Increased yield in cucumber

Abstract

The present invention relates to a Cucumis sativus plant having an increased fruit yield. The invention is further related to markers linked to the increased fruit yield phenotype and to the use of such markers to identify or select plants having such phenotype. The invention also relates to the seeds and progeny of such plants and to the propagation material for obtaining such plants.

Description

INCREASED YIELD IN CUCUMBERFIELD OF THE INVENTION

[0001] The present invention relates to a Cucumis sativus plant having an increased fruit yield. The invention is further related to markers linked to the increased fruit yield phenotype and to the use of such markers to identify or select plants having such phenotype. The invention also relates to the seeds and progeny of such plants and to the propagation material for obtaining such plants.BACKGROUND OF THE INVENTION

[0002] Cucumber (Cucumis sativus L. 2n = 2x = 14) belongs to the Cucurbitaceae family that includes more than 800 species and is an annual vine plant. Cucumber is one of the most important and widely cultivated vegetables crops, it originates from India, and China is extensively considered as its secondary center of origin. Despite its considerable morphological variability, cucumber showed a narrow genetic base and has a relatively small genome size (350 Mbp), which was sequenced in 2009 (Huang et al., 2009. Nat Genet. 2009 Dec; 41 (12): 1275-81 ). Cucumis sativus houses several botanical varieties including var. sativus, the cultivated cucumber and the wild, free-living var. hardwickii (R.) Alef. (Kirkbride, 1993. Biosystematic monograph of the genus Cucumis (Cucurbitaceae). 84).The cultivated cucumber genome has seven pairs of chromosomes (n=7) and a haploid genome size of about 367 Mb (Megabases) with an estimated total of about 26,682 genes. The cucumber genome was the first vegetable genome to be sequenced (Huang et al. 2009, Nature Genetics, Volume 41 , Number 12, p 1275-1283).

[0003] Yield of cultivated cucumber has not increased much over the last decades. Shetty and Wehner 2002 (CropSci. 42: 2174-2183) screened the USDA cucumber germplasm collection for fruit quality and fruit yield under field conditions in North Carolina (USA) and suggested that high yielding cultigens identified in their study can be used to develop high yielding cultivars.

[0004] Yuan et al. 2008 (Euphytica 164: 473-491) genetically mapped specific fruit traits in a cross between a Northern Chinese Cucumber S94 and a Northwest European Cucumber S06. Their linkage group 3 appears to correspond to the physical chromosome 2 and their linkage group 2 appears to correspond to the physical chromosome 6. They mapped a locus called 1w2.1 (fruit weight) to the top of chromosome 6 (LG2) and they mapped a locus called fw3.1 (fruit weight) to the bottom of chromosome 2 (LG3). However, they did not map total fruit yield.

[0005] Fazio et al. 2003 (Theor Appl Genet 107: 864-874) genetically mapped a number of traits, including cumulative fruits per plants over three harvests and morphological traits such as little leaf. Their linkage group 1 appears to correspond to the physical chromosome 6. A locus called fp1 1 .2 (fruits per plant) was consistent in both environments and mapped to the little leaf locus.

[0006] Still, there remains a need for identifying solutions for fruit yield in cucumber to be able to increase fruit yield of modern cucumber varieties.SUMMARY

[0007] The present invention relates to a Cucumis sativus plant comprising a mutant allele of a YieldPlus gene on chromosome 3 in its genome, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele, resulting in an increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

[0008] The following embodiments, can be optionally implemented, separately or in combination one with the others:

[0009] Embodiment 1 : A Cucumis sativus plant comprising a mutant allele of a YieldPlus gene on chromosome 3 in its genome, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

[0010] Embodiment 2: A plant according to embodiment 1 , wherein said mutant allele is present heterozygously in the genome of said plant.

[0011] Embodiment 3: A plant according to embodiment 1 , wherein said mutant allele is present homozygously in the genome of said plant.

[0012] Embodiment 4: A plant according to any one of embodiments 1 to 3 wherein said mutant allele comprises a nucleotide sequence having at least 90% identity with the sequence set forth in SEQ ID NO: 1 or SEQ ID NO: 2.

[0013] Embodiment 5: A plant according to any one of embodiments 1 to 4, wherein said mutation is a loss-of-function mutation and / or a mutation which decreases expression of the YieldPlus protein.

[0014] Embodiment 6: A plant according to any one of embodiments 1 to 5 wherein said at least one loss-of-function mutation is selected from a missense mutation, a nonsense mutation and a frameshift mutation.

[0015] Embodiment 7: The plant according to any one of embodiments 1 to 6, wherein said at least one mutation is located in the coding sequence of the YieldPlus gene.

[0016] Embodiment 8: The plant according to embodiment 7, wherein said at least one mutation results in at least one amino acid change, addition or deletion in the amino acid sequence set forth in SEQ ID NO:3.

[0017] Embodiment 9: The plant according to embodiment 8, wherein said at least one mutation results in at least one amino acid change, addition or deletion in the C-terminal portion of the protein encoded by the YieldPlus gene, preferably between the amino acid residues in position 150 and 277 of SEQ ID NO:3.

[0018] Embodiment 10: The plant according to embodiment 9, wherein said at least one mutation results in at least one amino acid change, addition or deletion in position 253 of SEQ ID NO:3.

[0019] Embodiment 11 : The plant according to embodiment 10, wherein said at least one mutation results in a G253S change in the amino acid sequence set forth in SEQ ID NO:3.

[0020] Embodiment 12: The plant according to embodiment 11 , wherein said at least one mutation is a G757A mutation in SEQ ID NO: 2.

[0021] Embodiment 13: The plant according to any one of embodiment 1 to 6, wherein said mutation is located in a regulatory sequence selected from a 3’-untranslated region (3 -UTR), a 5’-untranslated region (5 -UTR), a promoter, an enhancer and a polyA signal sequence.

[0022] Embodiment 14: The plant according to any one of embodiments 1 to 13, which produces at least 10 fruits or more, preferably at least 12 fruits or more.

[0023] Embodiment 15: The plant according to any one of embodiments 1 to 14, which produces at least 10%, preferably at least 20%, still preferably at least 50% more fruits in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene.

[0024] Embodiment 16: The plant according to any one of embodiments 1 to 15, wherein the fruit weight of said plant is at least 10%, preferably at least 20%, still preferably at least 50% higher in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene.

[0025] Embodiment 17: The plant according to any one of embodiments 1 to 16, wherein said plant is a plant from an inbred line or is a hybrid plant.

[0026] Embodiment 18 The plant according to any one of embodiments 1 to 17, which further comprises one or more traits of agronomical interest selected from resistance to ZYMV (Zucchini Yellow Mosaic Virus), resistance to CVYV (Cucumber Vein Yellowing Virus), resistance to PRSV (Papaya Ringspot Virus), resistance to WMV (Watermelon Mosaic Virus), resistance to CMV (Cucumber Mosaic Virus), resistance to powdery mildew, resistance to potyviruses, resistance to downy mildew, e.g. caused by Pseudoperonospora cubensis, resistance to Fusaria, e.g. caused by Fusariumoxysporum f.sp. cucumerinum or by Fusarium oxysporum f.sp. radicis cucumerinum, resistance to scab, e.g. caused by Cladosporium cucumerinum, resistance to CYSDV, resistance to angular leaf sport and resistance to anthracnose.

[0027] Embodiment 19: A seed for producing a plant of any one of embodiments 1 to 18.

[0028] Embodiment 20: A cell of a plant according to any one of embodiments 1 to 18, preferably a cell derived from an embryo, protoplast, meristematic cell, callus, pollen, leaf, anther, stem, petiole, root, root tip, fruit, seed, flower, cotyledon, and / or hypocotyl, comprising in its genome a mutant allele of the YieldPlus gene, wherein said mutant allele comprises one or more mutations in the sequence of the gene, or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 .

[0029] Embodiment 21 : A plant part of a C. sativus plant comprising at least one cell according to embodiment 20, preferably an embryo, protoplast, meristematic cell, callus, pollen, leaf, anther, stem, petiole, root, root tip, fruit, seed, flower, cotyledon, and / or hypocotyl, in particular a fruit.

[0030] Embodiment 22: An isolated polynucleotide, comprising a mutant allele of the YieldPlus gene, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , wherein said mutant allele confers increased fruit yield to a plant comprising said mutant allele, in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene.

[0031] Embodiment 23: The isolated polynucleotide of embodiment 22, which comprises a nucleotide sequence having at least 80% identity with SEQ ID NO: 1 or SEQ ID NO: 2.

[0032] Embodiment 24: The isolated polynucleotide of embodiment 22, which encodes a polypeptide with at least 95% identity with SEQ ID NO:3.

[0033] Embodiment 25: An isolated polypeptide, comprising an amino acid sequence with at least 95% identity with SEQ ID NO:3, wherein said polypeptide comprises at least one mutation in the amino acid sequence, in comparison to the sequence of the corresponding wild-type polypeptide as set forth in SEQ ID NO:3, wherein said mutation confers a I oss-of-fu notion phenotype to said polypeptide.

[0034] Embodiment 26: An in vitro cell or tissue culture of regenerable cells of the C. sativus plant according to any one of embodiments 1 to 18, wherein the regenerable cells are derived from an embryo, protoplast, meristematic cells, callus, pollen, leaf, anther, stem, petiole, root, root tip, seed, flower, cotyledon, and / or hypocotyl.

[0035] Embodiment 27: A method of producing a C. sativus plant, comprising: a) obtaining a part of a plant according to embodiments 1 to 18, b) vegetatively propagating said plant part to generate a plant from said plant part.

[0036] Embodiment 28: A method of producing a C. sativus plant, comprising the introduction of at least one mutation in the YieldPlus gene on chromosome 3 in the genome of a C. sativus plant, wherein said mutation is introduced in the sequence of the gene or in a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

[0037] Embodiment 29: The method of embodiment 28, wherein said mutation is introduced by mutagenesis or genome editing, in particular by a technique selected from ethyl methanesulfonate (EMS) mutagenesis, oligonucleotide directed mutagenesis (ODM), Zinc finger nuclease (ZFN) technology, Transcription Activator-Like Effector Nucleases (TALENs) the CRISPR / Cas system, the CRISPR / Cpf system engineered meganuclease, re-engineered homing endonucleases and DNA guided genome editing.

[0038] Embodiment 30: The method of embodiment 29, comprising:a) Introducing one or more mutations in cucumber plant(s), seed(s) or plant part(s), b) Optionally, determining if the plant, seed or plant part under a) presents an increased yield compared to a plant not having said at least one mutation; and c) Selecting a plant that comprises a mutant allele of a YieldPlus gene, wherein said mutant allele comprises at least one mutation in the sequence of the gene or in a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

[0039] Embodiment 31 : The method of embodiment 29 or 30, wherein the produced plant is according to anyone of embodiments 1 to 18.

[0040] Embodiment 32: A method of producing a C. sativus plant, comprising:(a) crossing a C. sativus plant according to any one of embodiments 1 to 18 with itself or with a second C. sativus plant, preferably of a different genotype, to produce one or more progeny plants;(b) selecting a progeny plant comprising a mutant allele of the YieldPlus gene; and(c) optionally self-pollinating and / or backcrossing one or several times the plant selected at step b) and selecting in the progeny thus obtained a plant comprising a mutant allele of the YieldPlus gene.

[0041] Embodiment 33: A method for detecting and / or selecting a cucumber plant, seed or plant part, comprising the steps of:(a) Providing at least one genomic DNA sample of a cucumber plant, seed, or plant part;(b) Identifying a mutant allele of the YieldPlus gene on chromosome 3, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 ; and(c) Selecting a plant comprising said mutant allele, wherein the selected plant has increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

[0042] Embodiment 34: The method of embodiment 33, wherein step (b) comprises carrying out a genotyping assay, using the DNA samples of a) as template, that discriminates between the wild type YieldPlus allele and the mutant allele, wherein said genotyping assay is based on nucleic acid amplification making use of YieldPlus allele specific oligonucleotide primers, and / or wherein said genotyping assay is based on nucleic acid hybridization making use of YieldPlus allele-specific oligonucleotide probes.

[0043] Embodiment 35: The method of embodiment 34, wherein said YieldPlus allele specific oligonucleotide primers or said YieldPlus allele-specific oligonucleotide probes comprise at least 10 nucleotides of SEQ ID NO: 1 or SEQ ID NO: 2 or of the complement strand of SEQ ID NO: 1 or SEQ ID NO: 2.

[0044] Embodiment 36: The method of embodiment 35, wherein said YieldPlus allele specific oligonucleotide primers comprise a sequence selected from the sequences set forth in SEQ ID NO: 6 and SEQ ID NO: 7.

