Polyploidization of interspecific tomato hybrids to create stable and fertile rootstocks.

Abstract

This disclosure relates to alloploidy plants and hybrid alloploidy plants having desirable properties such as resistance to abiotic or biological stressors, which can be used as rootstock for cultivated tomato varieties. This disclosure further relates to composite plants that include the alloploidy plants described herein as rootstock. This disclosure further relates to methods for producing alloploidy plants and plant parts, as well as hybrid alloploidy plants and plant parts. In some embodiments, the techniques described herein relate to methods for producing unnatural stress-tolerant composite tomato plants, comprising providing a stress-tolerant allotetraploid tomato plant as rootstock, providing a Solanum lycopersicum variety as scion, grafting the scion onto the rootstock, thereby producing an unnatural stress-tolerant composite tomato plant.

Description

Technical Field

[0001] Cross - reference to related applications This application claims the benefit of International Patent Application No. PCT / EP2023 / 057886, filed on March 27, 2023, and the entire content thereof is incorporated herein by reference for all purposes.

[0002] The present disclosure relates to the fields of agriculture, plant biotechnology, and molecular biology. More specifically, the present disclosure relates to allopolyploid plants, methods for producing allopolyploid plants having desirable traits, and methods of using them as a scion of tomato cultivars.

Background Art

[0003] For farmers, it is becoming increasingly difficult to meet the expanding global food demand. Although plants can be improved by conventional breeding methods and / or transgenic technologies, progress can be blocked and / or hindered by interspecific incompatibility, complex multifactorial and polygenic traits, and restrictions on genetically modified crops. One way to confer the advantages of desirable traits without using breeding or genetic modification is to graft two plants onto each other to form a composite plant.

[0004] Grafting has historically been used for grapes and trees, but is becoming more popular for other crops such as tomatoes. One advantage of grafting is that it can confer increased tolerance to abiotic stresses, such as drought tolerance, salt tolerance, waterlogging / water tolerance, heat and cold tolerance, etc., on cultivars by using a scion with desirable traits. In addition, since roots supply important nutrients to plants, the scion can further affect plant performance and yield. Therefore, there is a need for new scions with desirable traits.

Summary of the Invention

[0005] The following embodiments and their aspects are described together with the systems, tools, and methods intended to be illustrative, but are not intended to limit their scope.

[0006] In some embodiments, the technique described herein relates to a method for producing unnatural stress-tolerant complex tomato plants, comprising providing a stress-tolerant allotetraploid tomato plant as rootstock, a Solanum lycopersicum variety as scion, grafting the scion onto the rootstock, thereby producing unnatural stress-tolerant complex tomato plants. In some embodiments, the rootstock contains chromosomes derived from Solanum lycopersicum and chromosomes derived from at least one species selected from the Esculentum, Arcanum, Pervianum, Hirsutum, Lycopersicoides sections, and hybrid combinations thereof. In some cases, the scion is a commercially available variety.

[0007] In some embodiments, the technique described herein relates to a method for producing stress-tolerant allotetraploid plants, which involves crossing a Solanum lycopersicum variety with a stress-tolerant Solanaceae species to produce interspecific hybrid seeds (in which case the Solanum lycopersicum variety and the stress-tolerant Solanaceae species are essentially homozygous, and the stress-tolerant Solanaceae species are selected from the Esculentum group, Arcanum group, Peruvianum group, Hirsutum group, Lycopersicoides section, and hybrid combinations thereof), cultivating the interspecific hybrid seeds to produce interspecific hybrid plants, spraying the interspecific hybrid plants or their cuttings with a chromosome doubling agent to generate chimeric interspecific hybrids, cultivating the chimeric interspecific hybrids to produce tomato fruits, collecting seeds from the tomato fruits, and cultivating the seeds to produce stress-tolerant allotetraploid tomato plants.

[0008] In some embodiments, the technique described herein relates to a method for producing stress-tolerant allotetraploid tomato plants by fusing protoplasts isolated from Solanum lycopersicum with another protoplast isolated from a stress-tolerant Solanaceae species, isolating heterokaryon, and regenerating allotetraploid tomato plants from the heterokaryon.

[0009] In some embodiments, the techniques described herein relate to a method for selecting stress-tolerant Solanaceae species from the Esculentum group, Arcanum group, Peruvianum group, Hirsutum group, Lycopersicoides section, and hybrid combinations thereof.

[0010] In some embodiments, the techniques described herein relate to a method in which stress-tolerant Solanaceae species are derived from the Esculentum group and selected from S. galapagense, S. cheesemaniae, S. lycopersicum, S. pimpinellifolium, and hybrid combinations thereof.

[0011] In some embodiments, the techniques described herein relate to a method for selecting stress-tolerant Solanaceae species from the Arcanum group, specifically from S. neorickii, S. arcanum, S. chmielewskii, and hybrid combinations thereof.

[0012] In some embodiments, the techniques described herein relate to a method by which stress-tolerant Solanaceae species are selected from the Peruvianum group, specifically from S. huaylasense, S. peruvianum, S. corneliomulleri, S. chilense, and hybrid combinations thereof.

[0013] In some embodiments, the techniques described herein relate to a method for selecting stress-tolerant Solanaceae species from the Hirsutum group, specifically from S. habrochaites, S. pennellii, and hybrid combinations thereof.

[0014] In some embodiments, the techniques described herein relate to a method by which stress-tolerant Solanaceae species are derived from the Lycopersicoides section and selected from S. lycopersicoides, S. sitiens, and hybrid combinations thereof.

[0015] In some embodiments, the techniques described herein relate to methods for enabling stress-tolerant Solanaceae species to possess abiotic stress tolerance selected from the group consisting of cold tolerance, heat tolerance, drought tolerance, and salt tolerance.

[0016] In some embodiments, the techniques described herein relate to methods for enabling stress-tolerant Solanaceae species to possess biological stress tolerance selected from the group consisting of disease resistance, fungal resistance, pest and disease resistance, bacterial resistance, insect resistance, and nematode resistance.

[0017] In some embodiments, the techniques described herein relate to a method in which the protoplast fusion is asymmetric and the protoplast of S. lycopersicum is the acceptor.

[0018] In some aspects, this disclosure relates to non-natural compound tomato plants produced by the methods disclosed herein.

[0019] In some embodiments, the techniques described herein relate to non-natural compound tomato plants, wherein the heterotetraploid rootstock contains at least one allele derived from a scion variety of Solanum lycopersicum.

[0020] In some embodiments, the techniques described herein relate to a method for producing hybrid allotetraploid tomato seeds for producing hybrid allotetraploid plants, the method comprising the steps of crossing a first stress-tolerant allotetraploid tomato plant with a second stress-tolerant allotetraploid tomato plant (the second stress-tolerant allotetraploid containing chromosomes derived from at least one species different from the first allotetraploid tomato plant), and collecting hybrid allotetraploid tomato seeds (the first stress-tolerant allotetraploid tomato plant and the second stress-tolerant allotetraploid tomato plant are essentially homozygous). In some embodiments, the first or second stress-tolerant allotetraploid tomato plant comprises chromosomes derived from Solanum lycopersicum, as well as at least one species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, S. habrochaites, and their hybrids. In some embodiments, the first allotetraploid tomato plant provides at least one tolerance to at least one stressor that is not provided by the second allotetraploid tomato plant, and the stress tolerances provided by the first allotetraploid plant and the second allotetraploid plant are complementary. In some embodiments, the first stress-tolerant allotetraploid tomato plant has abiotic stress tolerance selected from the group consisting of cold tolerance, heat tolerance, drought tolerance, and salt tolerance. In some embodiments, the second stress-tolerant allotetraploid tomato plant has abiotic stress tolerance selected from the group consisting of disease tolerance, fungal tolerance, pest and disease tolerance, bacterial tolerance, insect tolerance, and nematode tolerance.

[0021] In some embodiments, the disclosure relates to hybrid allotetraploid tomato plants produced by cultivating the hybrid allotetraploid tomato seeds described herein.

[0022] In some embodiments, the disclosure relates to a non-natural compound tomato plant comprising a scion of Solanum lycopersicum grafted onto a hybrid allotetraploid rootstock produced by the method described herein. In some embodiments, the hybrid allotetraploid rootstock comprises an allele derived from at least one scion variety of Solanum lycopersicum.

[0023] In some embodiments, the disclosure relates to a chimeric plant tissue comprising a first plant cell and a second plant cell, wherein the first plant cell is an allotetraploid comprising chromosomes derived from Solanum lycopersicum, as well as chromosomes derived from at least one species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the second plant cell is a diploid Solanum lycopersicum.

[0024] In some embodiments, the disclosure relates to a hybrid allotetraploid tomato plant comprising chromosomes derived from Solanum lycopersicum, as well as chromosomes derived from at least one species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, S. habrochaites, and their hybrids, wherein the hybrid allotetraploid tomato plant provides at least one tolerance to at least one stressor that is not provided by one of its parent lines. [Brief explanation of the drawing]

[0025] [Figure 1] This is a flowchart illustrating the steps for generating heterotetraploid tomato plants.

[0026] [Figure 2] It is a flowchart showing the steps of generating an allopolyploid hybrid and using the obtained F1 as a scion of a commercial variety.

[0027] [Figure 3] The relationships among all wild tomato species are shown based on information from approximately half of the known genes in each of these species (in this case, more than 16,500 genes, and tomatoes have a total of approximately 34,000 genes each). The left side is a tree constructed from the average interspecific DNA differences of all these genes. The right side is a diagram showing many individual trees constructed from smaller groups of these genes. These indicate that the evolutionary emergence of current wild species is recent and complex. The genetic distance was calculated as the total transcriptome sequence divergence (%) between different species, and the maximum threshold was 3% (Pose et. al. 2016) Phylogenomics Reveals Three Sources of Adaptive Variation during a Rapid Radiation. PLoS Biol 14(2)). The phylogenetic species of the Lycopersicon section have a sequence divergence of 2.10% - 2.71% compared to the phylogenetic species of Lycopersicoides. The phylogenetic species within Lycopersicon have pairwise distances of 0.05% - 1.7%, and are the closest among different phylogenetic species within S. galapagense (gal - 3909 / gal - 0436) and different phylogenetic species within cultivated tomatoes (lyc - 3475 / lyc - ref).

[0028] [Figure 4] Fruits in plants of the allopolyploid variety "MX20 - 06" are shown.

[0029] [Figure 5] The whole harvested fruit of the allopolyploid variety "MX20 - 06" and a longitudinal section of the harvested fruit are shown.

[0030] [Figure 6] This shows the flowers of the heterotetraploid variety "MX20-06" in plants.

[0031] [Figure 7] This image shows a magnified view of a flower isolated from the heterotetraploid variety "MX20-06".

[0032] [Figure 8] The image shows stems and leaves excised from the heterotetraploid variety "MX20-06".

[0033] [Figure 9] This plot shows the relative nuclear DNA content of diploid (A) and heterotetraploid (B) strains, measured by flow cytometry. The x-axis represents propidium iodide (PI) fluorescence (i.e., relative DNA content), and the y-axis represents the number of nuclei measured (×100). An increase from 50 to 100 indicates diploidy.

[0034] [Figure 10] This shows the size difference between diploid seeds (left) and allotetraploid seeds (right).

[0035] [Figure 11] This shows the size difference between a diploid seedling (left) and an allotetraploid seedling (right).

[0036] [Figure 12] This shows the difference in shape between a diploid leaf (left) and an allotetraploid leaf (right).

[0037] [Figure 13] This shows the morphological differences between diploid roots (A, B) and allotetraploid roots (C, D).

[0038] [Figure 14A]This is a box plot showing the grams of harvested fruit per plant for Sugarino-MX20-06 and Sugarino-Maxifort composite plants grown in the experimental greenhouse facility of Red Sea Farms at King Abdullah University of Science and Technology, Thuwal, Saudi Arabia. [Figure 14B] This bar graph shows the yield (kilograms / plant) of ungrafted 'Midelyce', Midelyce-Maxifort compound plants, Midelyce-MX20-15 compound plants, and Midelyce-MX20-06 compound plants cultivated in a field in Saudi Arabia. The plants were irrigated with 6% seawater (EC=6.22 mS / cm) and exposed to temperatures up to 42°C.

[0039] [Figure 15] This bar graph shows the yield (kilograms / plant) of ungrafted 'Midelyce', Midelyce-Maxifort composite plants, and Midelyce-MX20-15 composite plants cultivated in a greenhouse trial in Saudi Arabia. 'Salt' treated plants were irrigated with 6% seawater (EC=6.22 mS / cm).

[0040] [Figure 16A] The images show field trials of ungrafted "CH7", the CH7-MX20-06 compound plant, and CH7-MX20-15, transplanted on September 10, 2022, and cultivated near Al-Dabaa Road in Egypt. The field is shown 74 days after transplanting, under vegetative growth conditions. [Figure 16B] The images show field trials of ungrafted 'CH7', the CH7-MX20-06 compound plant, and CH7-MX20-15, transplanted on September 10, 2022, and cultivated near Al-Dabaa Road in Egypt. The field is shown 178 days after transplanting, before the first harvest. [Figure 16C]The images show field trials of ungrafted "CH7", the CH7-MX20-06 compound plant, and CH7-MX20-15, transplanted on September 10, 2022, and cultivated near Al-Dabaa Road in Egypt. The field is shown 244 days after transplanting, following the final harvest.

