Peronospora resistance in Spinacia oleracea

Abstract

The present invention relates to a spinach plant which may comprise a single dominant gene which confers resistance to Peronospora farinosa f. sp. spinaciae races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410, wherein the gene is obtainable by introgression from a plant grown from seeds of which a representative sample was deposited with the NCIMB under NCIMB accession number 41857. The invention further relates to progeny of the plant, to propagation material therefore, such as seed, and to food products which may comprise the spinach leaves.

Claims

1. A method of producing a hybrid spinach seed comprising crossing a first parent spinach plant with a second parent spinach plant and harvesting the resultant hybrid spinach seed, wherein said first parent spinach plant comprises a R6 gene that confers resistance to a spinach plant to downy mildew races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410 and that requires stacking with another downy mildew resistance gene for the spinach plant to have resistance to downy mildew races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs7, Pfs8, Pfs9, Pfs10, Pfs11, Pfs12, Pfs13 and UA4410, wherein the R6 gene is as found in a plant grown from seed deposited with the NCIMB under NCIMB accession number 41857.

2. The method of claim 1 wherein the second parent spinach plant comprises the R6 gene.

3. The method of claim 1 wherein the second parent spinach plant comprises another downy mildew resistance gene whereby a plant grown from the seed produced by the method has resistance to downy mildew races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs7, Pfs8, Pfs9, Pfs10, Pfs11, Pfs12, Pfs13 and UA4410.

4. A hybrid spinach seed, a plant grown from which having resistance to downy mildew races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs7, Pfs8, Pfs9, Pfs10, Pfs11, Pfs12, Pfs13 and UA4410 produced from the method of claim 3.

5. A hybrid spinach plant grown from the spinach seed of claim 4, having resistance to downy mildew races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs7, Pfs8, Pfs9, Pfs10, Pfs11, Pfs12, Pfs13 and UA4410.

6. A method of conferring resistance to a spinach plant not having resistance against Peronospora farinosa f. sp. spinaciae races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410 comprising crossing a first parent spinach plant with a second parent spinach plant, wherein said first parent spinach plant comprises a R6 gene that confers resistance to the spinach plant to downy mildew races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410 and that requires stacking with another downy mildew resistance gene for the spinach plant to have resistance to downy mildew races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs7, Pfs8, Pfs9, Pfs10, Pfs11, Pfs12, Pfs13 and UA4410, and said second parent spinach plant does not contain the R6 gene, wherein the R6 gene is as found in a plant grown from seed deposited with the NCIMB under NCIMB accession number 41857.

7. The method of claim 6 wherein the second parent spinach plant comprises another resistance gene whereby a plant grown from the seed produced by the method has resistance to downy mildew races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs7, Pfs8, Pfs9, Pfs10, Pfs11, Pfs12, Pfs13 and UA4410.

Description

EXAMPLES

Example 1

Testing for the R6 Resistance Trait in Spinach Plants

(1) The resistance to downy mildew infection was assayed as described by Irish et al. (2008; Phytopathol. 98: 894-900), using a differential set. Spinach plants of the invention (R6) were planted along with spinach plants from different other genotypes (see Table 1) in trays containing Scotts Redi-Earth medium, and fertilized twice a week after seedling emergence with Osmocote Peter's (13-13-13) fertilizer (Scotts). Plants were inoculated with a sporangial suspension (2.510.sup.5/ml) of a pathogenic race of Peronospora farinosa f. sp. spinaciae at the first true leaf stage. In this manner, 11 pathogenic races were tested (as shown in Table 1).

(2) The inoculated plants were placed in a dew chamber at 18 C. with 100% relative humidity for a 24 h period, and then moved to a growth chamber at 18 C. with a 12 h photoperiod for 6 days. After 6 days, the plants were returned to the dew chamber for 24 h to induce sporulation, and they were scored for disease reaction.

(3) Plants were scored as resistant or susceptible based on symptoms of chlorosis and signs of pathogen sporulation on the cotyledons and true leaves, as described by Irish et al. (2007; Plant Dis. 91: 1392-1396). Plants exhibiting any evidence of chlorosis and sporulation were considered susceptible. Resistant plants were re-inoculated to assess whether plants initially scored as resistant had escaped infection, or whether they were truly resistant.

(4) Table 1 shows the differential set of spinach downy mildew races and the resistance of various spinach varieties (hybrids) to each one of these pathogenic races. A susceptible reaction is scored as + (indicating a successful infection by the fungus, with sporulation occurring on the entire cotyledon), and resistance is depicted as (absence of sporulation on the cotyledons). A weak resistance response is indicated as (+), which in practice means a slightly reduced level of infection (with sporulation only occurring on the tips of the cotyledons in the differential seedling test). R6 is a line exhibiting the resistance of the present invention, and the resistance patterns of the parental lines of hybrid variety Lion are also shown.

(5) Comparison of the parental lines of Lion to Lion itself reveals that the broad resistance pattern of Lion results from the combination of at least two resistance genes, coming from either of the parents, because both parents only possess parts of the resistance profile of the hybrid (Lion) that results from the crossing of these two lines. The genetic basis of the resistance in Lion is thus multigenic in nature, caused by the stacking of at least two resistance genes in the hybrid variety, and hence the genetic basis of the Peronospora resistance in Lion is entirely different from that in plants of the present invention.

