CA-3272860-A1 - DIDYMELLA BRYONIAE INTERNAL FRUIT ROT RESISTANCE IN CUCUMIS SATIVUS PLANTS

Abstract

The present invention relates to cultivated cucumber plants comprising modifications in their genome, whereby the modifications confer reduced susceptibility to internal fruit rot symptoms caused by Didymella bryoniae, and to methods for generating such plants, and their use.

Inventors

• Daniele LIBERTI
• Hans Peter KOELEWIJN
• Frank BEENDERS
• Laura REINIERS
• Wim VRIEZEN
• Lieke MERTENS

Assignees

• NUNHEMS B.V.

Dates

Publication Date

20260302

Application Date

20210315

Priority Date

20200318

Claims (1)

1. <pat:ClaimStatement>CLAIMS A plant cell of a Cucumis sativus var. sativus plant comprising an increased resistance against internal fruit rot symptoms caused by Didymella bryoniae, wherein said plant cell comprises an introgression fragment from a wild cucumber on chromosome 5, comprising Quantitative Trait Locus QTL5.1, wherein the introgression fragment comprising QTL5.1 comprises a SNP haplotype of at least 3 markers selected from: _ a Guanine for SNP 12 at nucleotide 51 of SEQ ID NO: 12 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 12, - an Adenine for SNP 45 at nucleotide 51 of SEQ ID NO: 45 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 45, - an Adenine for SNP 13 at nucleotide 51 of SEQ ID NO: 13 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 13, _ a Guanine for SNP_14 at nucleotide 51 of SEQ ID NO: 14 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 14, - a Thymine for SNP_46 at nucleotide 51 of SEQ ID NO: 46 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 46, - a Thymine for SNP 47 at nucleotide 51 of SEQ ID NO: 47 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 47, - a Cytosine for SNP 48 at nucleotide 51 of SEQ ID NO: 48 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 48, - a Guanine for SNP 49 at nucleotide 51 of SEQ ID NO: 49 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 49, - a Thymine for SNP_50 at nucleotide 51 of SEQ ID NO: 50 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 50, _ an Adenine for SNP 51 at nucleotide 51 of SEQ ID NO: 51 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 51, - a Guanine for SNP 15 at nucleotide 51 of SEQ ID NO: 15 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 15.</pat:ClaimStatement> <pat:Claims com:id="claims"> <pat:Claim com:id="CLM-00002"> <pat:ClaimNumber>2</pat:ClaimNumber> <pat:ClaimText>2. The plant cell according to claim 1, wherein said QTL5.1 is located in-between SNP 12 at nucleotide 51 in SEQ ID NO: 12 and SNP_15 at nucleotide 51 in SEQ ID NO: 15. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00003"> <pat:ClaimNumber>3</pat:ClaimNumber> <pat:ClaimText>3. The plant cell according to claim 1, wherein said QTL5.1 is located on chromosome 5 in- between SNP_12 at nucleotide 3823864 of chromosome 5 and SNP_15 at nucleotide 3967955. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00004"> <pat:ClaimNumber>4</pat:ClaimNumber> <pat:ClaimText>4. The plant cell according to claim 1, wherein said QTL5.1 is located physically in the region starting at 3.81 Mb and ending at 3.97 Mb of chromosome 5. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00005"> <pat:ClaimNumber>5</pat:ClaimNumber> <pat:ClaimText>5. The plant cell according to any one of claims 1 to 4, wherein said cell comprises said QTL5.1 in heterozygous form or in homozygous form in the genome. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00006"> <pat:ClaimNumber>6</pat:ClaimNumber> <pat:ClaimText>6. The plant cell according to any one of claims 1 to 5, wherein said cell comprises said QTL5.1 from seeds deposited under accession number NCIMB43530. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00007"> <pat:ClaimNumber>7</pat:ClaimNumber> <pat:ClaimText>7. The plant cell according to any one of claims 1 to 6, wherein the plant cell is from a plant which is of one of the following cucumber types: slicing cucumber, long cucumber, and European greenhouse cucumber. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00008"> <pat:ClaimNumber>8</pat:ClaimNumber> <pat:ClaimText>8. The plant cell according to any one of claims 1 to 7, wherein the plant cell is from a plant which is a single cross F1 hybrid or an inbred line. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00009"> <pat:ClaimNumber>9</pat:ClaimNumber> <pat:ClaimText>9. The plant cell according to any one of claims 1 to 8, wherein the plant cell is from a plant which produces seedless fruits without pollination. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00010"> <pat:ClaimNumber>10</pat:ClaimNumber> <pat:ClaimText>10. The plant cell according to any one of claims 1 to 9, wherein the plant cell is from a plant which is a vegetatively propagated plant. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00011"> <pat:ClaimNumber>11</pat:ClaimNumber> <pat:ClaimText>11. The plant cell according to any one of claims 1 to 10, wherein said introgression fragment comprises said SNP haplotype of at least 9 of said SNP markers. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00012"> <pat:ClaimNumber>12</pat:ClaimNumber> <pat:ClaimText>12. The plant cell according to any one of claims 1 to 11, wherein said introgression fragment comprises said SNP haplotype of at least 10 of said SNP markers. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00013"> <pat:ClaimNumber>13</pat:ClaimNumber> <pat:ClaimText>13. A method for identifying or detecting a cultivated C. sativus var. sativus plant or plant part comprising an introgression fragment on chromosome 5 comprising: a) providing a cultivated C. sativus var. sativus plant or plant parts or DNA of such plant or plant part, b) screening said plant, plant part or DNA using a molecular marker assay which detects at least one SNP marker selected from the group consisting of SNP_12 to SNP_15 for detecting the introgression fragment on chromosome 5; and c) identifying and/or selecting a plant comprising: i) at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more of the SNP markers of SNP 12 to SNP 15, for detecting the introgression fragment on chromosome 5; or ii) at least 2, 3, 4 5, 6, 7, 8, 9, 10 or more consecutive markers selected from SNP_12 to SNP_15, for detecting the introgression fragment on chromosome 5, wherein said SNP markers are selected from: _ a Guanine for SNP 12 at nucleotide 51 of SEQ ID NO: 12 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 12, - an Adenine for SNP 45 at nucleotide 51 of SEQ ID NO: 45 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 45, - an Adenine for SNP 13 at nucleotide 51 of SEQ ID NO: 13 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 13, - a Guanine for SNP 14 at nucleotide 51 of SEQ ID NO: 14 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 14, - a Thymine for SNP 46 at nucleotide 51 of SEQ ID NO: 46 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 46, - a Thymine for SNP 47 at nucleotide 51 of SEQ ID NO: 47 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 47, - a Cytosine for SNP 48 at nucleotide 51 of SEQ ID NO: 48 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 48, - a Guanine for SNP 49 at nucleotide 51 of SEQ ID NO: 49 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 49, - a Thymine for SNP 50 at nucleotide 51 of SEQ ID NO: 50 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 50, - an Adenine for SNP_51 at nucleotide 51 of SEQ ID NO: 51 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 51, _ a Guanine for SNP 15 at nucleotide 51 of SEQ ID NO: 15 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 15. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00014"> <pat:ClaimNumber>14</pat:ClaimNumber> <pat:ClaimText>14. A method of producing C. sativus F1 hybrid plants comprising an introgression fragment conferring enhanced internal fruit rot resistance is provided comprising: a) providing a first inbred cucumber plant comprising a recombinant chromosome 5 in homozygous form having an introgression fragment comprising an introgression fragment from a wild cucumber on chromosome 5, said introgression fragment comprises Quantitative Trait Locus QTL5.1 and a SNP haplotype of at least 3 markers selected from: - a Guanine for SNP 12 at nucleotide 51 of SEQ ID NO: 12 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 12, - an Adenine for SNP 45 at nucleotide 51 of SEQ ID NO: 45 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 45, - an Adenine for SNP 13 at nucleotide 51 of SEQ ID NO: 13 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 13, - a Guanine for SNP 14 at nucleotide 51 of SEQ ID NO: 14 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 14, - a Thymine for SNP 46 at nucleotide 51 of SEQ ID NO: 46 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 46, - a Thymine for SNP 47 at nucleotide 51 of SEQ ID NO: 47 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 47, - a Cytosine for SNP 48 at nucleotide 51 of SEQ ID NO: 48 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 48, - a Guanine for SNP 49 at nucleotide 51 of SEQ ID NO: 49 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 49, - a Thymine for SNP 50 at nucleotide 51 of SEQ ID NO: 50 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 50, - an Adenine for SNP 51 at nucleotide 51 of SEQ ID NO: 51 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 51, - a Guanine for SNP 15 at nucleotide 51 of SEQ ID NO: 15 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 15, b) providing a second inbred cucumber plant, c) crossing said cucumber plant of a) with said cucumber plant of b), d) collecting F1 hybrid seeds from said cross. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00015"> <pat:ClaimNumber>15</pat:ClaimNumber> <pat:ClaimText>15. A method for transferring the recombinant chromosome 5 comprising the QTL5.1 from one cultivated cucumber plant into another cultivated cucumber plant is provided, said method comprises the steps of: a) providing a first inbred cucumber plant comprising a recombinant chromosome 5 having an introgression fragment from a wild cucumber in homozygous form, said introgression fragment comprises Quantitative Trait Locus QTL5.1 and a SNP haplotype of at least 3 markers selected from: - a Guanine for SNP 12 at nucleotide 51 of SEQ ID NO: 12 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 12, - an Adenine for SNP 45 at nucleotide 51 of SEQ ID NO: 45 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 45, - an Adenine for SNP 13 at nucleotide 51 of SEQ ID NO: 13 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 13, - a Guanine for SNP 14 at nucleotide 51 of SEQ ID NO: 14 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 14, - a Thymine for SNP 46 at nucleotide 51 of SEQ ID NO: 46 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 46, - a Thymine for SNP 47 at nucleotide 51 of SEQ ID NO: 47 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 47, - a Cytosine for SNP 48 at nucleotide 51 of SEQ ID NO: 48 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 48, - a Guanine for SNP 49 at nucleotide 51 of SEQ ID NO: 49 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 49, - a Thymine for SNP 50 at nucleotide 51 of SEQ ID NO: 50 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 50, - an Adenine for SNP 51 at nucleotide 51 of SEQ ID NO: 51 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 51, - a Guanine for SNP 15 at nucleotide 51 of SEQ ID NO: 15 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 15, b) providing a second cultivated cucumber plant, c) crossing said cucumber plant of a) with said cucumber plant of b), d) collecting F1 hybrid seeds from said cross, and e) optionally selfing the plant grown from said F1 hybrid seeds to produce F2 seeds or further selfing generations, and optionally selecting the F2 seeds or further selfing generation seeds having the recombinant chromosome 5, and f) optionally breeding further with plants grown from said F1 or F2 or further generation selfing seeds to produce a cucumber plant having good agronomic characteristics and comprising the introgression fragment in homozygous or heterozygous form. </pat:ClaimText> </pat:Claim> <pat:Claim com:id="CLM-00016"> <pat:ClaimNumber>16</pat:ClaimNumber> <pat:ClaimText>16. A method for identifying a wild cucumber comprising a fruit rot resistance QTL on chromosome 5, said method comprising: A) providing a wild cucumber accession or several accessions; B) screening said accessions using a molecular marker assay which detects at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more SNP marker selected from SNP 12, SNP 45, SNP 13, SNP 14, SNP 46 to SNP 51 and SNP 15; C) identifying and/or selecting an accession from B) comprising a marker genotype or haplotype for at least 1, 2, 3, or more of the following markers selected from: - a Guanine for SNP 12 at nucleotide 51 of SEQ ID NO: 12 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 12, - an Adenine for SNP 45 at nucleotide 51 of SEQ ID NO: 45 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 45, - an Adenine for SNP 13 at nucleotide 51 of SEQ ID NO: 13 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 13, - a Guanine for SNP 14 at nucleotide 51 of SEQ ID NO: 14 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 14, - a Thymine for SNP 46 at nucleotide 51 of SEQ ID NO: 46 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 46, - a Thymine for SNP 47 at nucleotide 51 of SEQ ID NO: 47 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 47, - a Cytosine for SNP 48 at nucleotide 51 of SEQ ID NO: 48 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 48, - a Guanine for SNP 49 at nucleotide 51 of SEQ ID NO: 49 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 49, _ a Thymine for SNP 50 at nucleotide 51 of SEQ ID NO: 50 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 50, - an Adenine for SNP 51 at nucleotide 51 of SEQ ID NO: 51 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 51, a Guanine for SNP 15 at nucleotide 51 of SEQ ID NO: 15 or at nucleotide 51 of a sequence comprising at least 97% identity to SEQ ID NO: 15; and optionally D) introgressing said QTL from said wild accession into cultivated cucumber. </pat:ClaimText> </pat:Claim> </pat:Claims>