[0041] [Figure 17] This line graph shows the yield results (kilograms / plant) over four harvest periods (H1-H4) for ungrafted "CH7" plants, CH7-MX20-06 composite plants, and CH7-MX20-15 composite plants in a field trial conducted near Al-Dabaa Road in Egypt.

[0042] [Figure 18] This line graph shows the average fruit weight (kilograms / fruit) over four harvest seasons (H1-H4) for ungrafted "CH7" plants, CH7-MX20-06 composite plants, and CH7-MX20-15 composite plants in a field trial conducted near Al-Dabaa Road in Egypt.

[0043] [Figure 19] This image shows a CH7-MX20-06 compound plant uprooted from a field near Al-Dabaa Road in Egypt.

[0044] [Figure 20A] These are photographs of field trials conducted in Stockton, California, of plants grafted onto tetraploid rootstocks 'MX20-06' and 'Maxifort'. The commercially available variety 'UG161-12' was used as the scion. The transplanting date is indicated. [Figure 20B] This image shows a field trial conducted in Stockton, California, involving plants grafted onto tetraploid rootstocks 'MX20-06' and 'Maxifort'. The commercially available variety 'UG161-12' was used as the scion. The image shows the plants 32 days after transplanting. [Figure 20C]The images show field trials conducted in Stockton, California, of plants grafted onto tetraploid rootstocks 'MX20-06' and 'Maxifort'. The commercially available variety 'UG161-12' was used as the scion. The images show the plants 46 days after transplanting. [Figure 20D] This photograph shows field trials conducted in Stockton, California, of plants grafted onto tetraploid rootstocks 'MX20-06' and 'Maxifort'. The commercially available variety 'UG161-12' was used as the scion. The image shows the plants 52 days after transplanting. [Figure 20E] The images show field trials conducted in Stockton, California, of plants grafted onto tetraploid rootstocks 'MX20-06' and 'Maxifort'. The commercially available variety 'UG161-12' was used as the scion. The images show the plants 59 days after transplanting. [Figure 20F] This photograph shows field trials conducted in Stockton, California, of plants grafted onto tetraploid rootstocks 'MX20-06' and 'Maxifort'. The commercially available variety 'UG161-12' was used as the scion. The image shows the plants 84 days after transplanting.

[0045] [Figure 21] This bar graph shows the yield (tons / hectare) of ungrafted "UG161-12" plants, UG161-12-Maxifort composite plants, and UG161-12-MX20-06 composite plants grown at standard spacing and sparse planting in a field trial in Stockton, California.

[0046] [Figure 22] "MX20-15" shows the stem, leaves, flowers, and fruits of the plant.

[0047] [Figure 23] A magnified view of the "MX20-15" bundle is shown.

[0048] [Figure 24] This shows a magnified view of the upper part of the "MX20-15" fruit.

[0049] [Figure 25] This shows a longitudinal section of a harvested fruit of "MX20-15".

[0050] [Figure 26] This shows stems and leaves removed from the variety "MX20-15". [Modes for carrying out the invention]

[0051] definition The following terms are expected to be easily understood by those skilled in the art, but their definitions are provided below to facilitate the explanation of the subject matter disclosed herein.

[0052] In accordance with long-standing patent law practice, the terms “a,” “an,” and “the,” when used in this application, including in the claims, mean “one or more.” For example, the phrase “a cell” means one or more cells, and in some embodiments, it may mean tissue and / or organs. Similarly, the phrase “at least one,” when used herein to refer to an entity, means an entity including, but not limited to, all integer values ​​between 1 and 100, and integers greater than 100, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 75, 100, or more.

[0053] Unless otherwise indicated, all numbers used herein and in the claims to represent quantities of components, reaction conditions, etc., should be understood in all cases as being modified by the term “approximately.” As used herein with respect to measurable values ​​such as mass, weight, time, volume, concentration, or percentage, “approximately” means to include variations from a particular amount of ±20% in some embodiments, ±10% in some embodiments, ±5% in some embodiments, ±1% in some embodiments, ±0.5% in some embodiments, and ±0.1% in some embodiments, as appropriate for carrying out the disclosed method and / or using the disclosed composition, nucleic acid, polypeptide, etc. Therefore, unless otherwise indicated, the numerical parameters described herein and in the appended claims are approximations and may vary depending on the desired properties sought by the subject matter of this disclosure.

[0054] As used herein, the term “and / or” refers to entities that exist individually or in combination when used in the context of a list of entities. Thus, for example, the phrase “A, B, C, and / or D” includes not only A, B, C, and D individually, but also any combination and subcombinations of A, B, C, and D (e.g., AB, AC, AD, BC, BD, CD, ABC, ABD, and BCD). In some embodiments, one or more of the elements referred to by “and / or” may also exist individually, in single or multiple combinations and / or subcombinations.

[0055] The term "heteropolyploidy" refers to cells or plants that have two or more complete sets of chromosomes derived from different species.

[0056] The term "allotetraploid" refers to a hybrid cell or plant that originates from a different species and has four times the number of chromosomes as a haploid organism. For example, allotetraploids can be produced through interspecific hybridization followed by chromosome duplication. In some cases, allotetraploids can exhibit some degree of aneuploidy. For example, in a 2n=24 tomato, an allotetraploid may have a chromosome number ranging from 44 to 52.

[0057] As used herein, the term “aneuploid” refers to a cell or plant having an incomplete set of chromosomes. Aneuploids may, for example, have missing or extra chromosomes.

[0058] As used herein, “antimitotic” or “antimitotic agent” refers to a compound or chemical used to inhibit cell proliferation by halting mitosis (cell division), and is used in plant breeding to induce chromosome duplication. Examples of antimitotic agents include, but are not limited to, colchicine, trifluralin, oryzalin, and amiprophos-methyl (APM).

[0059] As used herein, the terms “at least a portion” or “fragment” of a nucleic acid or polypeptide mean the portion having the minimum size characteristics of such sequence, or any larger fragment of a full-length molecule less than or equal to the full-length molecule.

[0060] A "chimera," "chimeric tissue," or "chimeric plant" is a plant or tissue consisting of two or more genetically distinct groups of cells.

[0061] As used herein, the term "cisgenesis" refers to the genetic modification of a recipient organism with one or more genes (cis genes) derived from a mating and sexually compatible organism.

[0062] As used herein, “compound” or “compound plant” refers to a plant formed by grafting (rootstock + scion) two separate varieties into a single plant.

[0063] Colchicine is a pale yellow alkaloid obtained from autumn crocus, i.e., C 22 H 25 It is NO6 and is used in plant breeding to induce chromosome duplication.

[0064] "Finite-growth tomatoes" are tomato varieties that produce fruit all at once and then stop producing fruit. Because they can be harvested all at once, they are ideal for commercial cultivation and mechanical harvesting.

[0065] As used herein, the term “improved abiotic stress tolerance” refers to the ability of a plant or plant part to grow, reproduce, and / or survive under abiotic stress conditions compared to one or more controls (plants that are not stress-tolerant). "Improved abiotic stress tolerance" may refer to any improvement in the growth capacity and / or tolerance capacity of a plant or plant part grown under abiotic stress conditions, or to the ability of a plant to maintain growth and yield under abiotic stress conditions, including, but not limited to, reduced water loss, reduced accumulation of one or more reactive oxygen species, reduced accumulation of one or more salts, increased salt excretion, increased accumulation of one or more dehydrins, improved root structure, improved osmoregulation, increased accumulation of one or more proteins abundant in late embryonic development, improved viability, improved growth rate, increased plant height, increased chlorophyll content, improved fruit quality, and / or increased yield (e.g., increased biomass, increased seed yield, increased grain yield at standard moisture content, increased shoot length, reduced electrolyte leakage, increased grain weight per plot, increased yield recovery, reduced yield loss, and / or reduced barrenness). A plant or plant part that exhibits improved abiotic stress tolerance may be designated as “abiotic stress tolerance.”

[0066] As used herein, the term “improved drought tolerance” refers to an improvement in one or more water-optimizing traits compared to one or more controls (plants that are not stress-tolerant). Plants or plant parts that exhibit the above-mentioned reduction in water loss, reduction in the accumulation of one or more reactive oxygen species, reduction in the accumulation of one or more salts, increase in salt excretion, increase in the accumulation of one or more dehydrins, improvement in root structure, improvement in osmoregulation, increase in the accumulation of one or more proteins abundant in late embryonic development, improvement in viability, improvement in growth rate, increase in plant height, increase in chlorophyll content, and / or increase in yield, exhibit improved drought tolerance compared to control plants grown under the same drought stress conditions, and may be designated as “drought-tolerant.” In some embodiments, plants or plant parts exhibit increased viability after being subjected to drought stress conditions (e.g., irrigation withholding experiments).

[0067] As used herein, the term “improved osmotic stress tolerance” refers to an improvement in one or more osmotic optimization traits compared to one or more controls (plants that are not stress-tolerant). Plants or plant parts that exhibit the above-mentioned reduction in water loss, reduction in the accumulation of one or more reactive oxygen species, reduction in the accumulation of one or more salts, increase in salt excretion, increase in the accumulation of one or more dehydrins, improvement in root structure, improvement in osmoregulation, increase in the accumulation of one or more proteins abundant in late embryonic development, improvement in viability, improvement in growth rate, increase in plant height, increase in chlorophyll content, and / or increase in yield, when grown under the same osmotic stress conditions, exhibit improved osmotic stress tolerance and may be designated as “osmotic stress tolerant.” In some embodiments, plants or plant parts exhibit increased viability after being subjected to mannitol-induced osmotic stress conditions (e.g., culture in a 200 mM mannitol solution).

[0068] As used herein, the term “improved salt stress tolerance” refers to an improvement in one or more salt-optimizing traits compared to one or more controls (plants that are not stress-tolerant). Plants or plant parts that exhibit the aforementioned reduction in water loss, reduction in the accumulation of one or more reactive oxygen species, reduction in the accumulation of one or more salts, increase in salt excretion, increase in the accumulation of one or more dehydrins, improvement in root structure, improvement in osmoregulation, increase in the accumulation of one or more proteins abundant in late embryonic development, improvement in viability, improvement in growth rate, increase in plant height, increase in chlorophyll content, and / or increase in yield, are considered to exhibit improved salt stress tolerance compared to control plants grown under the same salt stress conditions, and may be designated as “salt stress tolerant.” In some cases, “improved salt stress tolerance” means that the reduction in total dry mass of a stress-tolerant plant under salt stress conditions is 75% or less of the total dry mass of a plant that is not salt-stress tolerant but exhibits the same dry mass as a stress-tolerant plant under normal conditions. In some cases, "improved salt stress tolerance" means that the yield reduction of stress-tolerant plants under salt stress conditions is less than 20% of the total yield of plants that are not salt-tolerant but exhibit the same dry mass as stress-tolerant plants under normal conditions. Salt tolerance can be assessed as described in Negrao et al. Annals of Botany 119.1,1-11 (2017) and Morton et al. The Plant Journal 97.1,148-163 (2019).

[0069] As used herein, the term “improved temperature stress tolerance” refers to an improvement in one or more temperature stress tolerance traits compared to one or more controls (plants that are not stress-tolerant). Plants or plant parts that exhibit the above-mentioned reduction in water loss, reduction in the accumulation of one or more reactive oxygen species, reduction in the accumulation of one or more salts, increase in salt excretion, increase in the accumulation of one or more dehydrins, improvement in root structure, improvement in osmoregulation, increase in the accumulation of one or more proteins abundant in late embryonic development, improvement in viability, improvement in growth rate, increase in plant height, increase in biomass, increase in chlorophyll content, or increase in grain yield may be designated as “temperature stress tolerant” compared to control plants grown under the same temperature stress conditions.

[0070] It should be understood that plants and plant parts exhibiting "drought tolerance," "osmotic stress tolerance," "salt stress tolerance," and "temperature stress tolerance" can also be referred to as "abiotic stress tolerance," since drought stress, osmotic stress, salt stress, and temperature stress are all abiotic stresses.

[0071] As used herein, the term “improved biological stress tolerance” means an improvement in the ability of a plant or plant part to grow, reproduce, and / or survive under biological stress conditions compared to one or more controls (plants that are not stress-tolerant). “Improved biological stress tolerance” may mean any improvement in the growth capacity and / or tolerance capacity of a plant or plant part grown under biological stress conditions, which include, but are not limited to, reduced plant vigor, increased cellular lignification, improved root structure, improved osmoregulation, increased accumulation of one or more proteins abundant in late embryonic development, improved viability, increased growth rate, increased plant height, increased chlorophyll content, and / or increased yield (e.g., increased biomass, increased seed yield, increased grain yield at standard moisture content, increased shoot length, reduced electrolyte loss, increased grain weight per plot, increased yield recovery, reduced yield loss, and / or reduced barrenness). A plant or plant part exhibiting improved biological stress tolerance may be designated “biostress-tolerant.”

