Disease resistance in vitis

Details Disease resistance in vitis Disease resistance in vitis

1997-06-19

2001-05-15

Hutzell, Paula (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

C514S002600, C800S278000, C800S280000, C800S290000, C800S292000, C800S293000, C800S294000, C800S295000, C800S298000, C800S301000, C435S069100, C435S410000, C435S419000

Reexamination Certificate

active

06232528

ABSTRACT:

BACKGROUND OF THE INVENTION
This application relates to disease resistance in Vitis.
Grape (Vitis spp.) is a deciduous temperate fruit crop of ancient origin. Grape production (65 ×10
6
metic tons) exceeds that of any other temperate fruit crop and ranks after Citrus and banana among all fruit crops worldwide (FAO Production Yearbook, 1990). Grape surpasses all other fruit crops in value due to its multiple uses for fresh fruit, juice, jelly, raisins, and wine. For example, in the United States, seedless grapes represent about 80% and 98% of the total table and raisin grape production; respectively (
In:
1994-95:
The Distribution and Per Capita Consumption of California Table Grapes By Major Varieties in the United States and Canada
, California Table Grape Commission, Fresno, Calif. 1995). Only a few seedless cultivars make up this production, of which ‘Thompson Seedless’ is the most important. This cultivar accounts for the most production of any single grape variety in the United States. In 1992, ‘Thompson Seedless’ was grown on 263,621 acres in California (
In: California Grape Acreage
, California Agricultural Statistics Service, Sacramento, Calif., 1993). Thirty-five percent of the table grape production in California in 1994 was ‘Thompson Seedless’ (23,244,683 boxes, 10 kg/box). In 1993, 97% of the grapes grown for raisin production was ‘Thompson Seedless’ (
In: Raisin Committee Marketing Policy
1994-95, Raisin Administrative Committee, Fresno, Calif., 1994).
Although Vitis spp. is generally considered to have desirable fruit quality, it is susceptible to many pests and diseases, including anthracnose, black rot, botrytis bunch rot, crown gall, downy mildew, eutypa dieback, various nematodes, phomopsis cane and leaf spot, phylloxera, Pierce's disease, and powdery mildew. Hybridization with resistant species has been the only method available to produce resistant cultivars (Galet and Morton,
In: Compendium of Grape Diseases
, R. C. Pearson and A. C. Goheen, eds., APS Press, St. Paul, 1990, pp. 2-3). While improving grape is possible by conventional breeding, it is difficult and time consuming due to the two- to three-year generation cycle, the long period of time required for reliable progeny testing and selection, and inbreeding depression that prohibits selfing (Gray and Meredith,
In: Biotechnology of Perennial Fruit Crops
, F. A. Hammerschlag and R. E. Litz, eds., C.A.B. Intl., Wallingford, U.K. 1992). These characteristics make introgression of desirable traits into existing grape cultivars difficult if not impossible to achieve in an individual breeder's lifetime. Thus, the alternative, and potentially less time-consuming, approach of using gene transfer to insert desirable genes is one approach for improving grapevine cultivars, even considering the time necessary for field testing transgenic lines. The ability to improve the disease or pest resistance or both of a major grape cultivar (e.g., ‘Thompson Seedless’) offers the possibility of improving a large portion of the grape production in a relatively short time, assuming that cultivar integrity would not be compromised by the transgene or the insertion event. Such a change could also reduce pesticide use for a significant portion of grape production.
SUMMARY OF THE INVENTION
In one aspect, the invention features a method for producing a transgenic plant of the genus Vitis having resistance to a plant pathogen. The method, in general, includes the step of transforming a plant cell with a nucleic acid which expresses a lytic peptide, where the expression of such a lytic peptide provides resistance to a plant pathogen. In preferred embodiments, the method further includes propagating a grape plant from the transformed plant cell. In other preferred embodiments, the method involves transforming a plant cell that is a part of a somatic embryo and propagating or regenerating a transgenic grape plant from the transformed somatic embryo. Expression of the lytic peptide confers disease resistance or tolerance or both to grapevine pathogens and pests including, without limitation, bacterial, fungal, and viral pathogens.