(6) In contrast, the R6 resistance trait of the present invention is conferred by a single dominant locus, which has the great advantage that the R6 resistance trait may be easily transferred to other spinach varieties by crossing/introgression, and that it may easily be combined with other resistance genes. When combined with selected other genes that e.g. confer resistance to downy mildew races Pfs7, Pfs8 and Pfs10, the R6 trait may be used to provide resistance to all downy mildew races known to date in spinach.

(7) TABLE-US-00001 TABLE 1 Viro- Resisto- Lion male Lion female Race flay flay Califlay Clermont Campania Boeing Lion Lazio Whale Polka Pigeon R6 parent parent Pfs1 + Pfs2 + + + Pfs3 + + Pfs4 + + + + + Pfs5 + + + Pfs6 + + + + + + + Pfs9 + + + + Pfs11 + + + + Pfs12 + + + + + + + Pfs13 + + (+) + (+) + + (+) (+) UA4410 + + + + + + + +

Example 2

Introduction of the R6 Resistance Trait into Other Spinach Plants

(8) A plant of the invention was crossed (as a father) with a plant that does not contain the R6 resistance trait, to obtain an F1. Thirty plants of the F1 population were tested for resistance to Peronospora race UA4410, as described in example 1. This particular resistance was absent from the mother plant used in the said cross. All 30 plants showed the resistance pattern of the invention, i.e. resistance to pathogenic race UA4410. This demonstrated that the R6 resistance gene inherits in a dominant manner.

(9) In another experiment, a plant of the invention was crossed (as a mother) with a different spinach plant that does not contain the R6 resistance trait of the invention. Plants of the F1 population were selfed, and a total of 76 plants of the F2 generation were tested for Peronospora resistance, as described in example 1. As a positive discriminator for the presence of the R6 trait, resistance to Pfs11 was assayed, because this resistance was present in the mother plant (R6) but not in the father plant of the cross.

(10) It was observed that Pfs11 resistance segregated in the F2 generation in a fashion that corresponds to dominant monogenic inheritance: 60 of the 76 F2 plants exhibited the R6-resistance pattern. Table 2 gives a detailed overview of the segregation of the R6 resistance trait in five F2 populations. Chi-square tests confirmed that the observed segregation in the F2 populations was consistent with a 3:1 segregation of the R6 resistance profile, as assayed here with resistance to Pfs11.

(11) TABLE-US-00002 TABLE 2 R6 trait R6 trait cross present absent Total Chi square >0.05 ? 1 observed 8 3 11 0.862 yes expected (3:1) 8.25 2.75 11 2 observed 14 3 17 0.484 yes expected (3:1) 12.75 4.25 17 3 observed 11 3 14 0.758 yes expected (3:1) 10.5 3.5 14 4 observed 7 2 9 0.847 yes expected (3:1) 6.75 2.25 9 5 observed 20 5 25 0.564 yes expected (3:1) 18.75 6.25 25 Table 2: segregation of the R6 resistance profile in five F2 populations from a cross between a spinach plant of the invention (mother) to a father plant of a different genotype, which lacket the R6 resistance trait. Chi-square tests confirm that the observed numbers of F2 plants that were resistant and sensitive were in agreement with what is expected if the trait segregates in a dominant monogenic fashion, namely 3:1 (resistant:sensitive). In all cases chi-square values are well above 0.05.

(12) Similar segregation results were obtained when the progeny of a cross between a plant that carries the R6 resistance trait and a plant not carrying the said trait were assayed for the races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410, which together constitute the R6 resistance profile.

(13) The invention is further described by the following numbered paragraphs:

(14) 1. Spinach plant comprising a single dominant gene which confers resistance to Peronospora farinosa f. sp. spinaciae races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410, wherein the gene is obtainable by introgression from a plant grown from seeds of which a representative sample was deposited with the NCIMB under NCIMB accession number 41857.

(15) 2. Progeny of a spinach plant of paragraph 1, which progeny is resistant to Peronospora farinosa f. sp. spinaciae races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410, wherein the resistance is the result of the single dominant gene as defined in paragraph 1, which is as found in seeds of which a representative sample was deposited with the NCIMB under NCIMB accession number 41857.

(16) 3. Propagation material of a plant of paragraph 1 or 2, wherein a plant grown or regenerated from the material is resistant to Peronospora farinosa f. sp. spinaciae races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410.

(17) 4. Cell of a spinach plant of any one of paragraphs 1-3, which cell comprises a single dominant gene which leads to resistance to Peronospora farinosa f. sp. spinaciae races Pfs1, Pfs2, Pfs3, Pfs4, Pfs5, Pfs6, Pfs9, Pfs11, Pfs12, Pfs13 and UA4410, wherein the said gene is as present in a spinach plant, representative seeds of which were deposited under NCIMB accession number 41857.

(18) 5. Seed of a spinach plant of paragraph 1 or 2.

(19) 6. Harvested leaves of a spinach plant of any one of paragraphs 1-3.

(20) 7. Food product comprising the harvested leaves of paragraph 6.

(21) 8. Container comprising one or more spinach plants of any one of paragraphs 1-3 in a growth substrate for harvest of leaves from the spinach plant in a domestic environment.

(22) 9. Use of spinach plant of any of paragraphs 1-3, representative seeds of which were deposited under NCIMB accession number 41857, in breeding to confer resistance against Peronospora farinosa f. sp. spinaciae.

(23) Having thus described in detail preferred embodiments of the present invention, it is to be understood that the invention defined by the above paragraphs is not to be limited to particular details set forth in the above description as many apparent variations thereof are possible without departing from the spirit or scope of the present invention.