Brassicaceae plants resistant to
The present invention relates to the identification of clubroot resistance genes from Brassica . Clubroot resistant Brassicaceae plants are provided, as well as clubroot resistance genes and methods and means to increase clubroot resistance in Brassicaceae.
1. A method for increasing clubroot resistance in a Brassica plant comprising: introducing a CRL clubroot resistance locus in said Brassica plant, and selecting said CRL clubroot resistant Brassica plant for the presence of the CRL clubroot resistance locus by analyzing genomic DNA from said plant for the presence of at least one molecular marker, wherein said at least one molecular marker is linked to the CRL clubroot resistance locus, wherein said CRL clubroot resistance locus comprises the CRL1 and CRL2 clubroot resistance genes, wherein
a) said CRL1 clubroot resistance gene comprises a nucleotide sequence
i) having at least 95% sequence identity to the sequence of nucleotide positions 32750 to 51049 of SEQ ID NO: 1;
ii) having a coding sequence having at least 95% sequence identity to the sequence of nucleotide positions 52 to 5343 of SEQ ID NO: 2, nucleotide positions 52 to 5340 of SEQ ID NO: 4, nucleotide positions 52 to 5361 of SEQ ID NO: 6, or to SEQ ID NO: 10; or
ii) encoding a protein having at least 95% sequence identity to SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, or SEQ ID NO: 11; and
b) said CRL2 clubroot resistance gene comprises a nucleotide sequence having at least 95% sequence identity to the sequence of nucleotide positions 220 to 2898 of SEQ ID NO: 8, or encoding a protein having at least 95% sequence identity to SEQ ID NO: 9.
2. A method for producing a clubroot resistant Brassica plant comprising:
introducing a CRL clubroot resistance locus in said Brassica plant, and selecting said CRL clubroot resistant Brassica plant for the presence of the CRL clubroot resistance locus by analyzing genomic DNA from said plant for the presence of at least one molecular marker, wherein said at least one molecular marker is linked to the CRL clubroot resistance locus, wherein said CRL clubroot resistance locus comprises the CRL1 and CRL2 clubroot resistance genes, wherein
a) said CRL1 clubroot resistance gene comprises a nucleotide sequence
i) having at least 95% sequence identity to the sequence of nucleotide positions 32750 to 51049 of SEQ ID NO: 1;
ii) having a coding sequence having at least 95% sequence identity to the sequence of nucleotide positions 52 to 5343 of SEQ ID NO: 2, nucleotide positions 52 to 5340 of SEQ ID NO: 4, nucleotide positions 52 to 5361 of SEQ ID NO: 6, or to SEQ ID NO: 10; or
iii) encoding a protein having at least 95% sequence identity to SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, or SEQ ID NO: 11; and
b) said CRL2 clubroot resistance gene comprises a nucleotide sequence having at least 95% sequence identity to the sequence of nucleotide positions 220 to 2898 of SEQ ID NO: 8; or encoding a protein having at least 95% sequence identity to SEQ ID NO: 9; and
generating progeny from the clubroot resistant Brassica plant comprising the CRL clubroot resistance locus, wherein said progeny is clubroot resistant and comprises said CRL clubroot resistance locus.
3. The method according to claim 2 , wherein said CRL clubroot resistance locus comprises a sequence having at least 95% sequence identity to SEQ ID NO: 1.
4. A Brassica plant or plant cell comprising a CRL1 and a CRL2 clubroot resistance gene as transgene, wherein
a) said CRL1 clubroot resistance gene
(i) comprises a coding sequence having at least 95% sequence identity to the sequence of nucleotide positions 52 to 5343 of SEQ ID NO: 2, nucleotide positions 52 to 5340 of SEQ ID NO: 4, nucleotide positions 52 to 5361 of SEQ ID NO: 6, or to SEQ ID NO: 10; or (ii) encodes a protein having an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, or SEQ ID NO: 11; and
b) said CRL2 clubroot resistance gene comprises a coding sequence having at least 95% sequence identity to the sequence of nucleotide positions 220 to 2898 of SEQ ID NO: 8, or encodes a protein having an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 9.
5. The Brassica plant or plant cell of claim 4 , wherein said CRL1 clubroot resistance gene comprises a sequence having at least 95% sequence identity to the sequence of nucleotide positions 32750 to 51049 of SEQ ID NO: 1.
6. The Brassica plant or plant cell of claim 4 , comprising a transgene comprising a sequence having at least 95% sequence identity to SEQ ID NO: 1.
7. A method for increasing clubroot resistance in a Brassica plant, comprising:
a) providing a CRL1 and a CRL2 clubroot resistance gene as transgene, b) introducing the CRL1 and CLR2 gene into a Brassica plant cell, to create transgenic cells; and c) regenerating transgenic plants from said transgenic cells; wherein
said CRL1 clubroot resistance gene i) comprises a coding sequence having at least 95% sequence identity to the sequence of nucleotide positions 52 to 5343 of SEQ ID NO: 2, nucleotide positions 52 to 5340 of SEQ ID NO: 4, nucleotide positions 52 to 5361 of SEQ ID NO: 6, or to SEQ ID NO: 10; or ii) encodes a protein having an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, or SEQ ID NO: 11; and wherein
said CRL2 clubroot resistance gene comprises a nucleotide sequence having a coding sequence having at least 95% sequence identity to the sequence of nucleotide positions 220 to 2898 of SEQ ID NO: 8, or encodes a protein having at least 95% sequence identity to SEQ ID NO: 9.