[0072] Grafting is the process of grafting a scion onto a rootstock. By grafting a susceptible scion onto a resistant rootstock, a resistant cultivar can be obtained without the need to incorporate resistance into the scion cultivar itself. Furthermore, grafting can enhance the tolerance of susceptible scions to abiotic stress, potentially leading to increased yields and more efficient use of water and nutrients.

[0073] As used herein, “intergeneric hybrid” refers to the hybridization of two individuals from different genera within the same family. “Intergeneric hybrid” refers to a plant, cell, or plant part resulting from an intergeneric hybrid.

[0074] As used herein, “interspecific hybridization” refers to the hybridization of two individuals from different species of the same genus. “Interspecific hybrid” refers to a plant, cell, or plant part resulting from interspecific hybridization.

[0075] The "infinite growth type tomato" produces leaves and flowers throughout its growing season.

[0076] The terms "modified" or "genetically altered" refer to any artificial manipulation of the genome of a target cell.

[0077] As used herein, the term “natural” refers to genes or plants derived from naturally occurring sources or methods. In some aspects, a natural gene refers to a wild-type (non-introduced) gene, whether it is located in its endogenous configuration within the source organism or in a “heterospecific” configuration when introduced into a different organism. A “non-natural” plant is an artificial plant created either by manipulating the number of chromosomes (e.g., allotetraploidy) and / or by grafting two distinct species together to form a single composite plant.

[0078] A "rootstock" is a plant whose lower part (including the roots) can receive a scion during the grafting process.

[0079] RHS refers to the Royal Horticultural Society of England, which publishes an official plant color chart that quantitatively identifies colors according to a prescribed numbering system. This chart can be purchased from Royal Horticultural Society Enterprise Ltd. RHS Garden; Wisley, Woking, Surrey GU236QB, UK.

[0080] "Salt stress" is the accumulation of excessive salt in soil or other growing media that can inhibit crop growth.

[0081] A "scion" is a plant whose upper part can be grafted onto a rootstock during the grafting process.

[0082] "Sequence identity" or "identity" of two nucleic acid or polypeptide sequences refers to the number of residues that are the same in the two sequences when aligned to maximize correspondence over a given comparison window. When the ratio of sequence identity is used for proteins, it is recognized that the positions of non-identical residues are often different due to conservative amino acid substitutions, but in conservative amino acid substitutions, the functional properties of the molecule do not change because an amino acid residue is replaced by another amino acid residue with similar chemical properties (e.g., charge or hydrophobicity). If the sequences differ due to conservative substitutions, the ratio of sequence identity may be adjusted upward to correct for the conservative nature of the substitutions. Sequences that differ due to such conservative substitutions are said to have "sequence similarity" or "similarity." Means for making this adjustment are well known to those skilled in the art and, for example, follow the algorithm in Meyers and Miller, Computer Applic. Biol. Sci., 4:11-17 (1988). The comparison of sequences and the determination of the degree of identity between two sequences can be performed using mathematical algorithms, such as the NCBI Basic Local Alignment Search Tool (BLAST®) (Altschul et al. 1990 J.Mol.Biol. 215:403-10), which is available from several sources, including the National Center for Biotechnology Information (NCBI, Bethesda, Md.) and the Internet, for use in conjunction with the sequence analysis programs blastp, blastn, blastx, tblastn, and tblastx, as well as Clustal W and Clustal X (Larkin et al. 2007 Bioinformatics, 23, 2947-294, Clustal W and Clustal X version 2.0), and Clustal Omega. Unless otherwise specified, references to sequence identity used herein refer to Clustal Omega.

[0083] Plant cells are cells of a plant, cells taken from a plant, or cells obtained by culturing cells taken from a plant. Therefore, the term "plant cells" is not limited to, but may include, cells in seeds, suspension cultures, embryos, meristematic regions, callus tissue, leaves, shoots, gametophytes, sporophytes, pollen, and microspores.

[0084] The term "plant part" refers to parts of a plant, including single cells and cellular tissues, such as intact plant cells, cell masses, and tissue cultures from which plants can be propagated. Examples of plant parts include, but are not limited to, single cells and tissues derived from pollen, ovules, leaves, embryos, roots, root tips, anthers, flowers, fruits, stems, shoots, and seeds, as well as scions, rootstocks, protoplasts, and calluses.

[0085] As used herein, the terms “resistant” or “tolerant” refer to a plant, strain, or variety that exhibits fewer or reduced symptoms than a susceptible (or highly susceptible) plant, strain, or variety. The term may also apply to plants that exhibit no symptoms, and may be referred to as “high / standard tolerant.”

[0086] As used herein, the terms “tolerance” or “tolerance” refer to a plant, strain, or variety that exhibits some symptoms but is still able to produce a commercially viable product at an acceptable yield. These strains may also be referred to as having “moderate / intermediate tolerance.”

[0087] As defined by the International Seed Federation (ISF), a non-governmental, non-profit organization representing the seed industry (see “Definition of the Terms Describing the Reaction of Plants to Pests or Pathogens and to Abiotic Stresses for the Vegetable Seed Industry”, May 2005), the perception of whether a plant is affected by or exposed to pests, pathogens, or abiotic stress can vary depending on the analytical method used. The ISF defines tolerance as the ability of a plant type to limit the proliferation and development of a particular pest or pathogen, and / or the damage they cause, compared to a susceptible plant variety under similar environmental conditions and pressure from pests or pathogens. Even resistant plant types may exhibit symptoms or damage from some diseases. Two levels of tolerance are defined. The term “high tolerance / standard tolerance” is used for plant varieties that, compared to susceptible varieties, highly limit the proliferation and occurrence of a particular pest or pathogen under normal pest or pathogen pressure. "Moderate / intermediate tolerance" applies to plant types that limit the growth and development of a particular pest or pathogen but exhibit more widespread symptoms or damage compared to highly resistant plant types. Plant types with moderate tolerance will not exhibit severe symptoms compared to susceptible plant varieties when grown under similar field conditions and pathogen pressure. Methods for assessing tolerance are well known to those skilled in the art. Such assessments may be made by visual observation of the plant or plant parts (e.g., leaves, roots, flowers, fruits, etc.) in determining the severity of symptoms. For example, if each plant is assigned a tolerance score on a scale of 1 to 5 based on the severity of the response or symptoms, a score of 1 is applied to the most resistant plant (e.g., asymptomatic or with the fewest symptoms), and a score of 5 is applied to the most severely affected plant. A strain is considered resistant if at least 75% of the plants have a tolerance score of 1, 2, or 3, while susceptible strains will have more than 25% of the plants scored at a level of 4 or 5.If a more detailed visual assessment is possible, a scale of 1 to 10 is used to broaden the score range, which is expected to allow for a wider range of scores among the plants being evaluated.

[0088] In addition to such visual assessments, disease can be evaluated by determining the biodensity of pathogens in a plant or plant part using electron microscopy and / or molecular biological methods such as protein hybridization (e.g., ELISA, measurement of pathogen protein density) and / or nucleic acid hybridization (e.g., RT-PCR, measurement of pathogen RNA density). Depending on a particular pathogen / plant combination, for example, a plant may be determined to be resistant to a pathogen if it has a pathogen RNA / DNA and / or protein density of approximately 50%, 40%, 30%, 20%, 10%, 5%, 2%, 1%, 0.1%, 0.01%, 0.001%, or 0.0001% of the RNA / DNA and / or protein density of a susceptible plant.

[0089] General methods of molecular and cellular biochemistry are found in: Molecular Cloning: A Laboratory Manual, 3rd Ed. (Sambrook et al., Harbor Laboratory Press 2001), Short Protocols in Molecular Biology, 4th Ed. (Ausubel et al. eds., John Wiley & Sons 1999), Protein Methods (Bollag et al., John Wiley & Sons 1996), Nonviral Vectors for Gene Therapy (Wagner et al. eds., Academic Press 1999), Viral Vectors (Kaplift & Loewy eds., Academic Press 1995), Immunology Methods Manual (I. Lefkovits ed., Academic Press 1997), Cell and Tissue Culture: Laboratory Procedures in Biotechnology (Doyle & Griffiths, John Wiley & Sons 1998), and Current Protocols. This can be found in standard textbooks such as *in Molecular Biology* (Ausubel et al. eds., John Wiley & Sons 2003) (including appendices 1-117), and these disclosures are incorporated herein by reference.

[0090] overview This disclosure relates to alloploidy tomato plants and hybrid alloploidy tomato plants having desirable properties such as resistance to abiotic or biological stressors, which can be used as rootstock for commercially available tomato varieties. This disclosure further relates to composite plants that include the alloploidy plants described herein as rootstock. This disclosure further relates to methods for producing alloploidy tomato plants and plant parts, as well as hybrid alloploidy tomato plants and plant parts.

[0091] tomato All cultivated forms of tomatoes belong to the species now known as Solanum lycopersicum L. This is the archetypal classification and is now considered more accurate than the earlier name Lycopersicon esculentum Miller, which is still widely used in older literature. Solanum is a large genus containing approximately 2,000 species, including potatoes, tomatoes, and eggplants, among others. The sequence difference between potatoes and tomatoes is approximately 8.7% (Verlaan et al. 2011).

[0092] The exact origins of cultivated tomatoes remain largely unknown, but they are believed to have originated in the Americas, native to Ecuador, Peru, and the Galapagos Islands, and were cultivated by the Aztec and Inca peoples around 700 AD. Mexico is thought to be the earliest place where domestication took place and tomatoes were introduced. The cherry tomato (L. esculentum var. cerasiforme) is the direct origin of modern cultivated forms.

[0093] Tomatoes are cultivated for their fruit and are widely used in fresh markets or as processed products. Tomatoes as a crop are grown commercially wherever environmental conditions allow for economically viable yield production. Although tomatoes are perennial plants, they are usually cultivated as annuals. The majority of fresh tomatoes on the market are harvested by hand at green ripeness, while still on the vine. Tomatoes for processing are used in many forms, including canned tomatoes, tomato juice, tomato sauce, puree, paste, and ketchup.

[0094] Tomatoes are typically diploid species with 12 pairs of chromosomes (n=12, 2n=24). Cultivated varieties have bisexual flowers and can self-pollinate. Fruit shapes vary from small to large and include cherry, plum, pear, block, round, and beefsteak shapes. Tomatoes can sometimes be classified by the time it takes for the plant to mature enough to harvest; cultivars are generally classified into early, mid-season, and late varieties. Tomatoes can also be classified by their growth characteristics (finite or infinite growth). Finite growth plants tend to grow leaves first, then flowers, and fruit ripens on the plant almost simultaneously. Infinite growth tomatoes initially grow a few leaves, and then continue to produce leaves and flowers throughout the growing season. These plants tend to produce tomatoes at various stages of maturation at any given time. In addition to the standard red, ripe color, tomatoes come in a variety of colors, such as milky white, lime green, pink, yellow, gold, orange, and purple.

[0095] Hybrid vigor has been demonstrated in tomatoes, and hybrids are becoming increasingly popular among farmers. Commercial hybrid tomato seeds can be produced through artificial pollination. Pollen from the male parent is collected and manually applied to the stigma of inbred females. Before and after artificial pollination, the flowers are covered to prevent insects from introducing foreign pollen or introducing impurities. Flowers are tagged to identify pollinated fruits, from which seeds are harvested.

[0096] Developing any new and desirable plant genetic resource involves numerous steps. Plant breeding begins with analyzing and defining the problems and weaknesses of existing genetic resources, establishing program objectives, and defining specific breeding goals. The next step is to select genetic resources that possess traits that satisfy the program objectives. The goal is to combine improved combinations of desirable traits derived from parent genetic resources into a single variety or hybrid.

[0097] In the case of tomatoes, these important traits may include improved tolerance to abiotic and / or biological stressors, increased fruit number, increased fruit size and weight, increased seed yield, improved color, resistance to diseases and insects, tolerance to drought and heat, improved uniformity, improved nutritional value and agricultural quality, growth rate, high seed germination rate, seedling vitality, early fruit maturation, ease of fruit setting, adaptability to soil and climatic conditions, hardness, soluble solids content, acidity, and viscosity. For mechanical harvesting of processing tomatoes, fruit density, harvestability, and field retention are also very important.

[0098] Generation of heteropolyploids Ploidy is the presence of more than two sets of homologous chromosomes within a cell's nucleus (Soltis et al. 2009). This phenomenon has a significant impact on plant evolution and speciation (Van de Peer, 2017). Some of the advantages of ploidy include increased organ size ("giga" effect), buffering of harmful mutations, and increased heterozygosity (Sattler et al., 2016). Previous attempts to develop tetraploid tomato lines have been reported (Saeed and Fatima, 2021). However, their offspring often fail to set fruit or produce little to no fruit, resulting in reduced fruit size or seed count (Rick and Butler, 1956, Nilsson 1950).

[0099] Autoploids (sets of chromosomes originating from the same species) are often sterile, while allopolyploids (sets of chromosomes originating from different species) exhibit fertility and restoration of hybrid vigor (Comai, 2005). However, the use of allopolyploid plants for tomato production has not been pursued. One reason is that the fruit size of wild tomato species is usually very small, and the resulting fruit size of hybrids with commercially available tomato plants is below average.