[0045] Embodiment 37: A method for screening and / or selecting plant, seeds or plant part, or DNA or RNA or protein derived therefrom, for the presence of a mutant allele of the YieldPlus gene on chromosome 3, conferring increased fruit yield in comparison to an isogenic plant comprising a wildtype allele of the YieldPlus gene, comprising one or more of the following steps: a) determining if the level of expression of the YieldPlus gene in the plant, seed or plant part is reduced or abolished in comparison to a plant comprising a wild-type allele of the YieldPlus gene, wherein the sequence of the wild-type YieldPlus gene is set forth in SEQ ID NO: 1 ; b) determining if the amounts of the protein encoded by the YieldPlus gene in the plant, seed or plant part thereof is reduced or abolished in comparison to a plant comprising a wild-type allele of the YieldPlus gene; c) determining the presence of at least one mutation in the YieldPlus gene or a regulatory sequence thereof, in nucleic acid sample, preferably a mRNA, cDNA and / or genomic DNA sample, from the plant, seed or plant part in comparison to the wild-type YieldPlus gene having the sequence set forth in SEQ ID NO: 1 ; d) determining the presence of at least one mutation in the YieldPlus protein from the plant, seed or plant part, in comparison to the wild-type YieldPlus protein of SEQ ID NO :3.

[0046] Embodiment 38: A method for improving the yield of cucumber production, wherein said method comprises growing C. sativus plants according to any one of embodiments 1 to 18 and harvesting fruits set by said plants.

[0047] Embodiment 39: A method of producing cucumber fruit comprising: a) growing a C. sativus plant according to any one of embodiments 1-18. b) allowing said plant to set fruit; and c) harvesting fruit of said plant.

[0048] Embodiment 40: A method of producing a foodstuff or feedstuff comprising: a) Obtaining fruits of a C. sativus plant according to any one of embodiments 1-18. b) Processing said fruits into a processed food or feedstuff or using said fruit as an ingredient into a foodstuff or feedstuff.

[0049] Embodiment 41 : Use of a C. sativus plant according to any one of embodiments 1-18 or some fruit thereof in the fresh cut market or for food processing.

[0050] Embodiment 42: A plant according to any one of embodiments 1-18, wherein the average individual fruit weight of fruits of said plant is decreased by less than 50%, in particular less than 40%, more particularly less than 30%, still more particularly less than 20%, even more particularly less than 10%, most particularly less than 5% in comparison to the average individual fruit weight of fruits of an isogenic plant which comprises a wild-type allele of the YieldPlus gene, or is not decreased.BRIEF DESCRIPTION OF THE DRAWINGS

[0051] Figure 1 shows the mean fruit weight per plant (A) and mean fruit number per plant(B) of progeny plants of the mutant plant which are homozygous for the mutant allele (Homo Mut) and progeny plants which are homozygous for the wild-type allele (Homo WT). Trial conducted for 32 days, 15 picks, 255 mutant fruits, 175 wild-type fruits. Fruit weight: Anova R2=0.32, P<0.001 . Fruit number: Anova R2=0.24, P<0.005.SEQUENCE DESCRIPTION

[0052] SEQ ID NO: 1 : wild-type allele of the YieldPlus gene

[0053] SEQ ID NO: 2: cds of wild-type YieldPlus gene

[0054] SEQ ID NO: 3: wild-type YieldPlus protein

[0055] SEQ ID NO: 4: mutant allele of the YieldPlus gene

[0056] SEQ ID NO: 5: mutant YieldPlus protein

[0057] SEQ ID NO: 6: KASP detection of the mutant: primer recognizing the wild-type allele

[0058] SEQ ID NO: 7: KASP detection of the mutant: primer recognizing the mutant allele

[0059] SEQ ID NO: 8: KASP detection of the mutant: common primerDEFINITIONS

[0060] In order that the present disclosure be more readily understood, certain terms are first defined. Additional definitions are set forth throughout the detailed description.

[0061] As used herein, the term "cucumber" refers to a plant, or any part thereof, of the species Cucumis sativus. This includes, without being limited to, plants commonly referred to as Cucumber American gherkin, Cassabanana, Cuke, Gherkin, Hothouse cucumber, Lemon cucumber, Mandera cucumber, Pickling cucumber, Serpent cucumber, Slicing cucumber, Snake cucumber, and West Indian gherkin.

[0062] As used herein, the terms “offspring” or “progeny” refers to any plant resulting as progeny from a vegetative or sexual reproduction from one or more parent plants or descendants thereof. For instance, an offspring plant may be obtained by cloning or selfing of a parent plant or by crossing two parental plants and include selfing as well as the F1 or F2 or still further generations. An F1 is a first-generation offspring produced from parents at least one of which is used for the first time as donor of a trait, while offspring of second generation (F2) or subsequent generations (F3, F4, etc.) are specimens produced from selfing of F1 ’s, F2's etc. An F1 may thus be (and usually is) a hybrid resulting from a cross between two true breeding parents (true-breeding is homozygous for a trait), while an F2 may be (and usually is) an offspring resulting from self-pollination of said F1 hybrids.

[0063] As used herein, the terms “cross”, “crossing”, “cross pollination” or “cross-breeding” refer to the process by which the pollen of one flower on one plant is applied (artificially or naturally) to the ovule (stigma) of a flower on another plant.

[0064] As used herein, the terms “molecular marker” or “marker” refer to an indicator that is used in methods for visualizing differences in characteristics of nucleic acid sequences. Examples of such indicators are restriction fragment length polymorphism (RFLP) markers, amplification fragment length polymorphism (AFLP) markers, single nucleotide polymorphisms (SNPs), insertion mutations, microsatellite markers (SSRs), sequence-characterized amplified regions (SCARs), cleaved amplified polymorphic sequence (CAPS) markers or isozyme markers or combinations of the markers described herein which defines a specific genetic and chromosomal location.

[0065] It is noted in this respect that specific positions in a chromosome can indeed be defined with respect to markers, such as SNPs, insofar as the flanking sequences of said markers are defined in order to unambiguously position them on the genome. The present inventors have used SNPs markers, identified by their flanking sequences, present in the cucumber genome, to discriminate between the mutant and wild type allele and to track down the mutation conferring the increased Yield in the cucumber genome. Their location in the cucumber genome refer to the assembly Cucumber (Chinese Long) v3 available at http: / / cucurbitgenomics.org / v2 / organism / 19 (based on Li et al, 2019 A chromosome-scale genome assembly of cucumber (Cucumis sativus L.). Gigascience, Volume 8, Issue 6, June 2019, giz072).

[0066] As used herein, the term “primer” refers to an oligonucleotide which is capable of annealing to the amplification target allowing a DNA polymerase to attach, thereby serving as a point of initiation of DNA synthesis when placed under conditions in which synthesis of primers extension product is induced, i.e., in the presence of nucleotides and an agent for polymerization such as DNA polymerase and at a suitable temperature and pH. The primer is preferably single stranded for maximum efficiency in amplification. Preferably, the primer is an oligodeoxyribonucleotide. The primer must be sufficiently long to prime the synthesis of extension products in the presence of the agent for polymerization. The exact length of the primers will depend on many factors, including temperature and composition (A / T and G / C content) of primer. A pair of bi-directional primers consists of one forward and one reverse primer as commonly used in the art of DNA amplification such as in PCR amplification.

[0067] As used herein, a single nucleotide polymorphism (SNP) is a DNA sequence variation occurring when a single nucleotide — A, T, C, or G — in the genome (or other shared sequence) differs between members of a biological species or paired chromosomes in an individual. For example, two sequenced DNA fragments from different individuals, AAGCCTA to AAGCTTA, contain a difference in a single nucleotide. In this case there are two alleles: C and T.

[0068] As used herein, the term “genotype” refers to the genetic makeup of an individual cell, cell culture, tissue, organism (e.g., a plant), or group of organisms.

[0069] As used herein, the term “heterozygous” refers to the presence of different alleles (forms of a given gene, genetic determinant or sequences) at a particular locus.

[0070] As used herein, the term “homozygous” refers to the presence of identical alleles at one or more loci in homologous chromosomal segments.

[0071] As used herein, “homologous chromosomes”, or “homologs” (or homologues), refer to a set of one maternal and one paternal chromosome that pair up with each other during meiosis. These copies have the same genes in the same loci and the same centromere location.

[0072] As used herein, the term “hybrid” refers to any individual cell, tissue or plant resulting from a cross between parents that differ in one or more genes. An F1 hybrid (HF1) results from the cross of two genetically different parent cultivars or lines.

[0073] As used herein, two plants are said “isogenic”, when they have the same or essentially the same set of chromosomes and genes, except for one gene, e.g. the YieldPlus gene. The two isogenic plants thus comprise different alleles of the gene. Comparing the phenotype of two isogenic plants allows the evaluation of the effect of an allelic variation of the gene.

[0074] As used herein, the term “locus” (plural: “loci”) refers to any site that has been defined genetically, this can be a single position (nucleotide) or a chromosomal region. A locus may be a gene, a genetic determinant, or part of a gene, or a DNA sequence, and may be occupied by different sequences (e.g. The YieldPlus locus is, thus, the location in the genome of cucumber, where the mutant allele and / or the wild type allele of the YieldPlus gene is found). A locus may also be defined by a marker, such as a SNP (Single Nucleotide Polymorphism), by several markers (e.g. SNPs), or by two flanking markers (e.g. SNPs). As used herein, the term “allele(s)” means any of one or more alternative forms of a gene at a particular locus, e.g. the YieldPlus locus. The alleles of the gene may be wild type (e.g. SEQ ID NO: 1), or mutant alleles, which alleles relate to one trait or characteristic at a specific locus (e.g. increased yield). In a diploid cell of an organism, alleles of a given gene are located at a specific location, or locus (loci plural) on a chromosome. One allele is present on each chromosome of the pair of homologous chromosomes. A diploid plant species may comprise a large number of different alleles at a particular locus. A triploid plant is referred to as homozygous for the gene if it has three identical alleles of a gene (e.g. three copies of the mutant allele) and a tetrapioid plant is referred to as homozygous for the gene if it has four identical alleles of the gene, e.g. four copies of the mutant allele.

[0075] As used herein, the term “gene” means a (genomic) DNA sequence comprising a region (transcribed region), which is transcribed into a messenger RNA molecule (mRNA) in a cell, and operably linked regulatory regions (e.g. a promoter, enhancer, 3’UTR region, 5’-UTR region, polyA sequence or intron). An example is the YieldPlus gene of the invention. Different alleles of a gene are thus alternatives forms of the gene, which may be reflected in differences in one or more nucleotides of the genomic DNA sequence (e.g. in a regulatory sequence or in a coding sequence etc.), in the transcribed mRNA and / or amino acid sequence of the encoded protein.

[0076] As used herein, the term “allele” refers to any of several alternative or variant forms of a genetic unit, such as a gene, which are alternative in inheritance because they are positioned at the same locus in homologous chromosomes. Such alternative or variant forms may be the result of single nucleotide polymorphisms, insertions, inversions, translocations or deletions, or the consequence of generegulation caused, for example, by chemical or structural modification, transcription regulation or post- translational modification / regulation. In a diploid cell or organism such as cucumber, the two alleles of a given gene or genetic element typically occupy corresponding loci on a pair of homologous chromosomes.

[0077] As used herein, the term “wild-type allele” such as in “wild type YieldPlus allele” refers herein to the functional allele of the gene, which causes the plant to develop a normal phenotype, e.g. a normal fruit yield.

[0078] As used herein, the term “mutant allele” such as in “mutant YieldPlus allele” refers herein to an allele of a gene comprising one or more mutations in comparison to a wild-type allele, which causes the cucumber plant to exhibit a modified phenotype, e.g. an increased fruit yield. The mutation(s) in the mutant allele can be any mutation or combination of mutations, including deletions, substitutions, truncations, insertions, point mutations, non-sense mutations, mis-sense mutations, frame shift mutations and / or mutations in the coding sequence and / or in one or more regulatory sequences such as a promoter sequence, a 3’UTR sequence, a 5’-UTR sequence, an enhancer sequence, a silencer sequence, a polyA sequence and an intron sequence.

[0079] As used herein, the term "Induced mutant alleles” are mutant alleles in which the mutation(s) is / are / have been induced by human intervention, e.g. by mutagenesis via physical or chemical mutagenesis methods or via e.g. tissue culture (as described in e.g. Zhang et al, Pios 9(5) e96879), including also targeted gene editing techniques (such as CRISPR based techniques, TALENS, etc.).

