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WO1993012239A1 - Modification des caracteristiques des plantes et de leur croissance cellulaire - Google Patents

Modification des caracteristiques des plantes et de leur croissance cellulaire Download PDF

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Publication number
WO1993012239A1
WO1993012239A1 PCT/GB1992/002340 GB9202340W WO9312239A1 WO 1993012239 A1 WO1993012239 A1 WO 1993012239A1 GB 9202340 W GB9202340 W GB 9202340W WO 9312239 A1 WO9312239 A1 WO 9312239A1
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WO
WIPO (PCT)
Prior art keywords
gene
cdc25
plant
plants
genome
Prior art date
Application number
PCT/GB1992/002340
Other languages
English (en)
Inventor
John Charles Ormrod
Dennis Francis
Mark Henri Bell
Original Assignee
Zeneca Limited
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 Zeneca Limited filed Critical Zeneca Limited
Publication of WO1993012239A1 publication Critical patent/WO1993012239A1/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)
    • C12N15/8241—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
    • C12N15/8262—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield involving plant development
    • C12N15/827—Flower development or morphology, e.g. flowering promoting factor [FPF]
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/37—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from fungi
    • C07K14/39—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from fungi from yeasts
    • 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
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/04—Plant cells or tissues

Definitions

  • This invention relates to plants and plant cells with altered growth characteristics and to a method for producing such alterations. More particularly, but not exclusively, the invention relates to plants which exhibit precocious flowering.
  • SUBSTITUTESHEET activity of a cell cycle protein The protein identified therein is known as p34 c and the level of that protein may be modulated directly by action on the gene itself or indirectly by action taken against one of a number of regulatory genes which regulate p34 c c . Examples of such regulatory genes are given as pl3suc , nim-1, wee-1, mik-1 and cdc25. These genes have long been known to exist in yeast but whether they have equivalents in the plant kingdom had remained a matter for speculation. The inventors of WO 92/09685 confirm that pl3 B does have a homologue in plants.
  • An object of the present invention is to provide plants with altered growth characteristics.
  • a method for altering the growth characteristics of plants and plant cells comprising incorporating into the genome of the plant or plant cell by genetic transformation a cdc25 gene or a gene of homologous function.
  • the said gene is the cdc25 gene or cdc25 homologue is derived from a yeast, for example, the yeast Schizosaccharomyces pombe.
  • the said cdc25 gene or cdc25 homologue may be placed under the control of a 35S promoter of Cauliflower Mosaic Virus (CaMV35S).
  • CaMV35S Cauliflower Mosaic Virus
  • the said gene may be placed under the control of a chemically inducible promoter enabling expression of the gene to be induced by external application of a chemical inducer.
  • the invention further provides a recombinant plant genome having stably incorporated within its genome an exogenous cdc25 gene or a functional homologue thereof which, preferably, is derived from a yeast such as Schizosaccharomyces pombe.
  • the invention also provides genetically modified plants which possess the ability to flower earlier than in the the unmodified form.
  • Whole plants with altered morphology, reflecting changes at the cellular level, may be produced by regeneration of plants from the said transformed cells.
  • the genetic alteration of the genome leads to improvements in the flowering characteristics such as precocious flowering and larger numbers of flowers.
  • the inserted gene may be derived from non— lant organisms which possess such genes or equivalents .
  • the gene be derived from the fission yeast Schizosaccharomyces pombe.
  • Functional equivalents of the cdc25 gene are those which participate in the control of expression of the p34cdc2 protein, for example weel, niml and sucl.
  • GSTII glutathione-S- transferase II
  • promoters can be used in this scenario. Some of these may be of plant origin, others may be of fungal, bacterial or yeast origin. It is implied in the present application that those promoters and chemical combinations suitable for the plant growth control procedure can be used in place of GSTII and safeners.
  • alcR activator gene and the alcA target promoter from Aspergillus.
  • the chemical inducer is cyclohexanone.
  • Another example of a chemically inducible gene is given in European Patent Application EP-A-0332104 (Ciba-Geigy) .
  • the cdc25 gene or its homologue By placing the cdc25 gene or its homologue under the control of such an inducible gene switch, expression of the gene may be controlled at will by the application of the appropriate chemical inducer to the plant. In the absence of the inducer, the gene does not express and when the inducer is applied the expression is switched on. In the context of this invention, then, the modified plant may be allowed to grow normally until an appropriate stage of its development at which the inducer may be applied and the benefit of the presence of the cdc25 or homologous gene may be obtained.