[0100] Allotetraploids are hybrid cells or plants that originate from different species and have four times the number of chromosomes as haploid organisms. As shown in Figure 1, allotetraploid tomatoes can be produced by crossing a commercially available tomato variety, such as Solanum lycopersicum, with a wild tomato variety that possesses a desired trait (e.g., abiotic stress tolerance) (103).

[0101] Following interspecific crosses, polyploidy is used to fix F1 hybrid vigor by chemically induced chromosome doubling (105). The resulting chimeric plants (107) have both diploid and tetraploid cells. Seeds derived from the fruits of the chimeric plants are collected (109) and sown. Alternatively, heterotetraploids may be produced via protoplast fusion.

[0102] The obtained plants are examined for ploidy, and allotetraploid plants are selected (111). The allotetraploid plants may be used as rootstock for commercially available tomato varieties for fruit production, or they may be crossed with another allotetraploid to produce a hybrid allotetraploid, as shown in Figure 2 (203). This hybrid allotetraploid (203) may be used as rootstock for commercially available tomato varieties (205) or as breeding material for fruit production. The allotetraploid and / or hybrid allotetraploid may be further subjected to a chromosome doubling agent to produce an allooctoploid.

[0103] Interspecies hybrids In one embodiment of this disclosure, a commercially available tomato variety is crossed with a wild tomato relative. In some embodiments, a first Solanum lycopersicum plant is crossed with a plant selected from the species shown in the phylogenetic tree of Figure 3 (from Pease et al. (2016) Phylogenomics Reveals Three Sources of Adaptive Variation during a Rapid Radiation. PLoS Biol 14(2)). Figure 3 shows the relationships between all wild tomato species based on information from about half of the known genes in each of these species (in this case, more than 16,500 genes, with each tomato having a total of about 34,000 genes). The left side is a tree constructed from the average species DNA differences from all these genes. The right side shows many individual trees constructed from smaller groups of these genes. These indicate that the evolutionary emergence of current wild species is recent and complex. Genetic distance was calculated as the percentage of total transcriptome sequence difference between different species, with a maximum threshold of 3% (Pose et. al. (2016) Phylogenomics Reveals Three Sources of Adaptive Variation during a Rapid Radiation. PLoS Biol 14(2)). Lineages in the Lycopersicon section have a sequence difference of 2.10% to 2.71% compared to lineages in Lycopersicoides. Lineages within Lycopersicon have a pairwise distance of 0.05% to 1.7%, with the closest relatives being different lineages within S. galapagense (gal-3909 / gal-0436) and different lineages within cultivated tomato (lyc-3475 / lyc-ref).

[0104] Chemically induced chromosome duplication After interspecific hybridization, chromosome doubling agents are sprayed. These chromosome doubling agents may include, but are not limited to, colchicine, trifluralin, oryzarin, amyprophos-methyl, and other polyploidy-inducing agents. Tetraploids can occur spontaneously or be induced using spindle-fiber inhibitors such as colchicine. The technique of colchicine-induced polyploidization has been used since the 1930s. Colchicine inhibits the association of tubulin subunits to the spindle, resulting in the inability of chromosome migration and thus the accumulation of cells in metaphase mitosis. If chromatids separate but are not divided into separate cells by the spindle, the chromosome number doubles, and isoploids are produced.

[0105] When creating polyploids for breeding purposes, it is necessary to double the layer of meristematic cells that give rise to gametophyte tissue. To optimize the probability of successful doubling, a large number of small, actively proliferating meristematic tissues are treated. Colchicine concentrations may vary depending on the tissue and species, but may be used at concentrations, for example, 0.1% to 2.0%. Methods of treating seeds with colchicine or other spindle inhibitors are well known in the art, as discussed in Poehlman, JM, Breeding Field Crops, University of Missouri, Holt, Rinehart and Winston Inc. (1966); Watts, L., Flower and Vegetable Plant Breeding, Grower Books (1980); Callaway DJ and Callaway MB, Breeding Ornamental Plants, Timber Press Inc. (2000).

[0106] In some embodiments, the present disclosure provides a method for producing stress-tolerant allotetraploid tomato plants, comprising crossing a Solanum lycopersicum variety with a stress-tolerant variety to produce interspecific hybrid seeds (in which case the stress-tolerant variety is selected from combinations of the Esculentum group, Arcanum group, Peruvianum group, Hirsutum group, section Lycopersicoides, and their hybrids), cultivating the interspecific hybrid seeds to produce interspecific hybrid plants, spraying the interspecific hybrid plants or their cuttings with a chromosome doubling agent to generate chimeric interspecific hybrids, cultivating the chimeric interspecific hybrids to produce tomato fruits, collecting seeds from the tomato fruits, cultivating the seeds, and selecting stress-tolerant allotetraploid tomato plants.

[0107] Protoplast Fusion In another embodiment, protoplast fusion can also be used to produce allopolyploids. Protoplast fusion is the inducible or spontaneous fusion, such as somatic hybridization, between two or more protoplasts (cells from which the cell wall has been removed by enzymatic treatment) to produce a single binucleated or multinucleated cell. Even plant species that cannot hybridize in nature can yield fused cells, which can then be tissue cultured to create hybrid plants exhibiting a desired combination of traits. In some embodiments, this disclosure shows a method for producing stress-tolerant allotetraploid tomato plants, comprising fusing a protoplast isolated from S. lycopersicum with another protoplast isolated from a stress-tolerant Solanaceae species, selecting a heterokaryon, and regenerating an allotetraploid tomato plant from the heterokaryon. In some embodiments, S. lycopersicum is the acceptor. In some embodiments, the Solanaceae species is sexually incompatible with S. lycopersicum. In some embodiments, the protoplast fusion is asymmetrical. In some embodiments, mitochondria and / or chloroplasts are provided solely by Solanum lycopersicum varieties. In some embodiments, mitochondria and / or chloroplasts are provided solely by wild tomato varieties. In some embodiments, the nucleus is provided solely by Solanum lycopersicum varieties. In some embodiments, the nucleus is provided solely by wild tomato varieties.

[0108] In some embodiments, this disclosure relates to allopolyploids produced by the methods disclosed herein. In some embodiments, this disclosure relates to allopolyploids described herein. In some embodiments, this disclosure relates to seeds of an allotetraploid tomato referred to as "MX20-06," a sample of which is deposited under NCMA accession number 202303001. In some embodiments, this disclosure relates to seeds of an allotetraploid tomato referred to as "MX20-15," a sample of which is deposited under NCMA accession number 202401032.

[0109] In some embodiments, the disclosure relates to stress-tolerant allotetraploid tomato rootstocks comprising chromosomes derived from Solanum lycopersicum and at least one stress-tolerant species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites.

[0110] Generation of heterotetraploid hybrids Further hybrid lines may be created by crossing an allotetraploid with another allotetraploid. In some embodiments, the disclosure provides a method for producing seeds to produce hybrid plants, comprising the step of crossing a first allotetraploid plant with a second allotetraploid plant and harvesting the resulting F1 hybrid seeds. In some embodiments, the first allotetraploid plant consists of a first Solanum lycopersicum variety and a first stress-tolerant variety of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the second allotetraploid plant consists of a second Solanum lycopersicum variety and a second stress-tolerant variety of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites. In some embodiments, the first stress-tolerant variety and the second stress-tolerant variety are different, and the first variety provides at least one tolerance to at least one stressor that is not provided by the second stress-tolerant variety, for example, one being stress tolerance to abiotic stress and the other being stress tolerance to abiotic stress. In another embodiment, the disclosure relates to a hybrid allotetraploid tomato plant and parts thereof grown from a hybrid allotetraploid seed.

[0111] Embryo Rescue Alternatively, embryo rescue may be used in the generation of interspecific hybrids and / or heterotetraploid hybrids. Embryo rescue can be used as a procedure to isolate embryos from the hybrid product and rapidly transition to the next generation for backcrossing or self-pollination, or to the next generation in which the plant cannot produce viable seeds. In this process, the fertilized ovary or immature seeds of the plant are tissue-cultured to create new plants (see Pierik, 1999, In ​​Vitro Culture of Higher Plants, Springer, ISBN 079235267X, 978-0792352679 (this document is incorporated herein by reference in its entirety)).

[0112] tissue culture As is well known in this field, tomato tissue culture can be used for in vitro regeneration of tomato plants. Tissue culture of various tomato tissues and the regeneration of plants from them are well known and publicly available. For example, as described in Girish-Chandel et al., Advances in Plant Sciences. 2000, 13:1, 11-17; Costa et al., Plant Cell Report. 2000, 19:3327-332; Plastira et al., Acta Horticulturae. 1997, 447, 231-234; Zagorska et al., Plant Cell Report. 1998, 17:12 968-973; Asahira et al., Breeding Science. 1995, 45:455-459; Chen et al., Breeding Science. 1994, 44:3, 257-262; and Patil et al., Plant and Tissue and Organ Culture. 1994, 36:2, 255-258, tissue culture including organs is used to produce regenerated plants. As is evident from the literature, these methods for obtaining plants are routinely used with very high success rates at the current level of technology. Therefore, another aspect of this disclosure is to provide cells that produce heterotetraploid tomato plants during propagation and differentiation.

[0113] As used herein, the term “tissue culture” refers to a composition comprising isolated cells of the same or different types, or aggregates of such cells organized into a plant part. Exemplary types of tissue cultures are protoplasts, calluses, plant masses, and plant cells that can produce tissue cultures that are intact in a plant or plant part, such as embryos, pollen, flowers, seeds, leaves, stems, roots, root tips, anthers, pistils, meristem cells, axillary buds, ovaries, seed coats, endosperm, hypocotyls, and cotyledons. Means for preparing and maintaining plant tissue cultures are well known in the art. As an example, tissue cultures containing organs have been used to produce regenerated plants. Specific techniques are described in U.S. Patents 5,959,185, 5,973,234, and 5,977,445, which are incorporated herein by reference.

[0114] Tomato varieties for use in the method of this disclosure Any number of wild, commercial, or cultivated tomato varieties may be used to produce the allopolyploid, hybrid, and grafted (compound) plants described herein. In some embodiments, a first Solanum lycopersicum plant is crossed with a plant selected from the Esculentum group. In some embodiments, a first Solanum lycopersicum plant is crossed with a plant selected from the Arcanum group. In some embodiments, a first Solanum lycopersicum plant is crossed with a plant selected from the Peruvianum group. In some embodiments, a first Solanum lycopersicum plant is crossed with a plant selected from the Hirsutum group. In some embodiments, a first Solanum lycopersicum plant is crossed with a plant selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites. In some embodiments, the Solanum lycopersicum plant is used as the mother in the cross. In some embodiments, the Solanum lycopersicum plant is used as the father in the cross.

[0115] In some embodiments, the wild tomato species is selected from S. galapagense, S. cheesemaniae, S. pimpinellifolium, S. neorickii, S. arcanum, S. chmielewskii, S. huaylasense, S. peruvianum, S. corneliomulleri, S. chilense, S. habrochaites, S. pennellii, S. lycopersicoides, S. sitiens, and combinations thereof.