[0080] As used herein, the term “Backcrossing” is a process in which a breeder repeatedly crosses hybrid progeny back to one of the parents, for example, a first - generation hybrid F1 , with one of the parental genotypes of the F1 , hybrid.

[0081] As used herein, the term “Marker assisted selection” or “MAS” is a process of using the presence of molecular markers (such as SNP markers), which are genetically and physically linked to a particular locus or to a particular chromosome region or allele specific markers, to select plants for the presence of the specific locus or region or allele. For example, a molecular marker genetically and physically linked to the mutant YieldPlus allele or an allele specific marker, can be used to detect and / or select e.g. cucumber plants, or plant parts, comprising the mutant YieldPlus allele. Allele specific markers are preferred markers, as they select for the allele directly.

[0082] As used herein, the term “Ml generation” or “Ml plant” in context with the present invention shall refer to the first generation that is produced directly from the mutagenic treatment. A plant grown from seeds treated with a mutagen e.g. is a representative of an Ml generation.

[0083] As used herein, the term “M2 generation” or “M2 plant” shall refer herein to the generation obtained from self-pollination of the Ml generation. A plant grown from seeds obtained from a selfpollinated Ml plant represents a M2 plant. M3, M4, etc. refers to further generations obtained after self- pollination. An “mRNA coding sequence” shall have the common meaning herein. An mRNA coding sequence corresponds to the respective DNA coding (cDNA) sequence of a gene / allele apart from that thymine (T) is replaced by uracil (U).

[0084] As used herein, the term “mutation” in a nucleic acid molecule (DNA or RNA) refers to a change of one or more nucleotides compared to the corresponding wild-type sequence, e.g. by substitution, deletion and / or insertion of one or more nucleotides. Examples of such a mutation are point mutation, nonsense mutation, missense mutation, frame shift mutation or a mutation in a regulatory sequence.

[0085] As used herein, the term “mutant protein” is herein a protein comprising one or more mutations in the nucleic acid sequence encoding the protein, whereby the mutation results in (the mutant nucleic acid molecule encoding) a "loss-of- function" protein, as e.g. measurable in vivo, e.g. by the phenotype conferred by the mutant allele.

[0086] The term “mutation” in an amino acid sequence (e.g. polypeptide or protein sequence) refers to a change of one or more amino acids compared to the corresponding wild-type amino acid sequence, e.g. by substitution, deletion and / or insertion of one or more amino acids. An amino acid substitution may be conservative or non-conservative. "Conservative" amino acid substitutions are those substitutions that do not substantially affect or decrease a function of a protein, such as the ability of the protein to induce an immune response when administered to a subject. The term conservative variation also includes the use of a substituted amino acid in place of an unsubstituted parent amino acid. Furthermore, deletions or additions which alter, add or delete a single amino acid or a small percentage of amino acids (for instance less than 5%, in some embodiments less than 1 %) in an encoded sequence are conservative variations where the alterations result in the substitution of an amino acid with a chemically similar amino acid.

[0087] The following six groups are examples of amino acids that are considered to be conservative substitutions for one another:

[0088] 1) Alanine (A), Serine (S), Threonine (T);

[0089] 2) Aspartic acid (D), Glutamic acid (E);

[0090] 3) Asparagine (N), Glutamine (Q);

[0091] 4) Arginine (R), Lysine (K);

[0092] 5) Isoleucine (I), Leucine (L), Methionine (M), Valine (V); and

[0093] 6) Phenylalanine (F), Tyrosine (Y), Tryptophan (W)-

[0094] Non-conservative substitutions are generally those that reduce an activity or function of a given protein, here the YieldPlus protein, such as the ability to induce an immune response when administered to a subject. For instance, if an amino acid residue is essential for a function of the protein, even an otherwise conservative substitution may disrupt that activity. Thus, a conservative substitution does not alter the basic function of a protein of interest.

[0095] As used herein, the term “point mutation” refers to the replacement, insertion or deletion of a single nucleotide.

[0096] As used herein, the term “nonsense mutation” refers to a mutation within a gene encoding a protein, that changes a sense codon which corresponds to an amino acid residue in a protein, into astop codon which ends synthesis of the protein at a premature position. A nonsense mutation shortens the length of the protein. A truncated protein may have loss of function.

[0097] As used herein, the term “missense mutation” refers to a mutation within a gene encoding a protein, that substitutes an amino acid of the protein by a different amino acid. A missense mutation is also called a nonsynonymous mutation. The resulting protein has a modified amino acid sequence and may have loss of function.

[0098] As used herein, the term “frame shift mutation” refers to a mutation within a gene encoding a protein by which the reading frame of the mRNA transcribed from the gene is shifted, i.e. changed, resulting in a different amino acid sequence. The resulting protein may have loss of function.

[0099] As used herein, the term “substitution” is when at least one nucleotide in the gene sequence is replaced by another nucleotide compared to the corresponding wild type nucleic acid sequence, or at least one amino acid in a protein sequence is different compared to the corresponding wild type amino acid sequence.

[0100] As used herein, the term “deletion” in the context of the invention shall mean that anywhere in a given nucleic acid sequence at least one nucleotide is missing compared to the nucleic sequence of the corresponding wild type sequence or anywhere in a given amino acid sequence at least one amino acid is missing compared to the amino acid sequence of the corresponding (wild type) sequence.

[0101] As used herein, the term “insertion” or “addition” shall mean that the nucleic acid sequence or the amino acid sequence of a protein comprises at least one additional nucleotide or amino acid compared to the corresponding wild type nucleic acid sequence or the corresponding wild type amino acid sequence, respectively.

[0102] As used herein, the term “premature stop codon” in context with the present invention means that a stop codon is present in a coding sequence (cds) which is closer to the start codon at the 5 ’-end compared to the stop codon of a corresponding wild type coding sequence.

[0103] As used herein, the term “truncation” shall be understood to mean that at least one nucleotide at either the 3’-end or the 5’-end of a nucleotide sequence is missing compared to the corresponding wild type sequence. In the context of an amino acid sequence, the term “truncation” shall be understood to mean that at least one amino acid, at either the N-terminus or the C -terminus of the amino acid sequence e.g. polypeptide or protein sequence, is missing compared to the amino acid sequence of the corresponding wild type protein.

[0104] As used herein, the term “knock-out” refers to the ablation of a gene expression (i.e., expression of the respective gene is not detectable anymore).

[0105] As used herein, the term “knock-down” refers to the decrease of a gene expression (i.e., expression of the respective gene is still detectable).

[0106] As used herein, a “loss-of -function mutation”, is a mutation which results in the gene product having a reduced function or no function at all (being partially or wholly inactivated). The degree to which the function is lost can vary. When the mutation induces a complete loss of function, it is also called anull mutation. Phenotypes associated with such mutations are generally recessive. Is also possible that some function may remain, but not at the level of the wild type allele. These are called leaky mutations. As a result of a I oss-of-fu notion mutation, the plant may produce a phenotypic change (e.g., increased fruit yield). As mentioned, it was found that the YieldPlus protein function directly reflects (and causes) a modification in fruit yield. The I oss-of-fu notion protein can thus be determined phenotypically.

[0107] In context of the present invention, a “decreased activity” of a protein shall mean a decrease in activity of a protein, e.g. the YieldPlus protein, when compared to a corresponding wild type protein. A “decreased activity” shall in one aspect comprises a decreased or abolished gene expression (e.g., knock-down or knock-out of the YieldPlus gene), in fine, leading to the apparition of the phenotype of interest (i.e., increased fruit yield). In another aspect, a “decreased activity” may refer to a loss-of- function YieldPlus protein (e.g., a mutant YieldPlus protein may have lost function compared to the wild type, functional YieldPlus protein). A loss-of-function of the protein is present when the mutant allele changes the phenotype from the wild type phenotype, i.e. normal yield when the wild type allele is present, into an increased fruit yield when the mutant allele is present.

[0108] As used herein, the term “Complementary strands” refers to two strands of complementary sequence and may be referred to as sense (or plus) and anti-sense (or minus) strands for double stranded DNA. The sense / plus strand is, generally, the transcribed sequence of DNA (or the mRNA that was generated in transcription), while the anti-sense / minus strand is the strand that is complementary to the sense sequence. For any of the sequences provided herein only one strand of the sequence is given, but the complementary strand of the given strand is also encompassed herein. The complementary nucleotides of DNA are A complementary to T, and G complementary to C. The complementary nucleotides of RNA are A complementary to U, and G complementary to C.

[0109] As used herein, the term “Yield” or “fruit yield” or “average yield” refers to the average number of fruits per plant (FrPP) and / or the average fruit weight (grams) per plant (GrPP) at a single harvest time-point. The single harvest time-point is in line with growers practice and chosen to maximize the number of fruits for each plant. For each typology, growers will identify the standard size (length and diameter) that need to be reached at each single harvesting time point. For example, for a European long type, standard size is defined as 34 to 38 cm long and 4cm to 4.5cm diameter. Thus, in one aspect all fruits per plant are harvested and only the ones fitting the standard size (length and diameter) are chosen. This is done for each plant line or variety grown under the same conditions and the average FrPP and / or GrPP of each line or variety is calculated. For clarity, the terms “average yield” or “fruit yield” do not refer to the average individual fruit weight, i.e. the average fruit weight (grams) per fruit in a particular plant.

[0110] As used herein, the term “average individual fruit weight” refers to the total fruit weight (grams) of a plant divided by the number of fruits of the plant, i.e. reflecting the average fruit weight (grams) per fruit in a particular plant.

[0111] As used herein, the term “increased fruit yield” refers to a cucumber plant, typically mutant, having a statistically significantly higher average number of fruits per plant (FrPP) and / or a significantlyhigher average fruit weight per plant (GrPP) compared to a reference plant, typically wild-type, at the same stage of development and when grown under the same environmental conditions. The reference plant is preferably an isogenic plant homozygously comprising a wild-type allele of the YieldPlus gene. Fruit yield comparisons are made in the same growth conditions, wherein the fruits are harvested at a same harvest time point. Preferably field trials are carried out in several replicates (2, 3, or more) in several locations (2, 3, or more), with sufficient plants (e.g. at least 10, 15, 20, 30, 40, or more plants per line) comprising a mutant allele (e.g., SEQ ID NO: 4) and the wild-type allele (e.g. SEQ ID NO: 1).

[0112] As used herein, the term “plant part” refers to any part of a plant including but not limited to the shoot, root, stem, seeds, fruits, leaves, petals, flowers, ovules, branches, petioles, internodes, pollen, stamen, rootstock, scion and the like.

[0113] The term “resistance” is as defined by the ISF (International Seed Federation) Vegetable and Ornamental Crops Section for describing the reaction of plants to pests or pathogens, and abiotic stresses for the Vegetable Seed Industry. Specifically, by resistance, it is meant the ability of a plant variety to restrict the growth and development of a specified pest or pathogen and / or the damage they cause when compared to susceptible plant varieties under similar environmental conditions and pest or pathogen pressure. Resistant varieties may exhibit some disease symptoms or damage under heavy pest or pathogen pressure.

[0114] As used herein, the term “susceptible” refers to a plant that is unable to restrict the growth and development of a specified pest or pathogen.

[0115] As used herein, the term “inbred” or “line” refers to a relatively true-breeding strain.

[0116] As used herein, the term “phenotype” refers to the observable characters of an individual cell, cell culture, organism (e.g. a plant), or group of organisms which results from the interaction between that individual genetic makeup (i.e. genotype) and the environment.

[0117] As used herein, the terms “introgression”, “introgressed” and “introgressing” refer to the process whereby genes of one species, variety or cultivar are moved into the genome of another species, variety or cultivar, by crossing those species. The crossing may be natural or artificial. The process may optionally be completed by backcrossing to the recurrent parent, in which case introgression refers to infiltration of the genes of one species into the gene pool of another through repeated backcrossing of an interspecific hybrid with one of its parents. An introgression may be also described as a heterologous genetic material stably integrated in the genome of a recipient plant.

[0118] In the present specification, a comparison between two or more plants or fruits, in particular a comparison between a cucumber plant according to the invention with an isogenic plant not comprising a mutant allele of the Yieldplus gene, is understood to be a comparison between plants or fruits at the same stage of maturity or at the same stage post-harvest, grown in the same environmental conditions.

[0119] Detailed description of the invention

[0120] The present inventors have identified that the presence of a mutant allele of a gene named YieldPlus on chromosome 3 in the genome of Cucumis sativus plants, provides to these plants an increased fruit yield.

[0121] The present invention thus provides cultivated cucumber plant or plant part that displays an increased fruit yield as well as a method for producing a cucumber plant that exhibits the increased yield. The present invention also discloses a method for screening and / or selecting a plant having an increased fruit yield.

