gene in a plant exhibiting resistance to at least one disease such as a disease caused by a plant pathogenic bacterium, i.e., a bacterium causing bacterial leaf blight of rice or bacterium causing bacterial seedling blight of rice or a plant pathogenic filamentous fungus i.e., a fungus causing late blight of potato has been reported (Honkura Ryosos et al., 1993).
  • EP0902089 patent application relates to a transgenic plant which exhibits resistance to at least one disease.
  • the present invention relates to a transgenic plant which comprises an expression cassette including a thionin gene and being capable of expressing the thionin gene, and which exhibits resistance to at least one disease.
  • EP1101771 patent application describes two cDNA clones, designated to PepDef (pepper defensin protein gene) and PepThi (pepper thionin-like protein gene) and individual component; thereof including its coding region and its gene product; modification thereto; application of said gene, coding region and modification thereto; DNA construct, , vectors and transformed plants each comprising the gene or part thereof
  • the present invention relates to the isolation of a nucleic acid sequences plants, the products of which, confer resistance to various biotic and abiotic stress like wounding, pathogen infection and drought in plants.
  • ⁇ -glucosidases that pertain to food processing and quality is that edible portions of some plants contain compartmentalized ⁇ -glucosidase- ⁇ -glucoside systems that produce toxic aglycones and/or HCN when tissue is macerated during preparation or by chewing.
  • This is exemplified by cassava roots and leaves, lima beans and flax seed.
  • cassava is a food staple in tropical regions of Africa, Asia and South America, consumption reaches about 1 kg/per capita/day in some parts of Africa (e.g., Congo). It contains the cyanogenic ⁇ -glucoside linamarin and the corresponding A-glucosidase linamarase.
  • the myrosinase-glucosinolate (or ⁇ -thioglucosidase- ⁇ -thioglucoside) system which occurs in cruciferous vegetables (e.g., mustard, cabbage, kale, broccoli, rapeseed, horseradish, etc.), has also importance for food quality and processing because the aglycone moiety and its breakdown products from enzymatic hydrolysis of glucosinolates are responsible for bitter, pungent taste and aroma associated with these vegetables, as well as the processed foods and relishes that include them (10).
  • the distinct flavor associated with glucosinolates comes primarily from isothiocyanates and is believed to have evolved to serve as a repellent against microorganisms and herbivores.
  • Glucosinolates and their breakdown products may impart undesirable flavors to milk, meat and eggs when farm animals graze on cruciferous plants or when their feed includes seed meals from such plants.
  • off-odors and flavors in foods of animal origin associated with glucosinolates direct ingestion of large amount of cruciferous vegetables is thought to cause endemic goiter in humans, as well as toxicity in laboratory animals.
  • claims have been made on anti-carcinogenic effects of glucosinolates and their breakdown products in humans.
  • raw or cooked cruciferous vegetables e.g., cabbage, broccoli, cauliflower and turnip
  • aryl hydrocarbon hydroxylase activity (11).
  • Biomass Conversion Polysaccharides, specifically cellulose, are the most abundant substances in the biosphere ( ⁇ 5x ⁇ o 10 tons produced/year) and are potential renewable sources of chemicals and fuels. Moreover, about 40% of typical municipal garbage includes newspaper and other paper products. Hydrolysis of cellulose using inorganic acids and high temperature is not ecologically sound and economically feasible.
  • An enzyme (cellulase) complex secreted by cellulolytic organisms, can hydrolyze cellulose to glucose, thus presenting itself as a suitable model for industrial processes that need to be developed (12).
  • the complex includes three enzymes: an endoglucanase, an exoglucanase (cellobiohydrolase) and a ⁇ -glucosidase.
  • Lignin Biosynthesis and Paper Quality Lignin is the second most abundant substance in the biosphere and its major precursor, coniferyl alcohol, is derived from coniferin (4-O-coniferyl glucoside) after hydrolysis by ⁇ -glucosidase (15), suggesting that some plant ⁇ -glucosidase isoforms are involved in lignin biosynthesis. This makes the enzyme a suitable target for improving wood strength and quality for paper production.
  • Anthocyanins are classes of pigments that determine flower color and plant pigmentation in angiosperm plants.
  • pelargonidin-based pigments confer bric- red/orange color to plants
  • cyanidin- and delphinidin-based pigments confer red and violet color each (Holton, et al. Plant Cell 7:1071-1083 (1995); Tanaka, et al. Plant Cell Physiol. 39: 1119-1126 (1998)).
  • Different ratio of these pigments confers a wide range of flower color.
  • Many anthocyanin bio synthetic genes have been identified.
  • One of key enzyme in the biosynthetic pathway is dihydroflavonol 4-reductase (DFR).
  • the enzyme converts dihydroflavonols (dihydrokaempferol (DHK), dihydroquercetin (DHQ), and dihydromyricetin (DHM)) to leucocyanidins.
  • DHK dihydrokaempferol
  • DHQ dihydroquercetin
  • DLM dihydromyricetin