[0116] Examples of commercially available tomato varieties of Solanum lycopersicum that can be used in the methods and rootstocks disclosed herein include: 42days, 506Bush, A Grappoli D'Inverno, Abracazebra, Ace, Amai, Amana Orange, Amarillo, Amelia, Amish Gold Slicer, Amish Paste, Amsterdam, Ananas Noire, Andiamo, Andrew Rahart's Jumbo Red, Andrina, Anna Aasa, Apero, Applause, Apple Yellow, Arbason, Argentina Cherry, Arkansas Traveler, Armenian, Artic Rose, Attention, Aubry's Special Pink, Aunt Gertie's Gold, Aunt Ginny's, Aunt Molly's Ground Cherry, Aunt Ruby's German Cherry, Aunt Ruby's German Green, Austin's Red Pear, Azoychka, Baby Bottle, and Baby Bottle Red. Pear, Baby Cakes, Baby Grape, Badiaa F1, Bali, Ball's Beefsteak, Banana Legs, Barnes Mountain Yellow, Bartelly, Basinga, Basket Vee, Basrawya, Baxter's Early Bush Cherry, Beall's Gourmet, Beam's Yellow Pear, Beauty King, Beauty Queen, Beefmaster, Beefsteak, Believe It Or Not, Bella Rosa, Bellestar, Bellini, Best Boy, Betalux, Better Boy, Better Bush, Betty, BHN 785, BHN 1021, BHN 189, BHN 268, BHN 444, BHN 543, BHN 589, BHN 602, BHN 624, BHN 762, BHN 826, BHN 871, BHN 901, BHN 961, BHN 964, BHNYC1、Bi-Color Cherry、Big Beef、Big Boy、Big Brandy、Big Bunch、Big League、Big Pink、Big Rainbow、Big Raspberry、Big Red、Big Tiger、Big White、Big White Pink Stripes、Big Yummy、Big Zebra、Bison、Black、Black Cherry、Black Icicle、Black Krim、Black Mauri、Black Opal、Black Pear、Black Pearl、Black Plum、Black Prince、Black Sea Man、Black Strawberry、Black Velvet、Black Zebra、Blondkopfchen、Bloody Butcher、Blue Beauty、Blue Beech、Blue Ribbon、Blush、Bobcat、Bolseno、Boondocks、Booty、Box Car Willie、Bradley、Brandymaster Pink、Brandymaster yellow、Brandysweet Plum、Brandywine、Brandywine Black、Brandywine OTV、Brandywine Pink、Brandywine Red、Brave General、Braveheart、Bronze Torch、Brown Berry、Buckbees New Fifty Day、Buffalo Steak、Bulgarian #7、Bulgarian Triumph、Burbank、Burgess Stuffing Tomato、Burpee’s Big Boy、Burpee’s Burger、Burpee’s Summer Choice、Burrell’s Special、Bush Beefsteak、Bush Big Boy、Bush Blue Ribbon、Bush Early Girl II、Bush Goliath、Cabernet、Cacady’s Folly、Caiman、Camaro、Camelia、Campbell’s 1327、Campbell’s33、Candyland、Capaya、Captain Lucky、Carbon、Carmelita、Carmello、Caro Rich、Carolina Gold、Casa del Sol、Caspian Pink、Celano、Celebration、Celebrity、Celebrity Supreme、Centiflor Red、Cerise Orange、Ceylon、Chadwick Cherry、Chalk’s Early Jewel、Champion、Chancha、Chapman、Charger、Chef’s Choice Black、Chef’s Choice Green、Chef’s Choice Orange、Chef’s Choice Pink、Chef’s Choice Purple、Chef’s Choice Red、Chef’s Choice Striped、Chello、Cherokee Carbon、Cherokee Chocolate、Cherokee Green、Cherokee Purple、Cherries Jubilee、Cherry Baby、Cherry Blossom、Cherry Bomb、Cherry Brandywine、Cherry Buzz、Cherry Ember、Cherry Pink、Cherry Roma、Cherry Sweetie、Chianti Rose Chile、Verde、Chiquita、Chocolate、Chocolate Cherry、Chocolate Pear、Chocolate Sprinkles、Chocolate Stripes、Christmas Grapes、Church、Classica、Clear Pink Early、Clementine、Clermon、Cloudy Day、Cluster Grande、Colonial、Conestoga、Copia、Corbarino、Cordova、Corona、Cosmonaut Volkov Red、Costoluto Fiorentino、Costoluto Genovese、Country Taste、Cour Di Bue、Coustralee、Coyote、Cream Sausage、CremeBrulee、Creole Original、Crimson Cushion Beefsteak、Crimson Sprinter、Crista、Crnkovic Yugoslavian、Crokini、Csiko Botermo、Cupid、Dacquiri、Dads Sunset、Dafel、Dagma’s Perfection、Damsel、Dark Galaxy、David Davidson’s、Daytona、Debaro、Debut、Defiant PhR、Delicious、Delizia、Dester、Dixie Red、Djena Lee’s Golden Girl、Dona、Dorma、Dorothy’s Green、Double Rich、Dr. Carolyn、Dr. Wyche’s Yellow、Druzba、DR7024TS、Earliana、Earl’s Faux、Early Blue Ribbon、Early Boy Bush、Early Cherry、Early Choice、Early Doll、Early First Prize、Early Girl、Early Goliath、Early Harvest、Early Treat、Early Wonder、Edkawi、Egg Yolk、El Dorado、El Fresco Hybrid、Elberta Girl、Elfin、Ella Bella、Emerald Evergreen、Emmy、Emmylou、Empire、Enchantment、Esterina、Estiva、Eva Purple Ball、Evil Olive、Fabulous、Fantastic、Fantastico、Fantome du Laos、Favorita、Fenda、Ferline、Finishline、Firecracker、Fireworks、First Light、First Prize、Five Star Grape、FLA 47R、FLA 7514、Flaming Burst、Floradade、Floralina、Florida 47、Florida 91、Fourth of July、Fox Cherry、Frazier’s Gem、Fresh Salsa、Fried GreenTomato、Front Runner、Frosted Green Doctors、Fruity Cherry、Gabrielle、Galina、Garden Gem、Garden Peach、Garden Treasure、Gardener’s Delight、Garnet、Genuwine、Georgia Streak、German Giant、German Head、German Johnson Pink、German Pink、German Queen、German Red Strawberry、Geronimo、Get Stuffed!、Giallo De Summer、Giant Belgium、Giant Syrian、Giant Tree、Gill’s All Purpose、Gin Fiz、Glacier、Glamour、Gold Medal、Gold Nuggets、Gold Spark、Golden Delight、Golden Gem、Golden Girl、Golden Jubilee、Golden Mama、Golden Peach、Golden Ponderosa、Golden Princess、Golden Queen、USDA Strain、Golden San Marzano、Golden Sunburst、Golden Sunshine、Golden Sweet、Goldene Konigin、Goldie、Golova Negra、Grandaddy、Grandero Plum、Grandeur、Grandma’s Little Girl、Grandma’s Pick、Grandma’s Pick、Grandpap’s Rose Wax、Granny Cantrell’s、Granny Smith、Great White、Greater Baltimore、Green Bell Pepper、Green Berkeley Tie-Dye、Green Doctors、Green Envy、Green Giant、Green Grape、Green Pear、Green Sausage、Green Tiger、Green Zebra、Green Zebra Cherry、Gremlin、GrinchDwarf、Grushovka、Gulf State Market、Gum Drop、Gypsy、Halley 3155、Hard Rock、Harlequin、Harless Creek Gold、Hartman’s Yellow Gooseberry、Hawaiian Pineapple、Health Kick、Heinz 1370 、Heirloom Green、Heirloom Orange、Heritage、High Carotene、Hillbilly、Holland、Homestead、Homesweet、Honey Bunch、Honey Bunch Yellow、Honey Delight、Honey Drop、Honey Hybrid、Honeybee、Honeycomb、Hugh’s、Huichol、Hungarian Heart、Husky Gold、Husky Pink、Hybrid 46、Hybrid Beef 9904、Hy-Brix、Igleheart Yellow Cherry、Ildi、Illini Star、Illinois Beauty、Indian Stripe、Indigo Cream Berries、Indigo Gold Berries、Indigo Kumquat、Indigo Rose、Indigo Ruby、Iron Lady、Isis Candy、Italian Giant Beefsteak、Italian Goliath、Italian Heirloom、Italian Ice、Ivory Pear、Janet’s Jewel、Japanese Trifele Black、Jasper、Jaune Flamme、Jazzy、Jelly Bean Red、Jersey Boy、Jersey Devil、Jet Star、Jetsonic、Joker、Jolly、Jolly Elf、Jolly Girl、Juanita、Jubilee、Jujube Cherry、Juliet、Jung’s Wayahead、Kalman’s Hungarian Pink、Kanner Hoell、Katana、KC 146、Kellogg’s Breakfast、Kimberly、Kobe Beefsteak、Kolb、Koralik、La Roma III、Lady Finger、Ladybug、Lake、Large Barred Boar、Legend、Lemon Boy、Lemon Cherry、Lemon Drop、Lemon Tree、Lime Green Salad、Limmony、LisaKing、Lizziebelle、Lollipop、Lucky Cross、Lucky Tiger、Lunch Box、Lyn’s Mahogany Garnet、Madame Marmande、Maglia Rosa、Magnum、Malakhitovaya Shkatulka、Malinowski、Mama Leone、Mamie Brown’s Pink、Mandarin Cross、Manitoba、Manyel、Margherita、Marglobe Improved、Margo、Mariana、Marion、Marizol Magic、Marizol Purple、Marmande、Marmara、Martian Giant、Martin、Martino’s Roma、Marvel Stripe、Marvelance、Marzinera、Matchless、Mater Sandwich、Matina、Matthew、Maya、Medford、Mega Tom Giant、Megabite、Mexico、Micado Violettor、Midelyce、Midnight Pear、Mighty Sweet、Mingle Mix、Mini Charm、Minibel、Mint Julep、Mirabelle Blanche、Miroma、Mision、Missouri Pink Love Apple、Momotaro、Moneymaker、Montesino、Moonbeam、Moonglow、Moonshadow、Moravsky Div、Moreton、Morning Light、Mortgage Lifter、Mortgage Lifter、bi-color strain、Mosaico、Moskvich、Mountain Delight、Mountain Fresh、Mountain Fresh Plus、Mountain Gem、Mountain Glory、Mountain Gold、Mountain Magic、Mountain Majesty、Mountain Man、Mountain Merit、Mountain Spring、Mountain Vineyard、Mr. Stripey、Mr. Ugly、Mrs. Maxwell’sBig Italian Hr、Napa Grape、Napa Rose Blush、Napoli、Nature Bites、Nature’s Riddle、Nebraska Wedding、Nectar、Nectarine、Neves Azorean Red、New Big Dwarf、New Girl、New Hampshire Red Pickling、New Yorker、Northern Lights、Nova、Nugget、Nyagous、Oaxacan Jewel、Oh Happy Day、Old Brooks、Old Fashioned Goliath、Old German、Old Ivory Egg、Old Yellow Candystripe、Olivade、Orange Banana、Orange Blossom、Orange Fizz、Orange Icicle、Orange Jazz、Orange King、Orange Minsk、Orange Oxheart、Orange Panuche、Orange Peach、Orange Queen、Orange Roma、Orange Russian 117、Orange Santa、Orange Slice、Orange Strawberry、Orange Sunshine、Orange Wellington、Orange Zinger、Oregon Spring、Oroshan、Out Damn Spot、Oxheart Pink、Pamella、Pantano Romanesco、Park’s Beefy Boy、Park’s Early Challenge、Park’s Season Starter、Patty’s Yellow Striped Beefsteak、Paul Robeson、Peacevine、Peach Blow Sutton、Pearly Pink、Pellicore、Peppermint、Perfect Flame、Peron、Persimmon、Phoenix、Picus、Pilcer Vesy、Pineapple、Pineapple Pig、Pink Accordion、Pink Beauty、Pink Berkeley Tie-Dye、PinkBoar、Pink Bumble Bee、Pink Champagne、Pink Cupcake、Pink Girl、Pink Peach、Pink Ping Pong、Pink Pounder、Pink Stuffer、Pink Tiger、Pink Wonder、Pink-a-Licious、Piriform、Pixie Stripe、Placero、Plum Crimson、Plum Lemon、Plum Regal、Polar Beauty、Polar Star、Polbig、Polish Dwarf、Poma Amoris Minora Lutea、Pony Express、Pork Chop、Porter、Porterhouse、Poseidon 43、Power Pops、Prairie Fire、Premio、Prime Beef Goliath、Primo Red、Princess Yum Yum、Principe Borghese、Pritchard、Prize of the Trials、Pruden’s Purple、Purple Boy、Purple Bumble Bee、Purple Russian、Purple Smudge、Quali T 23、Quarter Century、Quedlinburger Fruhe Liebe、Queens、Querida、QualiT-99、QualiT-27、Quick Pick、Quimbaya、RAF、Rally、Ramapo、Rambling Gold Stripe、Rambling Red Stripe、Ramsi、Ranger、Rapunzel、Raspberry Lyanna、Ravello、Razzle Dazzle、Rebekah Allen、Red Anjou、Red Brandywine、Red Candy、Red Cherry Large Fruited、Red Cup、Red Defender、Red Eclipse、Red Fig、Red Grape、Red House Free Standing、Red Lightning、Red Morning、Red Mountain、Red Pear、Red Pearl、Red Plum、Red Pride、RedRave、Red Robin、Red Rocket、Red、Rose、Red Star、Red Zebra、Redfield Beauty、Reisetomate、Ridge Runner、Riesentraube、Rio、Grande、Riviera、Roadster、Rocket、Rojita、Roma、Roman Candle、Rosalita、Rose、Rose De Berne、Rosella、Rosso Sicilian、Rostova、Rowdy Red、Royal Hillbilly、Royal Mountie、Royesta、RuBee Dawn、RuBee Prize、Rugged Boy、Russian Persimon、Russian Rose、Rutgers、Rutgers 250、Rutgers 39、Rutgers Improved PS、Rutgers Select、S 151496、Sakura Honey、Salt Spring Sunrise、Sanibel、Santa Clara Canner、Santiam、Sapho、Sara’s Galapagos、Sasha’s Pride、Scarlet Red、Scarlet Sunrise、Schimmeig Striped Hollow、Sean’s Yellow、Seattle’s Best of All、Seminis 0172-1432、Seminis 1236、Seminis Grape 9137、Serrat、Shady Lady、Shasta、Sheboygan、Shilling Giant、Sicilian Saucer、Siletz、Silvery Fir Tree、Sioux、Skorospelka、Skyreacher、Skyway、Slava、Sleeping Lady、Small Fry、Smarty、Snacker’s Delight、Snow White、Snowberry、Solar Fire、Solar Flare、Solar Power、Solid Gold、Sophie’s Choice、Sophya、Southern Night、Sparky XSL、Spear’s Tennesse Green、Speckled Roman、Spike、Spitfire、Sprite、St. Nick、St. Pierre、Steak House、Steak Sandwich、Stellar、Stone、Striped Cavern、Striped German、Striped Roman、Striped Stuffer、Subarctic、Sugar Lump、Sugar Pl um、Sugar Rush、Sugar Snack、Sugarino、Sugary、Summer Girl、Summer Pick、Summer Pink、Summer Sunrise、Sun Cherry、Sun Dried Cherry、Sun Gold、Sun King、Sunbrite、Sunchocola、Sungold Select II、Sungreen 4029、Sungreen Garden、Sunkist、Sunleaper、Sunlemon、Sunny Blue Ribbon、Sunny Boy、Sunny Goliath、Sunpeach、Sunray、Sunrise、Sunrise Bumble Bee、Sunrise Sauce、Sunset Falls、Sunshine Heirloom、Sunstart、SunSugar、Super Boy 785、Super Bush、Super Fantastic、Super Marmande、Super Snow White、Super Sweet 100、Supernova、SuperSauce、Supersonic、Supersteak、Supertasty、Supremo、SVR 1400、Sweet 100、Sweet Aperitif、Sweet Aroma、Sweet Baby Girl、Sweet Canary、Sweet Carnernos Pink、Sweet Chelsea、Sweet Cluster、Sweet Elite、Sweet Gold、Sweet Hearts、Sweet Million、Sweet Olive、Sweet Orange、Sweet Quartz、Sweet Seedless、Sweet Tangerine、Sweet Treats、Sweet Zen、Sweethearts、Sweetie、Talladega、Tami G、Tamina、Tangella、Tangerine Mama、Tappy’s Hertitage、Tasmanian Blushing、Tasmanian Chocolate、Tasti-Lee、Tasty Evergreen、Tasty Treat、Taxi、Ten Fingers of Naples、TennesseeBritches, Thai Pink Egg, Think Pink, Tidwell German, Tiffen Mennonite, Tiger Like, Tiger Tom, Tigerella, Tinkerbell, Tip-Top, Tocan, Tolstoi, Tomatoberry Garden, Tommy Toe, Tonopah, Top Gun, Topaz or Huan u, Torbay, Toronjina, Tough Boy, Tribeca, Tribute, Trophy, Tropic, Trucker’s Favorite, Tsungshigo Chinese, Tye-Dye, Tygress, Ukrainian Purple, Ultimate Opener, Ultra Pink, Ultra Sweet, Umamin, Umberto, Valencia, Valley Girl, Valleycat, Velvet Red, Vintage Wine, Violaceum Krypni-Rozo, Virginia Sweets, Viva Italia, Volante, Volantis, Wapsipinicon Peach, Washington Cherry, Watermelon Beefsteak, Weissbehaarte, Wes, Wherokowhai, White Beauty, White Cherry, White Currant, White Potato Leaf, White Queen, White Tomesol, White Wax, White Wonder, Whittemore, Wild Cherry, Wild Fred, Willamette, Wins All, Wonder Light, Woodle Orange, Yaqui, Yellow Belgium, Yellow Bell, Yellow Brandywine, Yellow Cherry, Yellow Fire, Yellow Magic, Yellow Mini, Yellow Peach, Yellow Pear, Yellow Perfection, Yellow Stuffer, Yellow Vernissage, Yukon Quest, Zapotec Pink Ribbed, and Zebra Cherry.