[0122] The present invention also discloses molecular genetic markers, especially KASP markers, linked the phenotype of interest (i.e., increased fruit yield). Plants obtained through the uses of such molecular markers are also provided. Methods for identifying further molecular markers linked to the phenotype of interest are also provided.

[0123] Said increased fruit yield is moreover easily transferable to different genetic backgrounds and the invention also extends to different methods allowing the transfer or introgression of the mutated allele conferring the phenotype as well as method for introducing the mutated allele in a cucumber plant.

[0124] Said increased fruit yield is characterized by increased total fruit weight per plant, increased fruit number per plant, and / or increased individual fruit weight per fruit of the plant.

[0125] The invention also provides methods for improving the yield of cucumber production and methods for producing cucumber fruits.

[0126] In one aspect, the invention relates to a Cucumis sativus plant comprising a mutant allele of a YieldPlus gene on chromosome 3 in its genome, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele. In one aspect, the mutation results in an increased fruit yield, particularly in an increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

[0127] The plant according to the invention is preferably a cultivated cucumber, namely a cultivated C. sativus var. sativus plant.

[0128] The YieldPlus gene has been mapped to chromosome 3 of the genome (nucleotide 31790724 to nucleotide 31792269 of reference genome Cucumis sativus (Cucumber 9930 (Chinese Long) v3.0). A sequence of a wild-type allele of the YieldPlus gene is set forth in SEQ ID NO: 1 . A wild-type coding sequence is set forth in SEQ ID NO: 2. Wild-type YieldPlus alleles also encompass any nucleotide sequence which retains the activity and expression level of the YieldPlus gene as encoded by SEQ ID NO: 1. In some embodiments the wild-type YieldPlus allele comprises a nucleotide sequence with at least 80%, preferably at least 90%, still preferably at least 95%, more preferably at least 98%, most preferably at least 99% identity with SEQ ID NO: 1. A translated sequence of the wild-type YieldPlus gene, i.e. the wild-type amino acid sequence of the YieldPlus protein is set forth in SEQ ID NO:3.

[0129] The YieldPlus protein (SEQ ID NO:3) is a zinc finger protein 8-like transcription factor. The YieldPlus protein has been annotated and reported to be involved in the cell’s response to gibberellin, trichome differentiation and trichome morphogenesis. In some instances, I oss-of-fu notion alleles of the YieldPlus gene can be identified by assessing a C. sativus plant or cell’s response to a gibberellin stimulus and / or trichome morphogenesis and / or trichome differentiation in a cell, and comparing said response to the response of a plant or cell comprising a wild-type allele of the YieldPlus gene to a gibberellin stimulus and / or trichome morphogenesis and / or trichome differentiation. Loss-of-fu notion alleles of the YieldPlus are also those which have the same effect, or a similar effect, as a reference loss-of-function allele, e.g. an allele comprising a YieldPlus gene sequence encoding a protein comprising a non-conservative substitution within the zinc finger domain, as located from amino acids 85 to 120 of the amino acid sequence of the YieldPlus protein, as set forth in SEQ ID NO:3. In some instances, the reference loss-of-function allele comprises a YieldPlus gene encoding a protein comprising a non-conservative substitution within the motif QAALGH in positions 99 to 104 of SEQ ID NO: 3. The same effect, or a similar effect refers, in particular to a same or similar effect on any phenotype distinguishing a plant or a cell with the reference allele and a wild-type plant or cell, in particular an effect on cell’s response to a gibberellin stimulus and / or trichome morphogenesis and / or trichome differentiation, and / or an effect on fruit yield as defined in the present specification. A same or similar effect may be an effect of identical nature (e.g. the presence or absence or a decrease or increase of a given phenotype), with the same intensity or a higher or lower intensity than the reference plant. Loss-of-function of the YieldPlus gene can also be assessed by a decrease or loss of the YieldPlus protein’s ability to bind target DNA. Wild-type YieldPlus protein also encompass any amino acid sequence which retains the activity and expression level of the YieldPlus protein as encoded by SEQ ID NO: 3. In some the wild-type YieldPlus protein comprises an amino acid sequence with at least 90%, preferably at least 95%, still preferably at least 98%, more preferably at least 99% identity with SEQ ID NO:3.

[0130] As shown in the present disclosure, a decreased activity of the YieldPlus protein causes a phenotype of interest, i.e. an increased fruit yield. A modification (i.e., mutation...) leading to a decreased or abolished activity of the YieldPlus protein, or a decreased or abolished expression of the YieldPlus protein, will cause the phenotype of interest. The presence of a mutant YieldPlus allele may result into a decreased or abolished activity or expression of the YieldPlus protein.

[0131] In one embodiment, the mutant allele of the YieldPlus gene is a loss-of-function allele, i.e. it comprises at least one loss-of-function mutation. The degree to which the function is lost can vary. The sequence of the mutant allele can differ from the wild-type sequence of the gene by at least one nucleotide substitution, insertion or deletion in said sequence. In particular, the mutation can be a point mutation. More particular, the mutation can be a single nucleotide polymorphism (SNP). The mutant allele of the YieldPlus gene can also differ from the wild-type allele by the insertion or the deletion of one or more nucleic acid segments, including the deletion of a part or the totality of the gene or a regulatory sequence of the gene, e.g. a promoter, enhancer, 3’-UTR, 5’-UTR and / or polyA sequence.

[0132] In some aspects, the loss-of-function mutation is a null mutation. A null mutation prevents expression of an active YieldPlus protein, i.e. the function is fully lost. Accordingly, the mutant allele of the YieldPlus gene may be a null allele or knockout allele.

[0133] In some embodiments, the mutation is a nonsense mutation. A nonsense mutation causes a premature stop in the translation of the mRNA into a protein, resulting into the expression of a truncated form of the YieldPlus protein.

[0134] In some aspects, the mutation is a missense mutation. The missense mutation may consist in the substitution of one amino acid of the YieldPlus protein by another. In some embodiments, more than one amino acid are substituted. A missense mutation preferably results in a protein with decreased activity or no activity at all. A preferred missense mutation is a mutation inducing the substitution of the glycine residue in position 253 of SEQ ID NO: 3. In particular, the glycine residue in position 253 of SEQ ID NO: 3 is substituted by a serine. More particularly, this missense mutation has been identified by the inventors in a mutant plant and introgressed into a different genotype. The amino acid sequence of the corresponding mutant allele is set forth in SEQ ID NO: 5. In some embodiments the at least one mutation is a G757A mutation in the coding sequence of the Yieldplus gene, as set forth in SEQ ID NO: 2. In some embodiments the at least one mutation is a G1077A mutation in the sequence set forth in SEQ ID NO: 1.

[0135] In some embodiments, the mutation is situated within the zinc finger domain of the YieldPlus protein, in particular from amino acid residues 85 to 120 of SEQ ID NO:3. In some embodiments, the mutation is located outside of the zinc finger domain of the YieldPlus, wherein said zinger domain is located from residues 85 to 120 of the amino acid sequence of the YieldPlus protein, as set forth in SEQ ID NO:3. In particular, the mutation is located at a position from amino acid residues 1 to 84 and / or from amino acid residues 121 to 277 C-terminally of the zinc finger domain of the amino acid sequence of the YieldPlus protein, as set forth in SEQ ID NO:3. More particularly, the mutation is located within the C-terminal disordered region of the YieldPlus protein. Preferably, the mutation is located at a position from amino acid residues 150 to 277 of the YieldPlus protein.

[0136] In some embodiments, the substitution is a non-conservative substitution. In the context of the present disclosure, non-conservative substitutions may be defined by substitutions between or outside the groups of amino acids reflected as follows:

[0137] 1) Alanine (A), Serine (S), Threonine (T);

[0138] 2) Aspartic acid (D), Glutamic acid (E);

[0139] 3) Asparagine (N), Glutamine (Q);

[0140] 4) Arginine (R), Lysine (K);

[0141] 5) Isoleucine (I), Leucine (L), Methionine (M), Valine (V); and

[0142] 6) Phenylalanine (F), Tyrosine (Y), Tryptophan (W)-

[0143] The importance of the hydropathic amino acid index in conferring interactive biologic function on a protein is generally understood in the art. It is accepted that the relative hydropathic character of the amino acid contributes to the secondary structure of the resultant protein, which in turn defines the interaction of the protein with other molecules, for example, enzymes, substrates, receptors, DNA, antibodies, antigens, and the like. Each amino acid has been assigned a hydropathic index on the basis of their hydrophobicity and charge characteristics, these are: isoleucine (+4.5); valine (+4.2); leucine (+3.8) ; phenylalanine (+2.8); cysteine / cystine (+2.5); methionine (+1.9); alanine (+1.8); glycine (-0.4); threonine (-0.7); serine (-0.8); tryptophane (-0.9); tyrosine (-1 .3); proline (-1 .6); histidine (-3.2); glutamate (-3.5); glutamine (-3.5); aspartate (-3.5); asparagine (-3.5); lysine (-3.9); and arginine (-4.5).

[0144] In some aspects, the at least one mutation is in the coding sequence of the YieldPlus gene. In other aspects, the at least one mutation is a mutation in a regulatory sequence of the YieldPlus gene. Regulatory sequences are sequences which control a gene expression. Mutations within such sequences are known to affect the expression of the gene that they control. In some aspects, the mutation is in a regulatory sequence selected from a promoter, an enhancer, a 3’-UTR sequence, a 5’- UTR sequence and a polyA sequence.

[0145] The mutation can be of any type. In particular, the mutation is selected from a substitution, a deletion or an addition of one or more nucleotides. In particular from 1 to 100 nucleotides, in particular from 1 to 50 nucleotides, more particularly from 1 to 20 nucleotides, most particularly from 1 to 10 nucleotides are mutated. In some embodiments, 1 , 2, 3, 4, 5 ,6, 7, 8, 9 or 10 nucleotides are mutated.

[0146] The mutation can affect one or more codons encoding the YieldPlus protein. In some embodiment, the mutant allele comprises at least one codon inserted or duplicated and / or at least one codon changed into a different codon (e.g., through a single nucleotide change), at least one codon changed into a stop codon, or at least one codon deleted. In particular, from 1 to 30 codons, in particular from 1 to 20 codons, more particularly from 1 to 10 codons, most particularly from 1 to 5 codons are mutated. In some embodiments, 1 , 2, 3, 4, 5 ,6, 7, 8, 9 or 10 codons are mutated.

[0147] Where the mutation affects the amino acid sequence of the YieldPlus protein, from 1 to 30 amino acids, in particular from 1 to 20 amino acids, more particularly from 1 to 10 amino acids, most particularly from 1 to 5 amino acids may be mutated. In some embodiments, 1 , 2, 3, 4, 5 ,6, 7, 8, 9 or 10 amino acids are mutated.

[0148] Mutant alleles and corresponding markers can be identified by methods known in the art. The mutation in the mutant YieldPlus allele can be induced via methods such as mutagenesis or by means of genetic engineering. Mutagenesis methods and methods of genetic engineering are known in the art and are described below in more details. Accordingly, the plants according to the invention may be obtained by different processes and are preferably not exclusively obtained by means of an essentially biological process.

[0149] In some embodiments, said mutant allele is introgressed in the genome of said Cucumis sativus plant.

[0150] In some embodiments, said mutant allele is in homozygous form. In alternative embodiments, said mutant allele is in a heterozygous form.

[0151] Said mutant allele confers the plant of the invention with an increased fruit yield. In particular, the fruit yield is increased in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

[0152] In some embodiments, the plant of the invention produces at least 10%, preferably at least 20%, still preferably at least 50% more fruits in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene. In some embodiments, said Cucumis sativus plant produces at least 1 more fruit, in particular at least 2 more fruits, more particularly at least 5 more fruits, still more particularly at least 10 more fruits in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene. In some embodiments, said Cucumis sativus plant produces at least 10 fruits or more, preferably at least 12 fruits or more.

[0153] In some embodiments, the fruit weight of said plant is at least 10%, preferably at least 20%, still preferably at least 50% higher in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene.

[0154] In some embodiments, the average individual fruit weight of the fruits of said plant is decreased by less than 50%, in particular less than 40%, more particularly less than 30%, still more particularly less than 20%, even more particularly less than 10%, most particularly by less than 5%, in comparison to the average individual fruit weight of the fruits of an isogenic plant which comprises a wild-type allele of the YieldPlus gene, or is not decreased. In some embodiments, the average individual fruit weight of the fruits of said plant is increased by at least 10%, in particular at least 20%, more particularly at least 30%, still more particularly at least 40%, even more particularly at least 50% in comparison to the average individual fruit weight of the fruits of an isogenic plant which comprises a wild-type allele of the YieldPlus gene.