  • One application is to apply the inducer only at the stage of seed development in order to alter the characteristics of the seed without substantially altering the plant stature or morphology.
  • An alternative method of obtaining such effects is to place the inserted- gene under the control of a tissue or organ specific promoter to direct expression to the tissue or organ which is selected for modification, or under control of a development-regulated promoter to restrict
  • the cell division cycle in plants appears to be regulated at key steps by a few genes, although exactly how the gene products interact to regulate cell division is poorly understood.
  • the product of cdc2, p34 c c plays an important role in both the onset of mitosis and the onset of DNA synthetic (or S-) phase, although there is evidence to indicate that the protein may exist in " two forms, one related to S-phase and one to mitosis.
  • the gene product, p34cdc2 is the catalytic sub-unit of the protein kinase MPF (maturation promoting factor), the other two components being the gene products of cdcl3 (a "cyclin").
  • At least three genes interact with the "cdc2 pathway" to regulate the onset of mitosis.
  • the product of cdc25, p80cdc25, is required for the dephosphorylation and activation of p34cdc2.
  • Weel is a negative regulator of the cdc2 protein kinase, delaying entry into mitosis. Nim acts to suppress weel, that is, it is also an activator of mitosis.
  • Cdc25, weel and niml are all dose-dependent genes, and therefore even if homologous sequences exist in plants, overexpression of any one of these genes may be expected to alter some aspect of cell division in a plant system.
  • a cassette containing cdc25 from Schizosaccharomyces pombe was obtained from Professor Paul Nurse FRS (ICRF Cell Cycle Group,
  • Regenerated tobacco plants were obtained following leaf disc transformation using Agrobacterium tumefaciens and the binary vector pBinl9. Southern blot analysis confirmed that the regenerated plants did contain the cdc25 construct. Transformed plants have shown several differences in phenotype compared with the wild-type. In particular, with the constitutive promoter (CaMV35S) the primary transformants are considerably dwarfed, as are the HMW promoter transformants but to a lesser degree.
  • the second phenotypic change of the CaMV transformants is in leaf character, in particular the leaves appear wrinkled and pocketed, as well as exhibiting altered shape.
  • the third phenotypic change is an increase
  • the presence of the CaMV/cdc25 construct does not affect seed viability.
  • the principal phenotypic change in transformants containing the cdc25 gene under control of the endosperm-specific HMW promoter is the size and viability of the seed, although there is some variability between and within pods. Much of the seed recovered is much smaller than that from the wild type; this is probably because of reduced endosperm size, although the endosperm of tobacco is relatively small. Seed viability varies from 9.4% in some pods to over 90% in pods from the same plant.
  • Figure 5 is a graph showing the relative frequencies of occurrence of a range of cells sizes (plotted as the logarithm) .
  • the statistical test used here is the Kolmogorov-Smirnoff two-sample test which is a nonparametric test to show whether two independent samples have been drawn from the same population. What the statistics show is strong evidence that the two distributions are significantly different at the 1% level. Flowering Characteristics
  • SUBSTITUTE SHEET expressed cassette CaMV-cdc25-nos were germinated and 80 plants potted on. Similarly, 80 plants from a backcross of one of the lines to wild-type (TlBCl) were grown up. There was no selection of germinating seed on kanamycin and, therefore, there should have been a predictable segregation of the T-DNA kanamycin marker in the T1S1 line between homozygotes (KK) and heterozygotes (KO) of 25% KK, 50% KO and 25% 00. The Mendelian segregation would result in a ration of 3:1.
  • Leaf discs were taken from surface sterilised leaves with a cork borer and placed on 200/t/g/ml kanamycin water agar plates. This was carried out at 67 days after potting out of the seedlings. At this point two plants in line D had still not flowered as observed at 45 days.

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

Abstract

Des plantes ayant une taille et d'autres caractéristiques phénotypiques modifiées, dont la plus avantageuse est une floraison précoce et un nombre accru de fleurs, sont produites par la transformation du génome de la plante avec le gène cdc25 ou un homologue fonctionnel de celui-ci. La source préférée de ces gènes est la levure fissipare Schizosaccharomyces pombe.
PCT/GB1992/002340 1991-12-18 1992-12-17 Modification des caracteristiques des plantes et de leur croissance cellulaire WO1993012239A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
GB9126818.5 1991-12-18
GB919126818A GB9126818D0 (en) 1991-12-18 1991-12-18 Alteration of plant and plant cell morphology

Publications (1)