[0117] In some embodiments, the Solanum lycopersicum variety is selected from the group consisting of Sugarino, QualiT-99, QuailT-27, Mision, Dorma, Midelyce, Badiaa F1, Volantis, DR7024TS, Marvelance, Juanita, Ramsi, RAF, Edkawi, Golden Princess, and St. Pierre.

[0118] Desirable traits In the case of tomatoes, desirable traits may include increased fruit number, increased fruit size and weight, increased seed yield, improved color, resistance to diseases, pests, and insects, tolerance to drought, heat, cold, and salt damage, improved uniformity, improved nutritional value and agricultural quality, growth rate, high seed germination rate, seedling vitality, early flowering, early fruit maturation, ease of fruit setting, adaptability to soil and climatic conditions, root vitality, plant vitality, fruit firmness, soluble solids content, acidity, and viscosity. For mechanical harvesting of processing tomatoes, fruit density, harvestability, and field retention are also very important.

[0119] In some embodiments, plants are selected based on particularly desirable traits that can be incorporated by the methods of the present disclosure. In some embodiments, the desirable trait is improved resistance to abiotic and biological stressors. Biological stressors include improved resistance to various viral pathogens, fungal pathogens, and bacterial pathogens, as well as to pests. Important diseases include, but are not limited to, tomato yellow leaf curl virus and tomato yellow necrotic virus. Improved resistance to pests is another desirable trait that can be incorporated into novel tomato plants developed by the present disclosure. Pests that affect various species of tomato include, but are not limited to, arthropod pests such as Tuta absoluta, Frankliniella occidentalis, and Bemisia tabaci.

[0120] In some embodiments, desirable traits are resistance to biological stressors such as disease resistance, fungal resistance, pest resistance, bacterial resistance, insect resistance, and nematode resistance.

[0121] In some embodiments, desirable traits are drought tolerance, salinity tolerance, flood / water tolerance, and tolerance to abiotic stressors such as heat tolerance and cold tolerance.

[0122] Salinization is a harmful consequence of irrigation (Tanji, 1990). Salt originates from major minerals in the soil. All surface and groundwater contains dissolved salts absorbed from the soil and geological materials it comes into contact with. Water used for irrigation leaves behind salts when it evaporates or is transpired by agricultural plants. Accumulated salts can adversely affect every stage of plant growth, from seed germination to fruiting. However, irrigation is necessary to increase agricultural productivity in response to the growing demand for food and fodder.

[0123] As freshwater supply decreases, irrigation increases, and more and more land around the world is exposed to the threat of salinization, the need for salinity and drought-tolerant crops is steadily increasing. This disclosure provides a method for transferring salinity tolerance from several wild tomato varieties to cultivated varieties.

[0124] Some wild tomato species are known from small, localized populations isolated in limited microhabitats distinguished by factors such as total rainfall, soil type, and elevation (Rick, 1973; Peralta and Spooner, 2001). Warnock points out that Andes encompasses a diverse set of habitats that give rise to the observed adaptive differentiation of wild tomato species phenotypes (1988).

[0125] The Galapagos Islands have been known as a hotbed of biodiversity since Charles Darwin's visit in 1835. Darwin's five weeks in the Galapagos are widely recognized as one of the most significant events contributing to the formation of his theory of natural selection as a mechanism of biological evolution (On the Origin of Species, 1859). Rich diversity was recorded in various populations of Galapagos tomatoes, including two endemic species (S. galapagense, S. cheesemaniae). The most complete collection of specimens from these tomato populations was created by Charles M. Rick in the 1950s and 1960s and is now maintained by the Tomato Genetic Resources Center (TGRC) at UC Davis.

[0126] Although this plant still grows wild in the Galapagos, many Galapagos tomato populations are threatened by pressure from human development and the invasion of a weed tomato variety (S. esculentum "Gal cer"). The Galapagos tomato has attracted the attention of tomato growers due to its extreme tolerance for high salinity levels and drought tolerance. In fact, the plant can still be found growing on the coast, rooted in small clumps of soil surrounded by dry lava and exposed to sea spray, thriving despite overwhelmingly salty and dry conditions.

[0127] Several researchers (Peralta and Spooner, 2001; Nesbitt and Talksley, 2002; Nuez et al., 2004; Peralta et al., 2006) have shown that the Galapagos tomato is closely related to wild Andean species. In fact, phylogenetic analysis has shown that the Galapagos tomato is deeply situated within a clade of wild Andean tomatoes, one of which was an ancestor of cultivated tomatoes. This clade includes Solanum pimpinellifolium, S. hirsutum, S. pennellii, S. chmieleswskii, S. peruvianum, and S. chilense.

[0128] In some embodiments, the desired trait is tolerance to abiotic stress, and the varieties are LA0089, LA0130, LA0166, LA0247, LA0317, LA0376, LA0400, LA0407, LA0421, LA0426, LA0438, LA0443, LA0453, LA0462, LA0522, LA0716, LA0751, LA1221, LA1237, LA1257, LA1278, LA1294, LA1301, LA1306, LA1310, LA1316, L A1325, LA1331, LA1340, LA1351, LA1352, LA1357, LA1363, LA1367, LA1373, LA1383, LA1392, LA1401, LA1408, LA1421, LA1449, LA1572, LA1579, LA1589, LA1590, LA1609, LA1610, LA1629, LA1630, LA1646, LA1648, LA1674, LA1676, LA1694, LA1722, LA1777, LA1778, LA1809 , LA1923, LA1926, LA1930, LA1932, LA1950, LA1958, LA1959, LA1961, LA1962, LA1969, LA1971, LA1972, LA1974, LA1986, LA2079, LA20 80, LA2081, LA2149, LA2150, LA2157, LA2163, LA2182, LA2309, LA2310, LA2311, LA2327, LA2403, LA2408, LA2425, LA2560, LA2661, LA2 662, LA2710, LA2711, LA2744, LA2747, LA2748, LA2755, LA2773, LA2876, LA2877, LA2878, LA2880, LA2884, LA2885, LA2931, LA2963, LA 2964, LA3120, LA3153, LA3320, LA3465, LA3799, LA3847, LA4023, LA4105, LA4108, LA4133, LA4321, LA4324, LA4330, LA4335, Arkansas Traveler, Delicious, Edkawi, German Johnson Pink, Golden Princess, Heinz 1370, Homestead, New Yorker, RAF, Ramsi, and St.The group is selected from the group consisting of Pierre.

[0129] In some embodiments, the desired trait is biological stress tolerance, and the varieties are LA0490, LA0655, LA0656, LA1783, LA1791, LA1792, LA1800, LA1802, LA1964, LA1995, LA2009, LA2356, LA2369, LA2396, LA2443, LA2444, LA2445, LA2446, LA2447, LA2448, LA2449, LA2458, LA2530, LA2531A, LA2531 B, LA2531C, LA2701, LA2818, LA2819, LA2820, LA2821, LA2822, LA2823, LA2824, LA2825, LA2826, LA2827, LA2828, LA2829, LA2830 , LA2968, LA3025, LA3026, LA3027, LA3028, LA3029, LA3038, LA3039, LA3040, LA3041, LA3042, LA3043, LA3044, LA3045, LA3046, L A3047, LA3048, LA3049, LA3050, LA3051, LA3118, LA3129, LA3130, LA3145, LA3151, LA3152, LA3158, LA3159, LA3160, LA3161, LA 3201, LA3202, LA3213, LA3214, LA3215, LA3216, LA3254, LA3258, LA3268, LA3269, LA3271, LA3273, LA3275, LA3276, LA3277, LA32 92, LA3297, LA3309, LA3310, LA3313, LA3314, LA3341, LA3342, LA3428, LA3432, LA3433, LA3471, LA3472, LA3473, LA3526, LA3667 , LA3839, LA3845, LA3846, LA3847, LA3856, LA3858, LA3859, LA3912, LA4025, LA4026, LA4285, LA4286, LA4441, LA4442, Arkansas selected from the group consisting of Traveler, Delicious, Edkawi, German Johnson Pink, Golden Princess, Heinz 1370, Homestead, New Yorker, RAF, Ramsi, and St. Pierre.

[0130] In some embodiments, a selected variety having one or more desirable traits is a variety that has at least 80% homozygosity and / or is strictly or preferentially self-pollinating.

[0131] In some embodiments, the desired trait is a single gene trait. A single gene trait may or may not be transgenic. Examples of such traits include male sterility (e.g., ms1, ms2, ms3, ms4, or ms5 genes), herbicide resistance (e.g., bar gene or PAT gene), bacterial disease resistance, fungal disease resistance (e.g., Cladosporium fulvum resistance gene Cf), or viral disease resistance (e.g., tomato yellow leaf curl virus (TYLCV) resistance gene Ty, tomato mosaic tobamovirus (ToMV) resistance genes Tm-1, Tm-2, and Tm2). 2 These include, but are not limited to, increased brix through the introduction of specific alleles such as insect resistance (nematode resistance gene Mi) and the hir4 allele of Lycopersicon hirsutum (S. habrochaites), improved storage life through the use of mutants such as the dg mutant described in US2008-0184382 (high lycopene), rin (ripening inhibitor), nor (unripened), or cnr (colorless unripened) alleles, increased fruit hardness or delayed softening due to mutations in the expansin gene, etc., the absence of gel (i.e., solid fruit with hollow regions lacking gel or liquid contents), improved fertility, nutritional value, sugar content, yield stability, and increased yield through the use of the PSAF allele. These genes are usually inherited via the nucleus. Some known exceptions to this are the genes for male sterility, some of which are inherited cytoplasmically but still act as single genetic traits. Some of these single genetic traits are described in U.S. Patents 5,777,196, 5,948,957, and 5,969,212, which are incorporated herein by reference.

[0132] Other desirable traits include those associated with improved tomato fruit. A non-restrictive list of fruit phenotypes used during breeding selection includes:

[0133] °Brix is ​​a measure of the total soluble solids (TSS) content in tomatoes or tomato products. The TSS in tomatoes is mainly sugar (fructose). 1°Brix means that 1 gram of soluble sugar is contained in 100 grams of solution. Tomato juice rated as having 20°Brix contains 200 g / liter of soluble sugar.