[0155] In some embodiments, the plant according to the invention comprises an increased average number of fruits, in particular commercial-sized fruits, per node, in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene. In some embodiments, the average number of fruits per node, in particular commercial-sized fruits, is increased by at least 5%, in particular at least 10%, more particularly at least 20%, still more particularly at least 30%, even more particularly at least 40%, most particularly at least 50%, in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene. In some embodiments, the average number of fruits per node is increased by 5- 100%, in particular 5-75%, more particularly 5-50% in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene. The average number of fruits per node, in particular commercialsized fruits, is measured at a single harvest time-point, at a commercial stage, which can be chosen in line with growers practice, as defined in the present specification in relation the measurement of yield parameters.

[0156] In some embodiments, the plant according to the invention has a faster fruit filling than an isogenic plant which comprises a wild-type allele of the YieldPlus gene. Faster fruit filling is assessedby measuring the time elapsed from anthesis to harvest. In some embodiments, the time elapsed from anthesis to harvest of a plant according to the invention is decreased by at least 5%, in particular at least 10%, more particularly at least 20%, still more particularly at least 30%, even more particularly at least 40%, most particularly at least 50%, in comparison to an isogenic plant which comprises a wildtype allele of the YieldPlus gene. In some embodiments, the time elapsed from anthesis to harvest of a plant according to the invention is decreased by 5-75%, in particular 5-50%, more particularly 5-30% in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene. The time of harvest is a single harvest time-point, at a commercial stage, chosen in line with growers practice, as defined in the present specification in relation to the measurement yield parameters.

[0157] In some embodiments, said cucumber plant is not exclusively obtained by means of an essentially biological process.

[0158] Preferably, a C. sativus plant according to the invention is a cultivated plant or line, more preferably a commercial plant or line or hybrid. Such a plant or line thus has generally 4 primary branches or less, generally less than 3 and more preferably has a single primary branch.

[0159] Such a commercial plant or line preferably also exhibits resistance to viruses. Preferably, a commercial plant is resistant to ZYMV (Zucchini Yellow Mosaic Virus) and / or to CVYV (Cucumber Vein Yellowing Virus). Resistances to PRSV (Papaya Ringspot Virus) and WMV (Watermelon Mosaic Virus) and CMV (Cucumber Mosaic Virus) are also generally found in commercial plants. A plant of the invention is thus advantageously resistant at least to powdery mildew and to potyviruses.

[0160] Other traits, such as resistances or tolerances, are also envisaged according to the invention, inter alia resistance to downy mildew caused Pseudoperonospora cubensis, resistance to Fusaria caused by Fusariumoxysporum f.sp. cucumerinum or by Fusarium oxysporum f.sp. radicis cucumerinum, resistance to scab caused by Cladosporium cucumerinum, resistance to WMV, to CYSDV and to CCYV, resistance to angular leaf sport and, resistance to anthracnose.

[0161] According to other aspects, the present invention is directed to one or more plant parts of a plant according to the invention. Such plant part comprises a mutant allele of the YieldPlus gene. The different features of said YieldPlus gene that have been defined in relation with the above aspects of the invention apply mutatis mutandis to this aspect of the invention.

[0162] According to another embodiment, the plant part is a seed, explant, reproductive material, scion, cutting, fruit, root, root tip, rootstock, pollen, ovule, embryo, meristem, callus, cotyledon, hypocotyl, protoplast, leaf, anther, stem, petiole or flower. A particularly preferred plant part according to the invention is a seed produced by the plant of the invention. Preferably, the seed comprises homozygously the mutant alleles of the YieldPlus gene.

[0163] In some embodiments, the plant part is a plant cell.

[0164] Accordingly, one aspect of the invention relates to a cell of a C. sativus plant according to the invention, i.e. a plant cell comprising a mutant allele of the YieldPlus gene. The different features of saidYieldPlus gene that have been defined in relation with the above aspects of the invention apply mutatis mutandis to this aspect of the invention.

[0165] A plant cell of the invention may have the capacity to be regenerated into a whole plant.

[0166] The invention is also directed to plant cells which are not regenerable, and thus are not capable of giving rise to a whole plant.

[0167] Preferably, a plant cell according to the invention is derived from a seed, reproductive material, scion, cutting, fruit, root, root tip, rootstock, pollen, ovule, embryo, meristem, callus, cotyledon, hypocotyl, protoplast, leaf, anther, stem, petiole or flower.

[0168] The invention further relates to a seed from a plant of the invention. The seed may be produced by such a plant after selfing or crossing. Preferably, said seed is capable of germinating into a plant with increased fruit yield. The invention also provides C. sativus plants grown from the seeds of the invention.

[0169] The invention further relates to a seed for producing a plant according to the invention. Said seed can be grown into a plant of the invention. In an embodiment, said plant, seed, cell or part comprises a mutant allele of a YieldPlus gene on chromosome 3 in its genome, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield of a plant grown from said seed, in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

[0170] The invention also relates to a population of C. sativus seeds according to the invention, wherein said population comprises at least 2 seeds, especially at least 10 seeds, particularly at least 100 seeds, even more particularly at least 105or 106seeds.

[0171] The present invention is also directed to an in vitro cell or tissue culture of regenerable cells of the plant as defined above according to the present invention. Preferably, the regenerable cells are derived from a seed, reproductive material, scion, cutting, fruit, root, root tip, rootstock, pollen, ovule, embryo, meristem, callus, cotyledon, hypocotyl, protoplast, leaf, anther, stem, petiole or flower of a plant of the invention, and comprise in their genome a mutant allele of the YieldPlus gene as described here above.

[0172] The tissue culture will preferably be capable of regenerating plants having the physiological and morphological characteristics of the foregoing C. sativus plant, and of regenerating plants having substantially the same genotype as the foregoing C. sativus plant. The present invention also provides C. sativus plants regenerated from the tissue cultures of the invention.

[0173] The invention also provides a protoplast of the plant defined above, or from the tissue culture defined above, said protoplast comprising in its genome a mutant allele of the YieldPlus gene, as described here above.

[0174] The present invention also encompasses asexual processes of propagation. Accordingly, one aspect of the invention relates to a method of producing C. sativus plant with an increased fruit yield, comprising:(a) obtaining a part of a plant according to the invention, such as a cutting; and(b) vegetatively propagating said plant part to generate a C. sativus plant from said plant part.

[0175] The generated C. sativus plant comprises in its genome a mutant allele of the YieldPlus gene, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield.

[0176] All the embodiments detailed above in connection with the first aspect of the invention are also embodiments according to this aspect of the invention, especially with regard to the features of the mutation(s) conferring the phenotype of interest.

[0177] The invention described above encompasses a plant, seed, plant part and cell of any ploidy levels. It encompasses inter alia diploid, triploid, tetrapioid and / or allopolyploid Cucumis sativus plant, plant part, cell or seed.

[0178] The invention further relates to an isolated polynucleotide comprising a mutant allele of the YieldPlus gene, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , wherein said mutant allele confers increased fruit yield to a plant comprising said mutant allele, in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene. In one embodiment, said polynucleotide is isolated or susceptible to be isolated from a plant according to the invention. By ’’susceptible to be isolated”, it is meant that said polynucleotide can be found, before isolation, within a plant according to the invention. Accordingly, in one embodiment the invention relates to a plant, plant part or plant cell comprising a polynucleotide according to the invention.

[0179] In some embodiments, said polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 2. In particular, said polynucleotide comprises a nucleotide sequence having at least 85%, more particularly at least 90%, even more particularly at least 95%, at least 96%, at least 97%, at least 98% or at least 99% with SEQ ID NO: 2. In some embodiments, said polynucleotide comprises a nucleotide sequence having at least 80% sequence identity with SEQ ID NO: 1. In particular, said polynucleotide comprises a nucleotide sequence having at least 85%, more particularly at least 90%, even more particularly at least 95%, at least 96%, at least 97%, at least 98% or at least 99% with SEQ ID NO: 1 .

[0180] In some embodiments, said polynucleotide encodes a polypeptide having at least 80% sequence identity with SEQ ID NO: 3. In particular, said polynucleotide encodes a polypeptide having at least 85%, more particularly at least 90%, even more particularly at least 95%, at least 96%, at least 97%, at least 98% or at least 99% with SEQ ID NO: 3.

[0181] In some embodiments, said polypeptide comprises the sequence of SEQ ID NO: 3 in which at least 1 %, in particular at least 2%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99% or 100% of said sequence has been modified. In someembodiments, said polypeptide comprises the sequence of SEQ ID NO: 3 in which 1-10%, 10-20%, 20- 30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90% or 90-100% of said sequence has been modified. In some embodiments, said polypeptide comprises the sequence of SEQ ID NO: 3, or a sequence with at least 95%, in particular at least 98%, more particularly at least 99% identity with SEQ ID NO:3, in which at least 1 %, in particular at least 2%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99% or 100% of said sequence has been deleted and / or truncated, and / or in which 1-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60- 70%, 70-80%, 80-90% or 90-100% of said sequence has been deleted and / or truncated. In some embodiments, said polypeptide comprises the sequence of SEQ ID NO: 3, or a sequence with at least 95%, in particular at least 98%, more particularly at least 99% identity with SEQ ID NO:3, in which at least 1 %, in particular at least 2%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99% or 100% of the C-terminal domain of the YieldPlus protein, preferably of the fragment from amino acid residues 150 to 277 of the YieldPlus protein, has been deleted and / or truncated, and / or in which 1-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90% or 90-100% of the C-terminal domain of the YieldPlus protein has been deleted and / or truncated.

[0182] In some embodiments, said polynucleotide comprises the sequence of SEQ ID NO:4.

[0183] In some embodiments, said polynucleotide encodes the sequence of SEQ ID NO:5.

[0184] The invention also relates to a polypeptide encoded by a polynucleotide of the invention. In particular, the invention relates to an isolated polypeptide comprising an amino acid sequence with at least 95%, in particular at least 98%, more particularly at least 99% identity with SEQ ID NO:3, wherein said polypeptide comprises at least one mutation in the amino acid sequence, in comparison to the sequence of the corresponding wild-type polypeptide as set forth in SEQ ID NO:3, wherein said mutation confers a loss-of-function phenotype to said polypeptide. In some embodiments, said polypeptide is as defined in the present specification.

[0185] Methods of measuring polynucleotide or polypeptide homology or identity are known in the art. For example, the UWGCG Package provides the BESTFIT program which can be used to calculate homology (e.g. used on its default settings) (Devereux et al, 1984, Nucleic Acids Research 12:387-395; the disclosures of which are incorporated herein by reference).

[0186] The PILEUP and BLAST algorithms can also be used to calculate homology or line up sequences (typically on their default settings), for example as described in Altschul, 1993, J Mol Evol 36:290-300; Altschul ef al, 1990, J Mol Biol 215:403-10, the disclosures of which are incorporated herein by reference). The BLAST program uses as defaults a word length (W) of 11 , the BLOSUM62 scoring matrix (see Henikoff & Henikoff, 1992, Proc. Natl. Acad. Sci. USA 89:10915-10919; the disclosures of which are incorporated herein by reference) alignments (B) of 50, expectation (E) of 10, M=5, N=4, and a comparison of both strands.

[0187] In some embodiments, said polypeptide has the sequence of SEQ ID NO:5.

[0188] The invention is also directed to the use of the information provided herewith by the present inventors, namely a mutant YieldPlus allele conferring an improved fruit yield to C. sativus plants compared to the wild type allele, as set forth in SEQ ID NO: 1 . This knowledge can be used inter alia to generate new mutant plants comprising a mutated YieldPlus allele and expressing the phenotype of interest (i.e., increased fruit yield). Indeed, new mutant can be generated de novo by e.g. targeted gene editing techniques, such as CRISPR based techniques or by mutagenesis, such as radiation induced mutagenesis or chemically induced mutagenesis.

[0189] The skilled person can for example generate a plant with a mutant YieldPlus gene and determine whether it results in an increased fruit yield compared to the same plant with the wild type YieldPlus gene, in particular in homozygous form (i.e. isogenic plants). A plant homozygous for the mutant allele can be generated by selfing the plant and then growing the homozygous plant to determine whether the yield is higher in the homozygous mutant plant in comparison to a wild-type control.

[0190] The invention is thus also directed to a method for generating or producing cucumber plant comprising a mutant allele of the YieldPlus gene. Especially, such method may comprise mutagenizing one or more Cucumis sativus seed, plant or plant part and screening / selecting the M1 or M2 generation for YieldPlus mutant alleles.