  • the leucocyanidins are subsequently converted to anthocyanins by other enzymes.
  • DFRs in most plants can convert all three dihydroflavonols to leucocyanidins
  • the ratio of three classes of anthocyanin pigments are mainly determined by the activity of F3 ⁇ and F3'5 ⁇ (Holton, et al. Plant Cell 7:1071-1083 (1995).
  • Dihydroflavanol 4-reductase (“DFR") reduces Dihydroflavanol to leuco-anthocyanin, which is then further converted into anthocyanin via anthocyanidin. This has been found to be one of the regulatory enzymes in the flavonoid biosynthetic pathway.
  • it has applications in increasing plant tolerance and plant defense to biotic stresses such as viruses, micro-organisms, insects and fungal pathogens; in improving plant forage quality, for example by disrupting protein foam and in conferring protection from rumen pasture bloat ; in reducing digestion rates in the rumen and reducing parasitic load ; in the production of plant compounds leading to health benefits, such as isoflavonoids, which have been linked to anticancer benefits, and stilbenes that are believed to contribute to reduced heart disease.
  • biotic stresses such as viruses, micro-organisms, insects and fungal pathogens
  • improving plant forage quality for example by disrupting protein foam and in conferring protection from rumen pasture bloat
  • in reducing digestion rates in the rumen and reducing parasitic load in the production of plant compounds leading to health benefits, such as isoflavonoids, which have been linked to anticancer benefits, and stilbenes that are believed to contribute to reduced heart disease.
  • the defense of plants to pathogens comprises constitutive barriers present in plants prior to any contact with pathogens or herbivores. Furthermore, exposure to various microorganisms or other forms of stress can lead to the activation of defense mechanisms. Induced resistance depends on the recognition of a pathogen or stress by the plant. This generates a cascade of events, eventually leading to the expression of defense mechanisms, which include physical barriers, metabolites and proteins that interfere with the spread of the invading microorganism.
  • the recognition process can vary in specificity. For instance, in its most extreme form, plants can distinguish subspecies or races of pathogenic organisms.
  • Thionins are highly abundant polypeptides with anti fungal activities. These polypeptides are located in cell walls of leaf cells and the synthesis of thionine mRNA was increased after fungal attack (Bohlmann et al., 1988). Expression of the a-thionin gene from barley in transgenic tobacco has been shown to confer resistance to bacterial pathogens (Carmona et al., 1993). The in vitro toxicity against plant pathogenic bacteria and fungi indicates the role of thionin in the resistance of plants to the said bacteria and fungi. The induction of the thionin gene in response to salt stress identifies this gene with the osmome of the complement essential for tolerance towards osmotic dessication. The usefulness of the emergence of the thionin gene under salt stress lies in the fact that the rice plant harbouring one or several anti fungal disease genes can control the fungal diseases, thereby minimising the use of chemical fungicides.
  • mRNA purification was performed by first, isolating high quality total RNA from 6 day old RASI seedlings and, subsequently by isolating mRNA from total RNA using oligo (dT) cellulose in a filter syringe by making use of a double purification method.
  • mRNA was converted into first and second strand cDNA followed by Sal I adapter addition, Not I digestion, cDNA vector ligation and transformation to obtain the cDNA library.
  • the superscript TM plasmid system with Gateway TM for cDNA cloning and synthesis was employed throughout. 4.
  • the clones obtained were picked, digested using Not I and Sal I enzymes, to obtain the inserts and these were further sequenced and checked for homology. 5.
  • the sequencing of the selected clones was done on ABI Prism, 377, DNA Sequencer (Perkin Elmer).
  • AGPs using recombinant gene technology ensures (a) a method of supplying AGP that is independent of harvesting or fermentation requirements and problems, (b) that enables high levels of quality control, (c) that provides a supply of substantially pure AGP product, (d) that permits an overproduction of AGP in a host cell, and (e) that can be adapted to produce a specifically engineered AGP having desired properties.
  • this invention provides a means for supplying the functions and utilities of plant gums, e.g., gum arabic, etc., without the need for finding renewable but shrinking natural sources of plant gums.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Molecular Biology (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biophysics (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Biomedical Technology (AREA)
  • Biochemistry (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Physics & Mathematics (AREA)
  • Microbiology (AREA)
  • Plant Pathology (AREA)
  • Cell Biology (AREA)
  • Botany (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Medicinal Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Agricultural Chemicals And Associated Chemicals (AREA)