[0134] Bostwick Average for Juice: A measurement of the Bostwick viscosity of the juice. The viscosity or concentration of a tomato product is influenced by the concentration of the tomato, the amount and degree of pectin breakdown, the size, shape, and quality of the pulp, and possibly to a lesser extent, by proteins, sugars, and other soluble components. Viscosity is measured in Bostwick centimeters using instruments such as a Bostwick viscometer.

[0135] pH: pH is a measure of the acidity of fruit puree. A pH of less than 4.5 is desirable to prevent bacterial spoilage of the finished product. pH increases as the fruit ripens.

[0136] Fruit color: Fruit color is measured as the Hunters a / b ratio, where a represents red / green, positive values ​​are red, negative values ​​are green, and 0 is neutral; b represents yellow / blue, positive values ​​are yellow, negative values ​​are blue, and 0 is neutral; a / b represents the intensity of redness, with larger values ​​indicating dark red and smaller values ​​indicating light red or yellowish-red.

[0137] Fruit weight: The average weight of a single fruit or a group of fruits, measured at ripeness at harvest and recorded in a convenient unit of measurement.

[0138] Ostwald Viscosity: Ostwald viscosity is a measure of serous viscosity, and the measurement is performed using an Ostwald viscometer. Serum is the non-solid portion of the tomato extract after centrifugation of tomato puree. Serum viscosity is affected by the amount and quality of soluble pectin. A higher number reflects a higher viscosity of the tomato serous viscosity.

[0139] Fruit hardness: Fruit hardness is the resistance to penetration and is measured using a digital durometer model DD-4-00 (Rex Gauge Company, Buffalo Grove, IL, USA). The tomato is placed on its side, and durometer readings are taken at four points approximately 90 degrees apart in the center of the tomato. Penetration resistance is measured using the durometer at four points for each of the nine individual fruit samples taken from a given location (a total of 36 independent measurements). The P5 value is calculated from the following formula: D-39 / 10, where D is the value obtained from the durometer.

[0140] It should be understood that in certain embodiments, plants may be selected based on the absence, suppression, or inhibition of certain features or traits (such as undesirable features or traits), rather than the presence of certain features or traits (such as desirable features or traits).

[0141] It is also conceivable to select plants based on genotypic information (e.g., the pattern of gene expression, genotype, or presence of genetic markers). When evaluating the presence of one or more genetic markers, those markers may be known and / or associated with specific characteristics of the plant (e.g., markers may be associated with increased growth rate or metabolic profile). This information can be used in the methods of this disclosure in combination with evaluations based on other characteristics to select different combinations of potentially desirable plant characteristics. Novel quantitative trait loci (QTLs) may be identified using such techniques. For example, plants may be selected based on growth rate, size (including, but not limited to, weight, height, leaf size, stem size, branching pattern, or size of any part of the plant), overall health, survival rate, tolerance to harmful physical environments, and / or any other characteristics described herein.

[0142] Further non-limiting examples include selecting plants based on seed germination rate, amount of biomass produced, improved root and / or leaf / shoot growth leading to increased yield (fruit) or biomass production, effects on plant growth leading to increased seed yield of crops, effects on plant growth leading to increased yield, effects on plant growth leading to increased resistance or tolerance to fungal, viral, or bacterial diseases, mycoplasma, or pests such as insects, mites, or nematodes (in this case, damage is measured by reduced leaf symptoms such as the incidence of bacterial or fungal lesions, or the area of ​​damaged leaves, or a decrease in the number of nematode cysts or galls present on the plant roots, or an improvement in plant yield in the presence of such plant pests and diseases), effects on plant growth leading to increased metabolite yield, and effects on plant growth leading to improved aesthetic appeal (this can be particularly important for plants cultivated for shape, color, or taste (e.g., the intensity of color of the outer peel of tomatoes)).

[0143] Compound (non-naturally grafted) plants Grafting is a widely used method of asexual plant propagation in agriculture and horticulture, involving the fusion of tissue from one plant with tissue from another. Grafting involves combining two independent plant parts into a single plant. Such combinations can be performed in a variety of ways, including but not limited to cleft grafting, stomach grafting, tongue grafting, stump grafting, slit grafting, cleft grafting, glove grafting, tongue grafting, snap grafting, leaf-bud cleft grafting, saddle grafting, joint grafting, and bud grafting (superficial bud grafting, slanted bud grafting, T-bud grafting) (for further details, see Garner RJ, The Grafter's Handbook, 5th Ed edition (March 1993), Cassell Academic; ISBN: 0304342742). Grafting produces non-natural composite plants.

[0144] One embodiment of this disclosure relates to a composite plant comprising an allopolyploid as a rootstock as described herein, and a method for producing the composite plant. In some embodiments, the scion is an elite commercial tomato variety. In some embodiments, the commercial variety is the same Solanum lycopersicum variety used for the initial interspecific hybridization or protoplast fusion to produce the allopolyploid rootstock, and thus the rootstock and scion share at least one allele. In some embodiments, the rootstock and scion share at least one chromosome. In some embodiments, the rootstock and scion share a set of chromosomes. In some embodiments, this disclosure relates to a chimeric plant tissue produced by grafting a Solanum lycopersicum variety as a scion onto an allopolyploid plant as described herein as a rootstock. In some embodiments, the chimeric plant tissue comprises a first plant cell and a second plant cell, the first plant cell being an allopolyploid containing chromosomes derived from Solanum lycopersicum, as well as chromosomes derived from at least one species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the second plant cell being a diploid Solanum lycopersicum.

[0145] The allopolyploid plants described herein are graft-compatible and suitable for use as rootstocks for tomato cultivars, as well as scions for other vegetable crops, including pepper and eggplant. The allopolyploid plants disclosed herein and methods for producing them possess and can confer any number of desirable traits, including but not limited to tolerance and / or tolerance to salt stress, cold stress, heat stress, and drought stress, disease resistance, fungal resistance, pest resistance, bacterial resistance, insect resistance, and nematode resistance, to scions. Furthermore, the allotetraploid plants described herein can further increase the yield of scion plants. Due to their high uniformity, the allotetraploid plants disclosed herein are particularly suitable for automatic grafting using automatic grafting machines. Accordingly, in another embodiment, this disclosure shows a method for conferring desirable traits derived from wild species of tomato to cultivated Solanum lycopersicum varieties by grafting.

[0146] Grafting is a process that has been used for many years in crops such as citrus plants and members of the Cucurbitaceae family, but its use in the production of some commercial tomatoes is relatively recent. Typically, a variety to be used as the scion (usually an elite commercial variety) is grafted onto a rootstock variety that possesses desirable traits such as resistance to abiotic or biological stress. Thus, a resistant rootstock maintains health and provides nutrients from the soil to the scion. Some recent developments have also shown that some rootstocks can also improve the agricultural value of the grafted plant, particularly in tomato cultivation, where balancing vegetative and reproductive growth is always difficult.

[0147] Several methods exist for grafting tomatoes. Examples of suitable grafting methodologies include, but are not limited to, cleft grafting, splice grafting, micrografting, tongue grafting, side grafting, and high grafting. Cleft grafting involves making a V-shaped incision in the rootstock and inserting a complementary wedge-shaped scion. The graft may then be held in place with a small clip until it heals. Splice grafting, also known as tongue grafting (TAG), involves making incisions on opposite sides of the stems of the rootstock and scion, and then holding the stems in place with a clip while the stems fuse. Once the graft has healed, the scion of the desired rootstock plant may be removed above the graft site, and the unused rootstock of the scion plant may be separated from the scion below the graft site. Micrografting, also known as splice grafting, is a technique that has recently been integrated into the micropropagation production of hybrid tomatoes. Micrografting involves utilizing micropropagated scion shoots that can be grafted onto rootstock seedlings approximately three weeks old. In some embodiments, micrografting is used for commercial-scale tomato grafting. Tube grafting involves cutting the scion and rootstock as seedlings and attaching the cut rootstock seedling to the cut scion seedling using a small silicone tube, with or without clips. Tube grafting can be performed when the plants are very small and can be very effective as it increases yield while eliminating the need for a large healing chamber. Although less frequently used on a commercial scale, side grafting and top grafting are also discussed herein. See also (Lee, 1994; Lee and Oda, 2003; Hanna, 2012; Lee and Oda, 2003; Oda, 1995; Rivard and Louws, 2006; Vu et al., 2015; Bausher, 2013; Rivard and Louws, 2006; Kubota et al., 2008; and Lee, 2003).

[0148] Deposit Information The deposit of heterotetraploid tomato seeds described herein is managed by Red Sea Farms LTD, 2435, Al Sila Tower, 24th Floor, Abu Dhabi Global Market Square, Al Maryah Island, Abu Dhabi, United Arab Emirates.

[0149] Furthermore, 625 seed samples of the “MX20-06” variety of this disclosure have been deposited with an international depositary established under the Budapest Treaty in accordance with 37 CFR 1.803(a)(1). The applicant deposited the seeds with the Provasoli-Guillard National Center for Marine Algae and Microbiota (NCMA) at Bigelow Laboratory for Ocean Science, 60 Bigelow Drive East Boothbay, ME 04544. The “MX20-06” seeds were deposited with and accepted by the Budapest Treaty on March 6, 2023, under NCMA number 202303001.

[0150] Furthermore, a sample of 625 seeds of the “MX20-15” variety of this disclosure has been deposited with an international depositary established under the Budapest Treaty in accordance with 37 CFR 1.803(a)(1). The applicant deposited the seeds with the Provasoli-Guillard National Center for Marine Algae and Microbiota (NCMA) at Bigelow Laboratory for Ocean Science, 60 Bigelow Drive East Boothbay, ME 04544. The “MX20-15” seeds were deposited with and accepted by the Budapest Treaty on January 30, 2024, under NCMA number 202401032.

[0151] To demonstrate that the enablement requirements of 35 USC112 are met and that the deposit of heteropolyploids in this disclosure meets the criteria set forth in 37 CFR 1.801-1.809 and the Patent Search Procedure Manual (MPEP) 2402-2411.05, the applicant hereby makes the following statement regarding the deposited seeds: 1. While this application is pending, access to this disclosure will be provided to the Director of the Patent Office upon request. 2. When a patent is granted under the conditions set forth in 37 CFR 1.808, all restrictions on general availability are irreversibly lifted. 3. The deposited items will be kept in the public repository for 30 years, or 5 years from the last claim, or the term of the patent, whichever is longer. 4. The viability of the biological material at the time of deposit will be tested by the public depositary in accordance with 37 CFR 1.807. 5. If a deposited item becomes unusable, it will be replaced.

[0152] Access to this deposit is permitted only to persons whom the Director of the Patent and Trademark Office has determined to have rights under 37 CFR §1.14 and 35 U.S.C §122 during the pendency of this application. If the request in this application is granted, all restrictions on the public availability of this variety will be irreversibly lifted by granting the NCMA access to the deposit of at least 625 seeds of the same variety.

[0153] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those generally understood by those skilled in the art to which the present invention belongs. Any methods and materials similar or equivalent to those described herein may be used in carrying out or testing the present invention, but non-limiting methods and materials are described herein.

[0154] All publications and patent applications referenced herein represent the level of skill of those skilled in the art to which the present invention pertains. All publications and patent applications are incorporated herein by reference to the same extent that each individual publication or patent application is shown to be incorporated specifically and individually by reference. Nothing herein should be construed as acknowledging that this disclosure does not qualify as prior to such publications by prior disclosure.

[0155] Many modifications and other embodiments of the disclosure described herein will be recalled by those skilled in the art to the extent of these disclosures, benefiting from the teachings presented in the foregoing description and the relevant drawings. Therefore, it should be understood that this disclosure is not limited to the specific embodiments disclosed, and modifications and other embodiments are intended to be included within the scope of the appended claims. Certain terms are used herein, but these are used only in a general and descriptive sense and are not intended to be limiting.

[0156] While this disclosure has been described in relation to specific embodiments, it is understood that further modifications are possible, and this application is intended to include any modifications, uses, or adaptations of this disclosure, including deviations from the disclosure in accordance with the appended claims, and which may be applied to the essential features set forth herein, and which may be applied in accordance with the principles of this disclosure and which may be known or customary practices in the art to which this disclosure belongs. [Examples]

[0157] Example 1 - Heterotetraploid variety "MX20-06" The heterotetraploid variety "MX20-06" was created through interspecific cross between the S. lycopersicum and S. pimpinellifolium varieties.

[0158] Tomato varieties of S. lycopersicum are cultivated locally in fields in Saudi Arabia due to their vigor and adaptability to drought environments. These flowers were used as mother flowers, the anthers were removed, and pollination was carried out with pollen extracted from S. pimpinellifolium strains selected for their salt tolerance. The crosses were performed under controlled conditions in a research greenhouse at King Abdullah University of Science and Technology (KAUST), Thuwal, Saudi Arabia. The resulting F1 interspecific hybrid seeds demonstrated vigorous growth and salt tolerance.

[0159] Salt tolerance of F1 interspecific hybrid seedlings was calculated as stress-weighted performance (SWP), as described in Saade et al. Sci Rep 6,32586 (2016). Using the same index, we evaluated the salt tolerance conferred to composite plants grafted onto F1 hybrid rootstocks using the Super Sweet commercial tomato variety as the scion.