8. A clubroot resistant Brassica plant or plant cell obtainable by the method according to claim 2 , said plant or plant cell further comprising at least one other disease resistance gene, said other disease resistance gene is a clubroot resistance gene, a blackleg resistance gene, a Sclerotinia resistance gene, a Verticillium resistance gene, a Fusarium resistance gene, an Aster Yellows resistance gene, an Altemaria resistance gene, or a Grey Stem resistance gene; wherein said other disease resistance gene is a transgene which is genetically linked with said CRL clubroot resistance gene or said clubroot resistance locus.
9. A chimeric gene comprising the following operably linked elements:
a) a heterologous plant-expressible promoter;
b) a DNA sequence coding for a CRL1 or for a CRL2 protein; and
c) optionally, a transcription termination and polyadenylation region functional in plant cells, wherein
a) said DNA sequence coding for a CRL1 protein comprises a nucleotide sequence i) having at least 95% sequence identity to the sequence of nucleotide positions 52 to 5343 of SEQ ID NO: 2, nucleotide positions of 52 to 5340 of SEQ ID NO: 4, nucleotide positions of 52 to 5361 of SEQ ID NO: 6, or to SEQ ID NO: 10; or ii) encoding a protein having an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, or SEQ ID NO: 11; and
b) said DNA sequence coding for a CRL2 protein comprises a nucleotide sequence having at least 95% sequence identity to the sequence of nucleotide positions 220 to 2898 of SEQ ID NO: 8; or encoding a protein having an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 9.
10. A method to produce clubroot free Brassica plants, comprising sowing seeds from the plants of claim 4 , and growing plants from said seeds.
11. A method of producing food, feed, or an industrial product comprising providing the Brassica plant or plant cell of claim 4 , and preparing food, feed or industrial product from the plant or plant cell.
12. The method of claim 11 wherein
a) the food or feed is oil, meal, grain, starch, flour or protein; or
b) the industrial product is biofuel, fiber, industrial chemicals, a pharmaceutical or a nutraceutical.
13. The method according to claim 1 , wherein said CRL1 clubroot resistance gene comprises a nucleotide sequence
said CRL1 clubroot resistance gene comprises a nucleotide sequence
i) having at least 98% sequence identity to the sequence of nucleotide positions 32750 to 51049 of SEQ ID NO: 1;
ii) having a coding sequence having at least 98% sequence identity to the sequence of nucleotide positions 52 to 5343 of SEQ ID NO: 2, nucleotide positions 52 to 5340 of SEQ ID NO: 4, nucleotide positions 52 to 5361 of SEQ ID NO: 6, or to SEQ ID NO: 10; and
iii) encoding a protein having at least 98% sequence identity to SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, or SEQ ID NO: 11; and
b) said CRL2 clubroot resistance gene comprises a nucleotide sequence
having a coding sequence having at least 98% sequence identity to the sequence of nucleotide positions 220 to 2898 of SEQ ID NO: 8, or encoding a protein having an amino acid sequence having at least 98% sequence identity to SEQ ID NO: 9.
14. The method according to claim 2 , wherein said CRL clubroot resistance locus comprises a sequence having at least 98% sequence identity to SEQ ID NO: 1.
15. The Brassica plant or plant cell of claim 4 , wherein said CRL1 clubroot resistance gene
i) comprises a coding sequence having at least 98% sequence identity to the sequence of nucleotide positions 52 to 5343 of SEQ ID NO: 2, nucleotide positions 52 to 5340 of SEQ ID NO: 4, nucleotide positions 52 to 5361 of SEQ ID NO: 6, or to SEQ ID NO: 10; or
(ii) encodes a protein having an amino acid sequence having at least 98% sequence identity to SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, or SEQ ID NO: 9; and
said CRL.2 clubroot resistance gene
comprises a coding sequence having at least 98% sequence identity to the sequence of nucleotide positions 220 to 2898 of SEQ ID NO: 8;
or encodes a protein having an amino acid sequence having at least 98% sequence identity to SEQ ID NO: 9.
16. The Brassica plant or plant cell of claim 4 , wherein said CRL1 clubroot resistance gene comprises a sequence having at least 98% sequence identity to the sequence of nucleotide positions 32750 to 51049 of SEQ ID NO: 1.
17. The Brassica plant or plant cell of claim 4 , comprising a transgene comprising a sequence having at least 98% sequence identity to SEQ ID NO: 1.
18. The chimeric gene of claim 9 , wherein
said DNA sequence coding for a CRL1 protein comprises a nucleotide sequence
i) having a coding sequence having at least 98% sequence identity to the sequence of nucleotide positions 52 to 5343 of SEQ ID NO: 2, nucleotide positions 52 to 5340 of SEQ ID NO: 4, nucleotide positions 52 to 5361 of SEQ ID NO: 6, or to SEQ ID NO: 10; or
iii) encoding a protein having at least 98% sequence identity to SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, or SEQ ID NO: 11; and
b) said DNA sequence coding for CRL2 protein comprises a nucleotide sequence having at least 98% sequence identity to the sequence of nucleotide positions 220 to 2898 of SEQ ID NO: 8, or encoding a protein having at least 98% sequence identity to SEQ ID NO: 9.