Abstract

La présente invention concerne l'isolation, dans des plantes, de séquences d'acide nucléique dont les produits confèrent aux plantes une résistance à divers stress biotiques et abiotiques tels que les lésions, les infections par agents pathogènes et la sécheresse.
EP04787517A 2003-09-24 2004-09-17 Proteines conferant aux plantes une resistance au stress biotique et abiotique Withdrawn EP1708558A4 (fr)

Applications Claiming Priority (17)

Application Number Priority Date Filing Date Title
IN769CH2003 2003-09-24
IN779CH2003 2003-09-24
IN774CH2003 2003-09-24
IN777CH2003 2003-09-24
IN771CH2003 2003-09-24
IN778CH2003 2003-09-24
IN780CH2003 2003-09-24
IN776CH2003 2003-09-24
IN773CH2003 2003-09-24
IN782CH2003 2003-09-24
IN768CH2003 2003-09-24
IN767CH2003 2003-09-24
IN775CH2003 2003-09-24
IN781CH2003 2003-09-24
IN770CH2003 2003-09-24
IN772CH2003 2003-09-24
PCT/IB2004/003017 WO2005027631A2 (fr) 2003-09-24 2004-09-17 Proteines conferant aux plantes une resistance au stress biotique et abiotique

Publications (2)

Publication Number Publication Date
EP1708558A2 true EP1708558A2 (fr) 2006-10-11
EP1708558A4 EP1708558A4 (fr) 2008-03-19

Family

ID=34382374

Family Applications (2)

Application Number Title Priority Date Filing Date
EP04787517A Withdrawn EP1708558A4 (fr) 2003-09-24 2004-09-17 Proteines conferant aux plantes une resistance au stress biotique et abiotique
EP04787519A Withdrawn EP1705987A2 (fr) 2003-09-24 2004-09-17 Proteines ayant un potentiel nutraceutique et biotherapeutique

Family Applications After (1)

Application Number Title Priority Date Filing Date
EP04787519A Withdrawn EP1705987A2 (fr) 2003-09-24 2004-09-17 Proteines ayant un potentiel nutraceutique et biotherapeutique

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Country Link
EP (2) EP1708558A4 (fr)
WO (2) WO2005027632A2 (fr)

Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1708558A4 (fr) * 2003-09-24 2008-03-19 Avestha Gengraine Tech Pvt Ltd Proteines conferant aux plantes une resistance au stress biotique et abiotique
CA2681515A1 (fr) * 2007-03-23 2008-10-02 Basf Plant Science Gmbh Plantes transgeniques offrant une amelioration de la tolerance au stressamelioree et du rendement exprimant une proteine lrp-2
DE102008041695A1 (de) * 2008-08-29 2010-03-04 Bayer Cropscience Ag Methoden zur Verbesserung des Pflanzenwachstums
WO2015054541A1 (fr) * 2013-10-11 2015-04-16 Portland State University Production de plantes résistantes via l'amélioration transgénérationnelle des défenses des plantes
CN104306989A (zh) * 2014-09-27 2015-01-28 郭和友 一种利用切割酶治疗细菌方法

Family Cites Families (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1329663A (zh) * 1998-10-09 2002-01-02 吉尼西斯研究及发展有限公司 改变植物木质素含量的材料和方法
US6433249B1 (en) * 1998-11-10 2002-08-13 Pioneer Hi-Bred International, Inc. Use of β-glucosidase to enhance disease resistance and resistance to insects in crop plants
WO2000075159A1 (fr) * 1999-06-03 2000-12-14 Cornell Research Foundation, Inc. EXO-β 1,3 GLUCOSIDASE FONGIQUE CODANT POUR UNE MOLECULE D'ADN ET SON UTILISATION COMME AGENT ANTI-FONGIQUE POUR VEGETAUX
JP4755769B2 (ja) * 2001-03-14 2011-08-24 独立行政法人理化学研究所 植物に対するストレス耐性付与方法
US20070011772A1 (en) * 2002-03-06 2007-01-11 Max-Planck-Gesellschaft Zur Forderung Der Wissenschaften E.V. Polynucleotides encoding a beta-glucosidase and uses thereof
EP1708558A4 (fr) * 2003-09-24 2008-03-19 Avestha Gengraine Tech Pvt Ltd Proteines conferant aux plantes une resistance au stress biotique et abiotique

Also Published As

Publication number Publication date
WO2005027632A2 (fr) 2005-03-31
EP1708558A4 (fr) 2008-03-19
EP1705987A2 (fr) 2006-10-04
WO2005027631A2 (fr) 2005-03-31
WO2005027631A3 (fr) 2006-08-31

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