[0191] The present invention also relates to a method for producing a C. sativus plant with an increased fruit yield, comprising the introduction of at least one mutation in the YieldPlus gene on chromosome 3 in the genome of a C. sativus plant, wherein said mutation is introduced in the sequence of the gene or in a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield.

[0192] In some embodiments, the present invention also relates to a method for producing a C. sativus plant with an increased fruit yield, comprising: a) Introducing one or more mutations in cucumber plant(s), seed(s) or plant part(s), b) Optionally, determining if the plant, seed or plant part under a) presents an increased yield compared to a plant not having said at least one mutation; and c) Selecting a plant that comprises a mutant allele of a YieldPlus gene, wherein said mutant allele comprises at least one mutation in the sequence of the gene or in a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele, as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield.

[0193] In some embodiments, said mutation is introduced by mutagenesis or genome editing. In particular, said mutation is introduced by a technique selected from ethyl methanesulfonate (EMS) mutagenesis, oligonucleotide directed mutagenesis (ODM), Zinc finger nuclease (ZFN) technology, Transcription Activator-Like Effector Nucleases (TALENs) the CRISPR / Cas system, the CRISPR / Cpf system engineered meganuclease, re-engineered homing endonucleases and DNA guided genome editing. In some embodiments, said mutation is introduced by ethyl methanesulfonate (EMS) mutagenesis. In other embodiments, said mutation is introduced by an endonuclease-mediatedtechnique selected from the Zinc finger nuclease (ZFN) technology, the TALEN technology, the CRISPR / Cas system and the CRISPR / Cpf system, preferably the CRISPR / Cas system, most preferably the CRISPR / Cas9 system.

[0194] Mutagenesis methods are known in the art and include chemical mutagenesis using ethyl methanesulfonate (EMS). Other chemical mutagenic agents include but are not limited to, diethyl sulfate (des), ethyleneimine (ei), propane sultone, N-methyl-N-nitrosourethane (mnu), N-nitroso-N-methylurea (NMU), N-ethyl-N-nitrosourea(enu), and sodium azide.

[0195] Alternatively, the mutations can be induced by means of irradiation, which is for example selected from x-rays, fast neutrons, UV radiation.

[0196] Mutagenesis techniques can be followed by an identification method such as TILLING. TILLING (Targeting Induced Local Lesions IN Genomes) is a general reverse genetics technique that uses traditional chemical mutagenesis methods to create libraries of mutagenized individuals that are later subjected to high throughput screens for the discovery of mutations. TILLING combines chemical mutagenesis with mutation screens of pooled PCR products, resulting in the isolation of missense and non-sense mutant alleles of the targeted genes. Thus, TILLING uses traditional chemical mutagenesis (e.g. EMS or MNU mutagenesis) or other mutagenesis methods (e.g. radiation such as UV) followed by high-throughput screening for mutations in specific target genes, such as the YieldPlus gene according to the invention. S1 nucleases, such as CEL1 or ENDO1 , are used to cleave heteroduplexes of mutant and wild type target DNA and detection of cleavage products using e.g. electrophoresis such as a Ll- COR gel analyzer system, see e.g. Henikoff et al. Plant Physiology 2004, 135: 630-636. TILLING has been applied in many plant species, including cucumber (Fraenkel et al., BMC Res Notes. 2014 Nov 26;7:846. doi: 10.1186 / 1756-0500-7-846). Also EcoTILLING, whereby mutants in natural populations are detected, has been widely used, see Till et al. 2006 (Nat Protoc 1 : 2465-77) and Comai et al. 2004 (Plant J 37: 778-86).

[0197] In another embodiment of the invention, the mutation(s) is(are) induced by means of genetic engineering.

[0198] The genetic engineering means which can be used include the use of all such techniques called New Breeding Techniques which are various new technologies developed and / or used to create new characteristics in plants through genetic variation, the aim being targeted mutagenesis, targeted introduction of new genes or gene silencing (RdDM). Example of such new breeding techniques are targeted sequence changes facilitated through the use of Zinc finger nuclease (ZFN) technology (ZFN- 1 , ZFN-2 and ZFN-3, see U.S. Pat. No. 9,145,565, incorporated by reference in its entirety), Oligonucleotide directed mutagenesis (ODM), Cisgenesis and intragenesis, RNA-dependent DNA methylation (RdDM, which does not necessarily change nucleotide sequence but can change the biological activity of the sequence), Grafting (on GM rootstock), Reverse breeding, Agro-infiltration (agro-infiltration "sensu stricto", agro-inoculation, floral dip), Transcription Activator-Like Effector Nucleases (TALENs, see U.S. Pat. Nos. 8,586,363 and 9,181 ,535, incorporated by reference in their entireties), the CRISPR / Cas system (see U.S. Pat. Nos. 8,697,359; 8,771 ,945; 8,795,965; 8,865,406;8,871 ,445; 8,889,356; 8,895,308; 8,906,616; 8,932,814; 8,945,839; 8,993,233; and 8,999,641 , which are all hereby incorporated by reference), engineered meganuclease re-engineered homing endonucleases, DNA guided genome editing (Gao et al., Nature Biotechnology (2016), doi: 10.1038 / nbt.3547), and Synthetic genomics. A major part of today’s targeted genome editing, another designation for New Breeding Techniques, is the applications to induce a DNA double strand break (DSB) at a selected location in the genome where the modification is intended. Directed repair of the DSB allows for targeted genome editing. Such applications can be utilized to generate mutations (e.g., targeted mutations or precise native gene editing) as well as precise insertion of genes (e.g., cisgenes, intragenes, or transgenes). The applications leading to mutations are often identified as site-directed nuclease (SDN) technology, such as SDN1 , SDN2 and SDN3. For SDN1 , the outcome is a targeted, non-specific genetic deletion mutation: the position of the DNA DSB is precisely selected, but the DNA repair by the host cell is random and results in small nucleotide deletions, additions or substitutions. For SDN2, a SDN is used to generate a targeted DSB and a DNA repair template (a short DNA sequence identical to the targeted DSB DNA sequence except for one or a few nucleotide changes) is used to repair the DSB: this results in a targeted and predetermined point mutation in the desired gene of interest. As to the SDN3, the SDN is used along with a DNA repair template that contains new DNA sequence (e.g. gene). The outcome of the technology would be the integration of that DNA sequence into the plant genome. The most likely application illustrating the use of SDN3 would be the insertion of cisgenic, intragenic, or transgenic expression cassettes at a selected genome location. A complete description of each of these techniques can be found in the report made by the Joint Research Center (JRC) Institute for Prospective Technological Studies of the European Commission in 2011 and titled “New plant breeding techniques - State-of-the-art and prospects for commercial development”, which is incorporated by reference in its entirety.

[0199] Applications of gene-editing techniques in cucumber have been reported, e.g. in Hu et al (2017 Mol Plant 10: 1575-1578), Sherman et al. (2016) Mol Plant Pathol 7: 1140-1153.

[0200] In an embodiment, the invention relates to a method for obtaining a C. sativus plant or seed carrying one or more mutations in its genome. Such a method is illustrated in the Examples and may comprise: a) treating M0 seeds of a C. sativus plant to be modified with a mutagenic agent to obtain M1 seeds; b) growing plants from the thus obtained M1 seeds to obtain M1 plants; c) producing M2 seeds by self-fertilisation of M1 plants; and d) optionally repeating step b) and c) n times to obtain M2+n seeds.

[0201] In this method, the M1 seeds of step a) can be obtained via chemical mutagenesis such as EMS mutagenesis. Other chemical mutagenic agents include but are not limited to, diethyl sulfate (des), ethyleneimine (ei), propane sultone, N-methyl-N-nitrosourethane (mnu), N-nitroso-N-methylurea (NMU), N-ethyl-N-nitrosourea(enu), and sodium azide. Alternatively, the mutations are induced by means of irradiation, which is for example selected from x-rays, fast neutrons, UV radiation.

[0202] In another embodiment of the invention, the mutation(s) is(are) induced by means of genetic engineering. Such mutations also include the integration of sequences, as well as the substitution of residing sequences by alternative sequences. A further aspect of the invention relates to the use of a C. sativus plant or seed according to the invention, as a breeding partner in a breeding program for conferring increased fruit yield to progeny C. sativus plants. By crossing a C. sativus plant of the invention with another plant, e.g. having a different genotype, for instance a susceptible or less resistant plant, it is possible to transfer the mutant YieldPlus allele, conferring the desired phenotype, to the progeny.

[0203] In such a breeding program, the selection of the progeny displaying the desired phenotype, or bearing sequences linked to the desired phenotype, can advantageously be carried out with markers, e.g. the markers disclosed in the present specification. The selection of the progeny having the desired phenotype can also be made by assessing the phenotype of the progeny plants.

[0204] The invention is also directed to the use of said plants in a program aiming at identifying, sequencing and / or cloning the genetic sequences conferring the desired phenotype.

[0205] According to another aspect, the invention also concerns methods for the production of a C. sativus plant having increased fruit yield, comprising the following steps:(a) crossing a C. sativus plant according to the invention with itself or with a second C. sativus plant, preferably of a different genotype, to produce one or more progeny plants;(b) selecting a progeny plant comprising a mutant allele of the YieldPlus gene; and(c) optionally self-pollinating and / or backcrossing one or several times the plant selected at step b) and selecting in the progeny thus obtained a plant a mutant allele of the YieldPlus gene.

[0206] The self-pollination and backcrossing steps may be carried out in any order and can be intercalated, for example a backcross can be carried out before and after one or several self-pollinations, and self-pollinations can be performed before and after one or several backcrosses.

[0207] The selection of the progeny can advantageously be carried out with markers, e.g. the markers disclosed in the present specification. The selection of the progeny having the desired phenotype can also be made by assessing the phenotype of the progeny plants.

[0208] The inventors have been able to develop a genotyping assay to distinguish between the wild type allele and a mutant allele of the YieldPlus gene. Thus, is it now possible to detect the presence of a mutant allele of the YieldPlus gene.

[0209] Three nucleotide sequences were designed, two forward and one reverse primers (SEQ ID NO: 6, SEQ ID NO: 7 and SEQ ID NO: 8). These three sequences were designed based on the following sequences: SEQ ID NO: 1 or 2 (i.e., wild type allele) and SEQ ID NO:4 (i.e., mutant allele). These are sequences of the reverse strand (- strand) of the alleles.

[0210] Similar genotyping assay can be developed for any mutant allele of the YieldPlus gene and so are encompassed herein.

[0211] The invention is also directed to a method for detection and / or selecting a cucumber plant, seed or, plant part having an increased fruit yield, comprising the steps of:(a) Providing at least one genomic DNA sample of a cucumber plant, seed, or plant part,(b) Identifying a mutant allele of the YieldPlus gene on chromosome 3, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 ; and(c) Selecting a plant comprising said mutant allele.

[0212] In some embodiments, step (b) comprises carrying out a genotyping assay, using the DNA samples of a) as template, that discriminates between the wild type YieldPlus allele and the mutant allele, wherein said genotyping assay is based on nucleic acid amplification making use of oligonucleotide primers and probes.

[0213] In some embodiments, said genotyping assay uses at least one pair of oligonucleotide primers specific for the mutant YieldPlus allele. Said genotyping assay may also use at least one pair of oligonucleotide primers specific for the wild-type YieldPlus allele. In other embodiments, said genotyping assay is based on nucleic acid hybridization making use of YieldPlus allele-specific oligonucleotide probes.

[0214] In some embodiments, said oligonucleotide primers comprise at least 10 nucleotides of SEQ ID NO: 1 or SEQ ID NO: 2 or of the complement strand of SEQ ID NO: 1 or SEQ ID NO: 2.

[0215] In some embodiments, said plant selected at step (c) comprises one ortwo copies of said mutant allele.

[0216] In some embodiments, step a) comprises the isolation of genomic DNA from the plant, seeds, plant part, cell or tissue to be analyzed in the genotyping assay. Crude DNA extractions methods can be used as known in the art.

[0217] In some embodiments, the plant may be mutagenized using any mutagenesis technique as described herein, e.g. chemical radiation, mutagens or gene editing techniques. Thus, the method may comprise, prior to step a), a step of mutagenizing a Cucumis sativus plant to induce one or more mutations in the YieldPlus gene or regulatory sequence thereof.

[0218] In some embodiments, in step b) the genotyping assay is carrying out using the DNA samples of a) as template.

