Multicellular living organisms and unmodified parts thereof and – Method of introducing a polynucleotide molecule into or... – The polynucleotide confers pathogen or pest resistance
Reexamination Certificate
1999-05-26
2001-09-25
Fox, David T. (Department: 1638)
Multicellular living organisms and unmodified parts thereof and
Method of introducing a polynucleotide molecule into or...
The polynucleotide confers pathogen or pest resistance
C800S278000, C800S298000, C800S306000, C800S317000, C800S301000, C435S069100, C435S320100, C435S419000, C435S468000, C536S023100, C536S023600
Reexamination Certificate
active
06294712
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention concerns a nucleic acid, which induces a resistance against sedentary nematodes in plants, preferably of the Solanaceae and/or Chenopodiaceae and/or the Brassicaceae families, especially preferably of the genus Beta and/or Brassica and/or Solanum.
The present invention additionally concerns the DNA sequence of a cDNA clone and a genomic clone of this nucleic acid. Further, the present invention concerns a vector, which can for example be a yeast artificial chromosome “YAC”, which contains the nucleic acid for a resistance against sedentary nematodes in plants. Finally, the present invention concerns the use of the nucleic acid or of the vector for the induction of a resistance against sedentary nematodes in plants, and also a transgenic plant, which contains the nucleic acid or the vector.
Further, the invention concerns the protein encoded by the nucleic acid, a test kit containing the nucleic acid and/or the vector and a process for producing a transgenic plant and also a process for producing a nematode resistance in plants.
Finally, the invention concerns the promoter of the resistance gene.
2. Description of Related Art
It is well known that plants are attacked by various pathogens and types of parasite. It is also well known that crop plants are mostly more susceptible to parasitic attack than their wild relatives. Often encountered are plant parasites from the nematode family, with a fluid-filled pseudocyloma sheath surrounded by a musculocutaneous sac. Nematodes are important parasites, which in the harvests throughout the world cause losses of ca 150 million DM per year. Particularly damaging are nematodes of the genera Meloidogyne, Heterodera and Globodera, which establish themselves permanently in the roots of the affected plants, after they have induced certain feeding structures. The nematode
Heterodera schachtii
has a broad host spectrum, which includes many species of various plant families, e.g. the Chenopodiaceae and Brassicaceae.
The life cycle of nematodes is subdivided into four larval stages (J1-J4). The roots are infected by J2 juvenile stages, which migrate to the central cylinder, where they induce the development of syncytia. These extensive feeding structures result from a partial cell-wall degradation between the cells of the xylem parenchyma. The nematode ends its life-cycle to the adult stage after three periods. The female nematodes swell up and finally destroy the root cortex, while they are still feeding from the syncytia. The male stages no longer feed after the end of the third stage, and when they are adult they move towards the female stages, by which they are attracted by sex pheromones. The mature female stages are filled with eggs. After their death, they form a cyst, in which the infectious larvae (J2) can survive in the soil for up to 10 years.
It is presumed that nematode resistance genes trigger an incompatibility reaction between the host and the parasites, which has already been described at the cellular level. The roots of plants which carry this or these gene(s) are admittedly attacked by J2 juvenile stages, but most of the nematodes die in the late J2 stage because of degradation of the initiated syncytium. In rare cases, female stages can develop, however they display a transparent appearance and cease growing. As a result, the nematodes are not able to complete their life cycle.
Since for environmental political reasons the use of nematicides is only possible to a limited extent, it is particularly desirable also to implement this resistance gene in crop plants.
In particular, root crops of the genus Beta (e.g. sugar-beet, fodder roots, mangold, beetroot) are highly susceptible to the root cyst nematode
Heterodera schachtii.
Efforts have already long been made to create resistance against
Heterodera schachtii
and other phytopathogenic nematodes (e.g. Globodera) in plants, in particular crop plants, since resistance genes corresponding to these are lacking. The only sources of resistance are the wild species
Beta procumbens
and its close relatives
B. webbiana
and
B. natellaris.
Genes for resistance against various nematode species are used in breeding in different useful plant species (e.g. potato, tomato, wheat, oil radish). A resistance gene from the wild species
Beta procumbens
has also been transferred into the sugar-beet by cross-breeding. From this, resistant sugar-beets could be selected; however, they had the disadvantage that they were characterised by inadequate quality and productivity properties. The resistant sugar-beet lines which derived from the cross-breeding with
Beta procumbens
have translocations of varying size from the wild beets of the Procumbentes section. Their low productivity and decreased quality are presumably due to the fact that as well as the resistance gene other productivity-decreasing genes from the wild species are present in these sugar-beet lines. Also, the transmission of the resistance property to subsequent generations is incomplete. These disadvantages cannot be eliminated purely by breeding methods, since breeding by crossing is unlikely to be able to select specific properties, without other, sometimes disadvantageous, properties also being transferred at the same time.
Further, many attempts have been made to induce artificial nematode resistance in plants by a combination of “suicide genes” with syncytia-specific promoters. So far, however, it has not been possible to breed any resistant plants from this.
Moreover, it has not previously been possible to identify a naturally resistant gene at the molecular level and to use it for the creation of a resistance in crop plants.
SUMMARY OF THE INVENTION
It is, therefore, one objective of the present invention to provide a nucleic acid which imparts a resistance against nematodes to plants. It is a further object of the present invention to provide the DNA sequence on which this gene is based.
Also, it is a further object according to the present invention to enable the use of such a gene for the induction of a resistance against sedentary nematodes.
Apart from this, it is an object according to the present invention to provide a transgenic plant which contains a nucleic acid imparting such resistance, and cells, seeds or plant parts which contain this nucleic acid.
Finally, it is an object of the present invention to provide vectors in which the gene for imparting resistance against nematodes can be effectively incorporated into plants and is contained therein.
It is a further object according to the present invention to provide the protein encoded by the nucleic acid and a test kit which contains the nucleic acid.
Finally, it is an object according to the present invention to provide a process for the production of a transgenic plant and a process for producing a resistance against nematodes.
It is also an object of the present invention to provide the promoter which controls the expression of the aforesaid resistance gene.
These and other objectives are achieved by the subject matter of the invention contained in the claims.
REFERENCES:
patent: 5750386 (1998-05-01), Conkling et al.
patent: WO 93/19181 (1993-09-01), None
patent: WO 92/15690 (1993-09-01), None
Salentija et al. Mol. Gen. Genet. vol. 235, pp. 432-440, 1992.*
Lindsey et al. Journal of Exp. Botany, vol. 41, No. 226, pp. 529-536, 1990.*
Sambrook J., et al. Molecular Cloning, A laboratory manual, 2nd edition, Cold Spring Harbor Laboratory Press, 1989.*
Lagudah et al.; “Map-Based Cloning of a Gene Sequence Encoding a Nucleotide-Binding Domain and a Leucine-Rich Region at the CRE3 Nematode Resistance Locus of Wheat”; Genome; vol. 40; 1997; pp. 659-665.
Kleine et al.; “Physical Mapping and Cloning of a Translocation in Sugar Beet (Beta Vulgaris L.) Carrying a Gene for Nematode (Heterodera Schachtii) Resistance from B. Procumbens”; Theor Appl. Genet; vol. 90; 1995; pp. 399-406.
Kleine et al.; Breeding for Nematode Resistance in Sugarbeet: A Molecular Approach; Klawer Academic Publishers; 1
Cai Daguang
Jung Christian
Kleine Michael
Foley & Lardner
Fox David T.
Ibrahim Medina A.
Jung Christian
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