Publication Number Publication Date
WO1993012239A1 true WO1993012239A1 (fr) 1993-06-24

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PCT/GB1992/002340 WO1993012239A1 (fr) 1991-12-18 1992-12-17 Modification des caracteristiques des plantes et de leur croissance cellulaire

Country Status (3)

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AU (1) AU3165793A (fr)
GB (1) GB9126818D0 (fr)
WO (1) WO1993012239A1 (fr)

Cited By (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1995024487A1 (fr) * 1994-03-09 1995-09-14 Hoechst Schering Agrevo Gmbh Procedes d'inhibition et de declenchement de la floraison des plantes
WO1997004116A1 (fr) * 1995-07-24 1997-02-06 Zeneca Limited Inhibition de la respiration cellulaire et production de plantes males steriles
WO1997025433A1 (fr) * 1996-01-09 1997-07-17 Eidg. Technische Hochschule Zürich Ethz Controle de la floraison de plantes
EP0784425A4 (fr) * 1994-10-14 1998-01-14 Univ Washington Regulation genique du developpement floral et de la dominance apicale dans des plantes
WO2000052171A1 (fr) * 1999-02-26 2000-09-08 Cropdesign N.V. Procede de modification de la morphologie, biochimie ou physiologie de plantes, a l'aide de cdc25
WO2000052168A1 (fr) * 1999-02-26 2000-09-08 Cropdesign N.V. Procede de selection de cellules et tissus transformes
WO2000037645A3 (fr) * 1998-12-23 2000-11-09 Pioneer Hi Bred Int Acides nucleiques intervenant dans le cycle cellulaire, polypeptides et leurs utilisations
WO2001023594A3 (fr) * 1999-09-27 2001-12-06 Pioneer Hi Bred Int Amelioration de la tolerance au stress dans le mais par manipulation des genes de regulation du cycle cellulaire

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1992009685A1 (fr) * 1990-11-29 1992-06-11 The Australian National University Procede de regulation de la proliferation et de la croissance des cellules vegetales

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1992009685A1 (fr) * 1990-11-29 1992-06-11 The Australian National University Procede de regulation de la proliferation et de la croissance des cellules vegetales

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
BIOLOGICAL ABSTRACTS vol. 90 , 1990, Philadelphia, PA, US; abstract no. 15284, OONO, Y., ET AL. 'Early flowering in transgenic tobacco plants possessing the rolC gene of Agrobacterium-rhizogenes Ri plasmid' *
CELL vol. 45, 11 April 1986, CAMBRIDGE, MA US pages 145 - 153 RUSSELL, P., ET AL. 'cdc25+ functions as an inducer in the mitotic control of fission yeast' *
PLANT MOLECULAR BIOLOGY. vol. 14, 1990, DORDRECHT, THE NETHERLANDS. pages 669 - 685 HILSON, P., ET AL. 'Yeast RAS2 affects cell viability, mitotic division and transient gene expression in Nicotiana species' *

Cited By (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1995024487A1 (fr) * 1994-03-09 1995-09-14 Hoechst Schering Agrevo Gmbh Procedes d'inhibition et de declenchement de la floraison des plantes
EP0784425A4 (fr) * 1994-10-14 1998-01-14 Univ Washington Regulation genique du developpement floral et de la dominance apicale dans des plantes
WO1997004116A1 (fr) * 1995-07-24 1997-02-06 Zeneca Limited Inhibition de la respiration cellulaire et production de plantes males steriles
WO1997025433A1 (fr) * 1996-01-09 1997-07-17 Eidg. Technische Hochschule Zürich Ethz Controle de la floraison de plantes
WO2000037645A3 (fr) * 1998-12-23 2000-11-09 Pioneer Hi Bred Int Acides nucleiques intervenant dans le cycle cellulaire, polypeptides et leurs utilisations
WO2000052171A1 (fr) * 1999-02-26 2000-09-08 Cropdesign N.V. Procede de modification de la morphologie, biochimie ou physiologie de plantes, a l'aide de cdc25
WO2000052168A1 (fr) * 1999-02-26 2000-09-08 Cropdesign N.V. Procede de selection de cellules et tissus transformes
WO2001023594A3 (fr) * 1999-09-27 2001-12-06 Pioneer Hi Bred Int Amelioration de la tolerance au stress dans le mais par manipulation des genes de regulation du cycle cellulaire

Also Published As

Publication number Publication date
AU3165793A (en) 1993-07-19
GB9126818D0 (en) 1992-02-19

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