[0219] Various genotyping assays can be used, as long as they can detect the at least one mutation present in the mutant allele of the YieldPlus gene, e.g. an insertion, deletion or substitution, and can differentiate between the wild type allele of SEQ ID NO: 1 being present in the genomic DNA sample (at the YieldPlus locus on chromosome 3) or a mutant allele of the YieldPlus gene being present in the genomic DNA sample. Genotyping assays are generally based on allele-specific primers used in PCR or thermal cycling reactions (polymerase chain reaction) to amplify either the wild type or mutant allele and detect the amplification product or on allele-specific oligonucleotide probes, which hybridize to eitherthe wild type allele or the mutant allele, or both. For example, genotyping with BHQplus probes uses two allele specific probes and two primers that flank the region of the polymorphism, and during thermal cycling the polymerase encounters the allele-specific probes bound to the DNA and releases a fluorescent signal. Allele discrimination involves competitive binding of the two allele- specific BHQPIus probes (see also biosearchtech.com).

[0220] Examples of genotyping assays are the KASP-assay (by LGC, see at LGCgenomics.com and also at biosearchtech.com / products / pcr-kits-and-reagents / genotyping-assays / kasp-genotyping- chemistry), based on competitive allele-specific PCR and end-point fluorescent detection, the TaqMan- assay (Applied Biosytstems), which is also PCR based, HRM assays (High Resolution Melting Assay), wherein allele-specific probes are detected using real time PCR, or the rhAmp assay, based on Rnase H2-dependent PCR, BHQplus genotyping, BHQplex CoPrimer genotyping and many others.

[0221] The KASP-assay is also described in He C, Holme J, Anthony J. ‘SNP genotyping: the KASP assay. Methods Mol Biol. 2014;1145:75-86’ and EP1726664B1 or US7615620 B2, incorporated by reference. The KASP genotyping assay utilizes a unique form of competitive allele-specific PCR combined with a novel, homogeneous, fluorescence-based reporting system for the identification and measurement of genetic variation occurring at the nucleotide level to detect single nucleotide polymorphisms (SNPs) or inserts and deletions (InDeis). The KASP technology is suitable for use on a variety of equipment platforms and provides flexibility in terms of the number of SNPs and the number of samples able to be analyzed. The KASP chemistry functions equally well in 96-, 384-, and 1 ,536-well microtiter plate formats and has been utilized over many years in large and small laboratories by users across the fields of human, animal, and plant genetics. As recommended in the KASPar assay and illustrated in the examples, the amplification may be carried out by PCR cycles, comprising a first denaturation step at 94°C during around 15 minutes, at least 10 cycles of around 20 seconds at 94°C followed by around 60 second at a decreasing temperature from 65°C for the 1st cycle to 57°C for the last cycle, and around 35 cycles of around 20 seconds at 94°C followed by around 60 seconds at 57°C. This protocol can easily be adapted by a skilled person, depending on the type of primers used.

[0222] Various genotyping assays can, therefore, be used, which can differentiate between the presence of the wild type allele of the YieldPlus gene, encoding the protein of SEQ ID NO: 3, and a mutant allele of the YieldPlus gene.

[0223] As mentioned preferably a bi-allelic genotyping assay is used, e.g. a KASP-assay, a TaqMan assay, a BHQplus assay, PACE genotyping (see world wide web at idtdna.com / pages / products / qpcr- and-pcr / genotyping / pace-snp- genotyping-assays) or any other bi-allelic genotyping assay.

[0224] In one aspect the genotyping assay in step b) of the methods above is a KASP-assay. Thus, in step b) a competitive PCR is carried out using two forward primers and one common reverse primer. The two forward primers comprise at least 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19 or 20 nucleotides complementary to SEQ ID NO: 1 or SEQ ID NO:2 (or the complement strand thereof). In addition, the two forward primers comprise 1 , 2, 3 or more nucleotides (preferably at the 3’-end of the primers) which provide specificity to the SNP or INDEL which differentiates the wild type sequence from the mutantsequence of the allele. The two forward primers thereby have different binding specificity (or preference) to either the wild type allele or to the mutant allele. As illustrated herein with the oligonucleotide primers set forth in SEQ ID NO: 6, 7 and 8, the Fam- primer comprises 17 nucleotides of the wild type sequence and 1 nucleotide specific for the insertion allele, and the VIC -primer comprises 18 nucleotides of the wild type allele and 1 nucleotide specific to the ‘deletion’ allele. A KASP-assay can easily be designed to differentiate between the wild type allele of SEQ ID NO: 1 and any mutant allele of the YieldPlus gene which differs from the wild type allele in one or more nucleotides being inserted, deleted or substituted, so e.g. the assay can be designed for any SNP or INDEL that differentiates two alleles.

[0225] In a further aspect, the invention relates to a method for the production of C. sativus plantlets or plants, which method comprises:(a) culturing in vitro an isolated cell or tissue of the C. sativus plant according to the invention to produce C. sativus micro-plantlets, and(b) optionally further subjecting the C. sativus micro-plantlets to an in vivo culture phase to develop into C. sativus plants.

[0226] The isolated cell or tissue used to produce a micro-plantlet is an explant obtained under sterile conditions from a C. sativus parent plant of the invention to be propagated. The explant comprises or consists, for instance, of a cotyledon, hypocotyl, stem tissue, leaf, embryo, meristem, node bud, shoot apice, or protoplast. The explant can be surface sterilized before being placed on a culture medium for micropropagation. Conditions and culture media that can be suitably used in plant micropropagation are well known to those skilled in the art of plant cultivation and are described, for example, in "Plant Propagation by Tissue Culture, Handbook and Directory of Commercial Laboratories, eds. Edwin F George and Paul D Sherrington, Exegetics Ltd, 1984".

[0227] Micropropagation typically involves: axillary shoot production: axillary shoot proliferation is induced by adding cytokinin to the shoot culture medium, to produce shoots preferably with minimum callus formation; adventitious shoot production: addition of auxin to the medium induces root formation, in order to produce plantlets that are able to be transferred into the soil. Alternatively, root formation can be induced directly into the soil.

[0228] Plantlets can be further subjected to an in vivo culture phase, by culture into the soil under lab conditions, and then progressive adaptation to natural climate, to develop into C. sativus plants.

[0229] The invention is thus also directed to a method for improving the yield of C. sativus fruits and / or for increasing the number of harvestable C. sativus fruits comprising growing C. sativus according to the invention. The invention is also directed to the use of the C. sativus plants of the invention for improving the yield of C. sativus plants and / or fruits, and / or for increasing the number of harvestable cucumber fruits. The invention is also directed to a method of increasing the productivity of a C. sativus field, tunnel or glasshouse.

[0230] Preferably, the method comprises a first step of choosing or selecting a C. sativus plant comprising a mutant allele of the YieldPlus gene.

[0231] Preferably, the method comprises:(a) identifying C. sativus plants according to the invention, comprising a mutant allele of the YieldPlus gene, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield.(b) growing said resistant C. sativus plants in said infested environment.

[0232] By this method, the yield of cucumber production is increased, inter alia more marketable cucumbers can be harvested, and / or more seeds are obtained.

[0233] In still a further aspect, the invention relates to a method of producing cucumber fruits comprising:(a) growing a C. sativus plant of the invention;(b) allowing said plant to set fruit; and(c) harvesting fruit of said plant, preferably at maturity and / or before maturity.

[0234] All the preferred embodiments regarding the C. sativus plant are already disclosed in the context of the previous aspects of the invention.

[0235] The method may comprise a step of processing said cucumber fruit into a processed food and / or a step of mixing the cucumber fruits or part thereof with one or more food ingredients.

[0236] The present invention also relates to a method of producing a food product, comprising mixing a cucumber fruit of the invention, or part thereof, with one or more food ingredients. Optionally, the method comprises cooking and / or processing the cucumber fruit of the invention, alone or in mixture with the one or more food ingredients.

[0237] The present invention also relates to a food product made of a cucumber fruit of the invention or parts thereof, optionally in processed form.

[0238] In another aspect, the invention relates to the use of a C. sativus plant according to the invention or a fruit thereof in the fresh cut market or for food processing.

[0239] Throughout the instant application, the term “comprising” is to be interpreted as encompassing all specifically mentioned features as well optional, additional, unspecified ones. As used herein, the use of the term “comprising” also discloses the embodiment wherein no features other than the specifically mentioned features are present (i.e. “consisting of’).EXAMPLESExample 1 : Generation of an EMS population.

[0240] A cucumber (Cucumis sativus) mutant TILLING population was developed from a cucumber pickling line using an EMS (Ethyl methanesulfonate) treatment, in order to introduce random point mutations in the genome.

[0241] A screening analysis was then performed on this EMS population. One mutant was identified having a mutation in a gene named YieldPlus. More precisely, this mutant comprises a point mutation located in the coding sequence of the gene.

[0242] The YieldPlus gene encodes a zinc finger protein 8-like transcription factor. In the cucurbitgenomics.org database V2 genome, this gene is located on the chromosome 3 (negative strand) from 31 ,790,724 nucleotide to 31 ,792,269 nucleotide (Cucumis. sativus L. var. sativus var. 9930 v3; http: / / cucurbitgenomics.org / feature / gene / CsaV3_3G038590).

[0243] Table 1 : Description of the mutant plant generated. Nucleotide mutated position refers to the nucleotide number in the corresponding sequence number, i.e., SEQ ID NO. The genome position mentioned with respect to the cucumber as published by Li et al, 2019 A chromosome-scale genome assembly of cucumber (Cucumis sativus L.). Gigascience, Volume 8, Issue 6, June 2019, giz072).

[0244] Table 1 : mutations of the generated mutant

[0245] A KASP marker-assay was developed to distinguish between the wild type allele of the gene, shown in SEQ ID NO: 1 , and the mutant allele of the YieldPlus gene comprising a mutation, in the present case a substitution, shown in SEQ ID NO: 4.

[0246] Table 2: markers and primers for mutation detection on minus DNA strand

[0247] It is noted that the DNA sequences for the KASP assay were designed on the reverse DNA strand (minus strand) but can be equally designed based on the plus strand of the allele. Plus and minus strands are complementary strands of the double stranded DNA. Nucleotide G corresponds to a C in the complementary strand and nucleotide A corresponds to a T in the complementary strand.Example 2: Introgression of the mutant allele

[0248] The introgression of the mutant allele (SEQ ID NO: 4) into a wild type Cucumber (i.e., not bearing SEQ ID NO: 4) is achieved by crossing and genotypic screening, using the markers developed (SEQ ID NO: 6 to 8) to specifically follow the mutant alleles.

[0249] The family that was identified(i.e., mutant family) was selfed to get cucumber plants bearing the mutant allele homozygously (MutantHOM).

[0250] These homozygote plants were used as trait donor for the introgression into a wild type cucumber (Digitus, a Beit-Alpha parthenocarpy cucumber). Two backcrosses were done with Digitus as a recurrent parent (used as female) until BC2F2 stage. Hence a BC2F2 population issued from crossing MutantHOM with Digitus has been created.

[0251] The fruit yield of cucumber plants bearing homozygously the mutant allele, i.e., SEQ ID NO: 4 and wild type cucumber plants homozygous forthe wild-type allele (i.e., not bearing SEQ ID NO: 4) were compared. To this end, parthenocarpic fruits were harvested 3 times a week for three and a half weeks (10 harvests).

[0252] The BC2F2 conversions of the mutant family show a significant difference in fruit weight per plant (Figure 1A) and fruit number per plant (Figure 1 B) between the homozygote conversion (A:A) and the one that does not have the mutation (G:G). The mutant allele of the YieldPlus gene (SEQ ID NO: 4) is responsible forthe increased yield of the cucumber plants comprising the mutant allele in homozygous form.

[0253] Additionally, the BC2F2 conversions of the mutant family do show a difference below 10%, even below 5%, in average individual fruit weight between the plants with the homozygote conversion (A:A) and the ones that do not have the mutation (G:G). Plants having the mutant allele demonstrate an average individual fruit weight of 136g whereas plants having the wild-type allele demonstrate an average individual fruit weight of 142g.

[0254] The point mutation in the mutant corresponds to a substitution of a G by a A at position 757 of SEQ ID NO: 2. This substitution corresponds to a change of a glycine into a serine in position 253 of the protein sequence (SEQ ID NO:3). This amino acid residue is located in a C-terminal disordered portion of the protein and not in the zinc finger domain. This mutation is predicted to have a neutral impact using the Provean (1.528) and SIFT (0.551) scores.Example 3: Introduction of mutations by genome editing

[0255] Cas9 / sgRNA constructs were designed to target sequences in the coding sequence of the YieldPlus gene (SEQ ID NO: 1) or in 3’-UTR or 5’-UTR sequences.