In general, Vitis is transformed by introducing into a plant cell or somatic or zygotic embryos a nucleic acid that includes a lytic peptide by using
A. tumefaciens
, microprojectile bombardment, or any combination of these methods (for example, by bombarding the plant cell with microprojectiles, followed by infecting the bombarded cells with
Agrobacterium tumefaciens
including a nucleic acid which expresses the lytic peptide).
In preferred embodiments, the method of the invention involves the use of the lytic peptides Shiva-1 or cecropin B or both.
The methods of the invention are useful for providing disease resistance or tolerance or both to a variety of grape plants (for example, Vitis spp., Vitis spp. hybrids, and all members of the subgenera Euvitis and Muscadinia), including scion or rootstock cultivars. Exemplary scion cultivars include, without limitation, those which are referred to as table or raisin grapes and those used in wine production such as Cabernet Franc, Cabernet Sauvignon, Chardonnay (e.g., CH 01, CH 02, CH Dijon), Merlot, Pinot Noir (PN, PN Dijon), Semillon, White Riesling, Lambrusco, Thompson Seedless, Autumn Seedless, Niagrara Seedless, and Seval Blanc. Rootstock cultivars that are useful in the invention include, without limitation,
Vitis rupestris
Constantia,
Vitis rupestris
St. George,
Vitis california, Vitis girdiana, Vitis rotundifolia, Vitis rotundifolia
Carlos, Richter 110 (
Vitis berlandieri x rupestris
), 101-14 Millarder et de Grasset (
Vitis riparia x rupestris
), Teleki 5C (
Vitis berlandieri x riparia
), 3309 Courderc (
Vitis riparia x rupestris
), Riparia Gloire de Montpellier (
Vitis riparia
), 5BB Teleki (selection Kober,
Vitis berlandieri x riparia
), SO
4
(
Vitis berlandieri x rupestris
), 41 B Millardet (
Vitis vinifera x berlandieri
), and 039-16 (
Vitis vinifera x Muscadinia
).
In another aspect, the invention features a transgenic plant or plant cell of the genus Vitis transformed with a nucleic acid which expresses a lytic peptide, wherein expression of the lytic peptide provides resistance to a plant pathogen. In preferred embodiments, the transgenic grapevine or cell with the nucleic acid includes an expression vector. Preferably, the transgenic grapevine or cell is
Vitis vinifera
‘Thompson Seedless’ and the expression of the lytic peptide provides resistance to the bacterium
Xylella fastidiosa
, the causative agent of Pierce's Disease. In other preferred embodiments, the transgenic grapevine is a somatic embryo, a scion, or a rootstock.
In still another aspect, the invention features a method of transforming Vitis with a nucleic acid which expresses a tomato ringspot virus coat protein (TomRSV-CP) gene, where the expression of such a coat protein gene provides resistance to a plant pathogen.
In still another aspect, the invention features a method of transforming Vitis with a nucleic acid which expresses a TomRSV-CP gene and a lytic peptide gene, where the expression of such genes in a grapevine provides resistance to a plant pathogen.
The invention also features scions, rootstocks, somatic or zygotic embryos, cells, or seeds that are produced from any of the transgenic grape plants described herein.
By “lytic peptide” is meant a gene encoding a polypeptide capable of lysing a cell. Exemplary lytic peptides include, without limitation, apidaceins, attacins, cercropins (e.g., cercropin B), caerulins, bombinins, lysozyme, magainins, melittins, sapecin, sarcotoxins, and xenopsins.
By “peptide” is meant any chain of amino acids, regardless of length or post-translational modification (for example, glycosylation or phosphorylation).
By “positioned for expression” is meant that the DNA molecule is positioned adjacent to a DNA sequence which directs transcription and translation of the sequence (i.e., facilitates the production of, for example, a lytic peptide).
By “operably linked” is meant that a gene and a regulatory s

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