[0256] Independent TO transgenic lines were generated by Agrobacterium-mediated transformation. The presence of the transgene (Cas9 / sgRNA) was confirmed by kanamycin resistance and PCR using sgRNA specific primers. To evaluate the types of mutations generated in sgRNA transgenic plants, PCRwas performed in TO plants using primers flanking the sgRNA target and subsequently digested with a restriction enzyme, at a site that will disappear if CAS9 and NHEJ were active in this site. A distinct undigested fragment is observed in mutant plants following restriction. A partial digestion observed indicates a heterozygous genome with both wild-type and mutant alleles. The uncut digested fragment is cloned and sequence to identify the mutations.

[0257] For propagation by seeds, the Cucumis sativus TO-mutant plant are cross-pollinated with a wild type cucumber (Beit-Alpha parthenocarpy cucumber). The mutant alleles are followed by molecular marking, evidencing stable transmission of the mutation. The T1 mutant plants are grown and crosspollinated to produce mutant plants, with a homozygous mutation in the YieldPlus gene. T2 and T3 plants are further generated.

[0258] The edited plants are evaluated for their fruit yield phenotype. Plants showing increased fruit yield with respect to wild-type control plants are selected for further introgression of the edited allele in cucumber genotypes of agricultural interest.Example 4: Effect of the mutation on the number of fruits per node

[0259] To investigate mechanisms responsible for increased yield, the number of fruits per node was measured. This mechanism can be observed by recording the commercial-sized fruits on different nodes, and compared to a reference line which does not comprise the mutant allele, e.g. an isogenic wild-type line. In various cucumber types / lines and hybrids, some have one fruit per node, while others have multiple fruits per node. For a given genotype, having additional fruits per node corresponds to an increase in yield, as assessed by the measurements carried out on the cucumbers plants of the invention.Example 5: Effect of the mutation on fast fruit filling

[0260] Another mechanism investigated as possibly responsible for increased yield is faster fruit filling. When a fruit reaches commercial size and is picked, it contributes to increased yield. To measure fast fruit filling, the time elapsed from anthesis to harvest at a commercial stage is measured. Specifically, the time elapsed from anthesis to harvest at a commercial stage is measured in a mutant line according to the invention, as compared to a reference line which does not comprise the mutant allele, e.g. an isogenic wild-type line. In a favorable case, the line with the favorable allele, e.g. the mutant allele, has a decreased time from anthesis to harvest, and has more fruits at a commercial stage vs. the line with the non-favorite allele.

Claims

CLAIMS

1. A Cucumis sativus plant, or a seed, cell or part thereof, wherein said plant, seed, cell or part comprises a mutant allele of a YieldPlus gene on chromosome 3 in its genome, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

2. A plant, seed, cell or part according to claim 1 , wherein said mutant allele is present homozygously in the genome of said plant.

3. A plant according to claim 1 or 2, wherein said mutant allele comprises a nucleotide sequence having at least 80% identity with the sequence set forth in SEQ ID NO: 1 or SEQ ID NO: 2.

4. A plant, seed, cell or part according to claim 1 or 2, wherein said mutation is a loss-of- function mutation and / or a mutation which decreases expression of the YieldPlus protein.

5. A plant according to any one of claims 1 to 4 wherein said at least one loss-of-function mutation is selected from a missense mutation, a nonsense mutation and a frameshift mutation.

6. The plant according to any one of claims 1 to 5, wherein said at least one mutation results in at least one amino acid change, addition or deletion in the amino acid sequence of the protein encoded by the YieldPlus gene, as set forth in SEQ ID NO:3.

7. A plant, seed, cell or part according to any one of claims 1 to 6, wherein said at least one mutation results in at least one amino acid change, addition or deletion in the C-terminal portion of the protein encoded by the YieldPlus gene, preferably between the amino acid residues in position 150 and 277 of SEQ ID NO:3.

8. A plant, seed, cell or part according to any one of claims 1 to 7, wherein said at least one mutation results in at least one amino acid change, addition or deletion in position 253 of the amino acid sequence of the protein encoded by the YieldPlus gene, as set forth in SEQ ID NO:3.

9. A plant, seed, cell or part according to any one of claims 1 to 8, wherein said at least one mutation results in a G253S change in the amino acid sequence of the protein encoded by the YieldPlus gene, as set forth in SEQ ID NO:3.

10. A plant, seed, cell or part according to any one of claims 1 to 9, wherein said at least one mutation is a G757A mutation in the coding sequence of the Yieldplus gene, as set forth in SEQ ID NO: 2.

11. A plant, seed, cell or part according to any one of claims 1 to 10, wherein said plant produces at least 10% more fruits and / or has a fruit weight at least 10% higher in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene.

12. A plant, seed, cell or part according to any one of claims 1 to 11 , wherein the average individual fruit weight of fruits of said plant is decreased by less than 50%, in particular less than 10%,or is not decreased, in comparison to the average individual fruit weight of fruits of an isogenic plant which comprises a wild-type allele of the YieldPlus gene.

13. A plant, seed, cell or part according to any one of claims 1 to 12, wherein said plant is a plant from an inbred line or is a hybrid plant.

14. A plant, seed, cell or part according to any one of claims 1 to 13, wherein said plant further comprises one or more traits of agronomical interest selected from resistance to ZYMV (Zucchini Yellow Mosaic Virus), resistance to CVYV (Cucumber Vein Yellowing Virus), resistance to PRSV (Papaya Ringspot Virus), resistance to WMV (Watermelon Mosaic Virus), resistance to CMV (Cucumber Mosaic Virus), resistance to powdery mildew, resistance to potyviruses, resistance to downy mildew, e.g. caused by Pseudoperonospora cubensis, resistance to Fusaria, e.g. caused by Fusariumoxysporum f.sp. cucumerinum or by Fusarium oxysporum f.sp. radicis cucumerinum, resistance to scab, e.g. caused by Cladosporium cucumerinum, resistance to CYSDV, resistance to angular leaf sport and resistance to anthracnose.

15. A seed for producing a plant according to any one of claims 1 to 14.

16. An isolated polynucleotide, comprising a mutant allele of the YieldPlus gene, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , wherein said mutant allele confers increased fruit yield to a plant comprising said mutant allele, in comparison to an isogenic plant which comprises a wild-type allele of the YieldPlus gene.

17. The isolated polynucleotide of claim 16, which comprises a nucleotide sequence having at least 80% identity with SEQ ID NO: 1 or SEQ ID NO: 2.

18. The isolated polynucleotide of claim 16 or 17, which encodes a polypeptide with at least 98% identity with SEQ ID NO:3.

19. An isolated polypeptide, comprising an amino acid sequence with at least 98% identity with SEQ ID NO:3, wherein said polypeptide comprises at least one mutation in the amino acid sequence, in comparison to the sequence of the corresponding wild-type polypeptide as set forth in SEQ ID NO:3, wherein said mutation confers a loss-of-function phenotype to said polypeptide.

20. An in vitro cell or tissue culture of regenerable cells of the plant according to any one of claims 1 to 7, wherein the regenerable cells are derived from an embryo, protoplast, meristematic cells, callus, pollen, leaf, anther, stem, petiole, root, root tip, seed, flower, cotyledon, and / or hypocotyl.

21. A method of producing a plant, comprising: a) obtaining a part of a plant according to any one of claims 1 to 14, b) vegetatively propagating said plant part to generate a plant from said plant part.

22. A method of producing a C. sativus plant, comprising the introduction of at least one mutation in the YieldPlus gene on chromosome 3 in the genome of a C. sativus plant, wherein said mutation is introduced in the sequence of the gene or in a regulatory sequence thereof, in comparisonto the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

23. The method of claim 22, wherein said mutation is introduced by mutagenesis or genome editing, in particular by a technique selected from ethyl methanesulfonate (EMS) mutagenesis, oligonucleotide directed mutagenesis (ODM), Zinc finger nuclease (ZFN) technology, Transcription Activator-Like Effector Nucleases (TALENs) the CRISPR / Cas system, the CRISPR / Cpf system engineered meganuclease, re-engineered homing endonucleases and DNA guided genome editing.

24. The method of claim 22 or 23, comprising:(a) Introducing one or more mutations in cucumber plant(s), seed(s) or plant part(s),(b) Optionally, determining if the plant, seed or plant part under a) presents an increased yield compared to a plant not having said at least one mutation; and(c) Selecting a plant that comprises a mutant allele of a YieldPlus gene, wherein said mutant allele comprises at least one mutation in the sequence of the gene or in a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 , resulting in an increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

25. A method of producing a C. sativus plant, comprising:(a) crossing a C. sativus plant according to any one of claims 1 to 14 with itself or with a second C. sativus plant, preferably of a different genotype, to produce one or more progeny plants;(b) selecting a progeny plant comprising a mutant allele of the YieldPlus gene; and(c) optionally self-pollinating and / or backcrossing one or several times the plant selected at step b) and selecting in the progeny thus obtained a plant comprising a mutant allele of the YieldPlus gene.

26. A method for detecting and / or selecting a cucumber plant, seed or plant part, comprising the steps of:(a) providing at least one genomic DNA sample of a cucumber plant, seed, or plant part,(b) identifying a mutant allele of the YieldPlus gene on chromosome 3, wherein said mutant allele comprises at least one mutation in the sequence of the gene or a regulatory sequence thereof, in comparison to the sequence of the corresponding wild-type allele as set forth in SEQ ID NO: 1 ; and(c) selecting a plant comprising said mutant allele, wherein the selected plant has increased fruit yield in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene.

27. The method of claim 26, wherein step (b) comprises carrying out a genotyping assay, using the DNA samples of a) as template, that discriminates between the wild type YieldPlus allele and the mutant allele, wherein said genotyping assay is based on nucleic acid amplification making use of YieldPlus allele specific oligonucleotide primers, and / or wherein said genotyping assay is based on nucleic acid hybridization making use of YieldPlus allele-specific oligonucleotide probes, preferably wherein said YieldPlus allele specific oligonucleotide primers or said YieldPlus allele-specificoligonucleotide probes comprise at least 10 nucleotides of SEQ ID NO: 1 or of the complement strand of SEQ ID NO: 1 .

28. The method of claim 27, wherein said YieldPlus allele specific oligonucleotide primers or said YieldPlus allele-specific oligonucleotide probes comprise at least 10 nucleotides of SEQ ID NO: 1 or SEQ ID NO: 2 or of the complement strand of SEQ ID NO: 1 or SEQ ID NO: 2.

29. The method of claim 27 or 28, wherein said YieldPlus allele specific oligonucleotide primers comprise a sequence selected from the sequences set forth in SEQ ID NO: 6 and SEQ ID NO: 7.

30. A method for screening and / or selecting plant, seeds or plant part, or DNA or RNA or protein derived therefrom, for the presence of a mutant allele of the YieldPlus gene on chromosome 3, conferring increased fruit yield to a plant comprising said mutant allele in comparison to an isogenic plant comprising a wild-type allele of the YieldPlus gene, said method comprising one or more of the following steps:(a) determining if the level of expression of the YieldPlus gene in the plant, seed or plant part is reduced or abolished in comparison to a plant comprising a wild-type allele of the YieldPlus gene, wherein the sequence of the wild-type YieldPlus gene is set forth in SEQ ID NO: 1 ;(b) determining if the amounts of the protein encoded by the YieldPlus gene in the plant, seed or plant part thereof is reduced or abolished in comparison to a plant comprising a wild-type allele of the YieldPlus gene, wherein the sequence of a wild-type allele of the YieldPlus gene is set forth in SEQ ID NO: 1 ;(c) determining the presence of at least one mutation in the YieldPlus gene or a regulatory sequence thereof, in nucleic acid sample, preferably a mRNA, cDNA and / or genomic DNA sample, from the plant, seed or plant part in comparison to the wild-type YieldPlus gene having the sequence set forth in SEQ ID NO: 1 ;(d) determining the presence of at least one mutation in the YieldPlus protein from the plant, seed or plant part, in comparison to the wild-type YieldPlus protein of SEQ ID NO:3.

31. A method of producing cucumber fruit, comprising:(a) growing a C. sativus plant according to any one of claims 1-14.(b) allowing said plant to set fruit; and(c) harvesting fruit of said plant.

32. A method of producing a foodstuff or feedstuff, comprising:(a) obtaining fruits of a C. sativus plant according to any one of claims 1-14; and(b) processing said fruits into a processed food or feedstuff or using said fruit as an ingredient into a foodstuff or feedstuff.

33. Use of a C. sativus plant according to any one of claims 1-14 or some fruit thereof in the fresh cut market or for food processing.