[0160] At the seedling stage, the shoots of selected F1 interspecific hybrids were cut, and most of the leaves were removed without damaging the apical meristem. The cuts were immersed overnight in a 5 mM colchicine solution with gentle shaking. After washing three times to remove the colchicine, the cuts were transplanted into soil to grow roots and regenerate as chimeric plants. Tomato fruits obtained from these plants were then collected, and the seeds were sown. The polyploidy was examined by flow cytometry, and the nuclear DNA content was measured compared to that of diploid samples (Figures 9A and 9B). The heterotetraploid variety "MX20-06" was selected based on uniformity and seed yield.

[0161] The commercially available rootstock variety most closely related to the allotetraploid variety "MX20-06" is "Maxifort." However, as shown in Table 1 below, the allotetraploid variety "MX20-06" has a different morphology compared to the commercially available rootstock variety "Maxifort." See also Figures 4-8 and 13C. [Table 1-1] [Table 1-2] [Table 1-3]

[0162] The heterotetraploid variety "MX20-06" exhibits trait uniformity and stability because its heterozygosity is fixed (achieved by a chromosome doubling agent).

[0163] The allotetraploid variety "MX20-06" has been tested as a rootstock. Approximately 84 compound plants were produced by grafting the commercially available variety "Sugarino" onto the allotetraploid "MX20-06" rootstock. For comparison, "Sugarino" was also grafted onto the commercially available rootstock variety "Maxifort," producing approximately 84 compound plants. All grafting was performed manually using standard techniques. The compound plants were cultivated in the experimental greenhouse at Red Sea Farms, Thuwal, Saudi Arabia. The Sugarino-MX20-06 compound plants yielded an average fruit yield of 274.57g per plant, while the Sugarino-Maxifort compound plants yielded an average fruit yield of 237.71g per plant. Therefore, the Sugarino-MX20-06 compound plants showed a 16% increase in yield compared to the Sugarino-Maxifort compound plants (Figure 14A).

[0164] The allotetraploid variety "MX20-06" was tested as a rootstock under combined heat and salt stress. Approximately 10 compound plants were produced by grafting the commercially available variety "Midelyce" onto the allotetraploid "MX20-06" rootstock. For comparison, "Midelyce" was also grafted onto the commercially available rootstock variety "Maxifort," producing approximately 10 compound plants. All grafting was performed manually using standard techniques. The compound plants were cultivated in the experimental field at King Abdullah University of Science and Technology (KAUST), Thuwal, Saudi Arabia. The experimental period was 120 days. The plants were irrigated with 6% seawater (EC=6.22 mS / cm) and exposed to temperatures up to 42°C. The Midelyce-MX20-06 composite plant yielded an average fruit yield of 86g per plant, while the Midelyce-Maxifort composite plant yielded an average fruit yield of 55g per plant, and the ungrafted Midelyce plant yielded an average fruit yield of 50g per plant. Therefore, the Midelyce-MX20-06 composite plant showed a 56% increase in yield compared to the Midelyce-Maxifort composite plant and a 72% increase compared to the ungrafted Midelyce plant (Figure 14B).

[0165] In a 150-day field trial conducted in the Al-Dabaa Road region behind Wadi Al-Natrun, Egypt, the allotetraploid variety 'MX20-06' was tested as a commercial-scale rootstock (Figures 16A-16C). Commercial variety 'CH7' was grafted onto the allotetraploid 'MX20-06' rootstock to produce approximately 3,000 compound plants. For comparison, ungrafted 'CH7' plants were planted. All grafting was performed manually using standard techniques. The field consisted of poorly drained loamy soil with a salinity of 4.1 EC (ds / m) and a pH of 8.2. Drip irrigation was provided with water with a salinity of 434.3 ppm and a pH of 7.3. The seedling density was 30 cm. Fruit was harvested over four harvest periods (H1-H4).

[0166] In total, the CH7-MX20-06 composite plants yielded an average of 3.37 kg of fruit per plant, compared to an average of 1.55 kg per plant from ungrafted "CH7" plants. Therefore, the CH7-MX20-06 composite plants showed a 117% increase in yield compared to ungrafted "CH7" plants (Figure 17). The average fruit weight in the CH7-MX20-06 composite plants was 140 g per fruit, compared to 127 g per fruit in ungrafted "CH7" plants. Therefore, the CH7-MX20-06 composite plants showed a 10% increase in fruit size compared to ungrafted "CH7" plants (Figure 18). The uprooted CH7-MX20-15 composite plants showed a noticeable increase in vitality and root size (Figure 19).

[0167] In Stockton, California, the allotetraploid variety 'MX20-06' was tested commercially at two different planting densities as a rootstock for processing tomato production (Figures 20A-F). The tests were conducted in a delta area containing fine alluvial soil (Kingile Much) with an organic matter content of 40%. The commercially available variety 'UG161-12' was grafted onto the allotetraploid variety 'MX20-06' rootstock. For comparison, 'UG161-12' was grafted onto the commercially available rootstock variety 'Maxifort', and ungrafted 'UG161-12' plants were also planted. All grafting was performed manually using standard techniques. The composite plant was planted at two different planting densities: standard spacing (row spacing): 7,500 plants per acre, 13.94 inches apart (18,533 plants per hectare), and sparse planting: 3,750 plants per acre, 27.88 inches apart (9,266 plants per hectare). The experiment lasted four months.

[0168] At standard planting densities, the UG161-12-MX20-06 rootstock compound yielded an average of 156.22 tons of fruit per hectare, compared to 140.43 tons per hectare for the UG161-12-Maxifort compound and 144.92 tons per hectare for the ungrafted UG161-12 plants. Therefore, the UG161-12-MX20-06 compound yielded an 11% increase compared to the UG161-12-Maxifort compound and an 8% increase compared to the ungrafted UG161-12 plants. At sparse planting densities, the UG161-12-MX20-06 compound yielded an average of 155.17 tons of fruit per hectare, compared to an average of 117.74 tons per hectare from the UG161-12-Maxifort compound and an average of 122.39 tons per hectare from ungrafted UG161-12 plants. Therefore, the UG161-12-MX20-06 compound yielded a 32% increase compared to the UG161-12-Maxifort compound and a 27% increase compared to ungrafted UG161-12 plants. The yield difference for the UG161-12-MX20-06 composite plant was only 1.05 t / ha, while the yield differences for UG161-12-Maxifort and ungrafted UG161-12 plants were 22.69 t / ha and 22.53 t / ha, respectively. This indicates that the UG161-MX20-06 composite plant maintains good yields even at low planting densities (Figure 21).

[0169] Example 2 - Heterotetraploid variety "MX20-15" The heterotetraploid variety "MX20-15" was created through interspecific crossbreeding of a S. pimpinellifolium strain and a S. lycopersicum variety.

[0170] The S. pimpinellifolium strain was selected based on its salt tolerance. Using this flower as the mother flower, the anthers were removed, and pollination was performed with pollen extracted from a S. lycopersicum tomato variety locally grown in Saudi Arabia due to its vigor and adaptability to drought environments. The crosses were carried out under controlled conditions in a research greenhouse at King Abdullah University of Science and Technology, Thuwal, Saudi Arabia. The resulting F1 interspecific hybrid seeds demonstrated vigorous growth and salt tolerance.

[0171] Salt tolerance of F1 interspecific hybrid seedlings was calculated as stress-weighted performance (SWP), as described in Saade et al. Sci Rep 6,32586 (2016). Using the same index, we evaluated the salt tolerance conferred to composite plants grafted onto F1 hybrid rootstocks using the Super Sweet commercial tomato variety as the scion.

[0172] At the seedling stage, shoots of selected F1 interspecific hybrids were cut, and most leaves were removed without damaging the apical meristem. The cuts were immersed overnight in a 5 mM colchicine solution with gentle shaking. After washing three times to remove the colchicine, the cuts were transplanted into soil to grow roots and regenerate as chimeric plants. Tomato fruits obtained from these plants were then collected, and the seeds were sown. The polyploidy was examined by flow cytometry, and the nuclear DNA content was measured compared to that of diploid samples. The heterotetraploid variety "MX20-15" was selected based on uniformity and seed yield.

[0173] The commercially available rootstock variety most closely related to the allotetraploid variety "MX20-15" is "Maxifort." However, as shown in Table 2 below, the allotetraploid variety "MX20-15" has a different morphology compared to the commercially available rootstock variety "Maxifort." See also Figures 22-26 and 13D. [Table 2-1] [Table 2-2] [Table 2-3]

[0174] The heterotetraploid variety "MX20-15" exhibits trait uniformity and stability because its heterozygosity is fixed (achieved by a chromosome doubling agent).

[0175] The allotetraploid variety "MX20-15" was tested as a rootstock under combined heat and salt stress. Approximately 10 compound plants were produced by grafting the commercially available variety "Midelyce" onto the allotetraploid "MX20-15" rootstock. For comparison, "Midelyce" was also grafted onto the commercially available rootstock variety "Maxifort," producing approximately 10 compound plants. All grafting was performed manually using standard techniques. The compound plants were cultivated in the experimental field at King Abdullah University of Science and Technology (KAUST), Thuwal, Saudi Arabia. The experimental period was 120 days. The plants were irrigated with 6% seawater (EC=6.22 mS / cm) and exposed to temperatures up to 42°C. The Midelyce-MX20-15 composite plant yielded an average fruit yield of 115g per plant, while the Midelyce-Maxifort composite plant yielded an average fruit yield of 55g per plant, and the ungrafted Midelyce plant yielded an average fruit yield of 50g per plant. Therefore, the Midelyce-MX20-15 composite plant showed a 109% increase in yield compared to the Midelyce-Maxifort composite plant and a 130% increase compared to the ungrafted Midelyce plant (Figure 14B).

[0176] The allotetraploid variety "MX20-15" was tested as a rootstock under salt stress. Approximately 40 compound plants were produced by grafting the commercially available variety "Midelyce" onto the allotetraploid "MX20-15" rootstock. For comparison, "Midelyce" was also grafted onto the commercially available rootstock variety "Maxifort," producing approximately 40 compound plants. All grafting was performed manually using standard techniques. The compound plants were cultivated in an experimental greenhouse at Red Sea Farms, Thuwal, Saudi Arabia. The experimental period was 120 days. Under salt stress, the plants were irrigated with 6% seawater (EC=6.22 mS / cm).

[0177] Under control conditions, the Midelyce-MX20-15 compound plant yielded an average fruit yield of 235g per plant, while the Midelyce-Maxifort compound plant yielded an average fruit yield of 228g per plant, and the ungrafted Midelyce plant yielded an average fruit yield of 186g per plant. Therefore, the Midelyce-MX20-15 compound plant showed a 3% increase in yield compared to the Midelyce-Maxifort compound plant and a 26% increase in yield compared to the ungrafted Midelyce plant.

[0178] Under salt stress, plants were irrigated with 6% seawater (EC=6.22 mS / cm). Midelyce-MX20-15 composite plants yielded an average fruit yield of 191g per plant, while Midelyce-Maxifort composite plants yielded an average fruit yield of 115g per plant, and ungrafted Midelyce plants yielded an average fruit yield of 111g per plant. Therefore, Midelyce-MX20-15 composite plants showed a 66% increase in yield compared to Midelyce-Maxifort composite plants and a 72% increase compared to ungrafted Midelyce plants (Figure 15).

[0179] In a 150-day field trial conducted in the Al-Dabaa Road region behind Wadi Al-Natrun, Egypt, the allotetraploid variety 'MX20-15' was tested as a commercial-scale rootstock (Figures 16A-16C). Commercial variety 'CH7' was grafted onto the allotetraploid 'MX20-15' rootstock to produce approximately 3,000 compound plants. For comparison, ungrafted 'CH7' plants were planted. All grafting was performed manually using standard techniques. The field consisted of poorly drained loamy soil with a salinity of 4.1 EC (ds / m) and a pH of 8.2. Drip irrigation was provided with water with a salinity of 434.3 ppm and a pH of 7.3. The seedling density was 30 cm. Fruit was harvested over four harvest periods (H1-H4). In total, the CH7-MX20-15 composite plants yielded an average of 2.99 kg of fruit per plant, compared to an average of 1.55 kg per plant from ungrafted "CH7" plants. Therefore, the CH7-MX20-15 composite plants showed a 93% increase in yield compared to ungrafted "CH7" plants (Figure 17). The average fruit weight in the CH7-MX20-15 composite plants was 155 g per fruit, compared to 127 g per fruit in ungrafted "CH7" plants. Therefore, the CH7-MX20-15 composite plants showed a 22% increase in fruit size compared to ungrafted "CH7" plants (Figure 18).

[0180] The examples of the related technology described above and the limitations thereto are illustrative and not exclusive. Other limitations of the related technology will become apparent to those skilled in the art by reading this specification.

[0181] Numbered Embodiments 1. A non-naturally grafted plant, A heterotetraploid rootstock consisting of a first Solanum lycopersicum variety and a stress-tolerant variety of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and At least one scion of a second Solanum lycopersicum variety grafted onto the aforementioned rootstock. The aforementioned non-natural grafted plant, consisting of the above.

[0182] 2. The aforementioned heterotetraploid rootstock Interspecific crosses were performed between a first Solanum lycopersicum variety and stress-tolerant varieties of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the seeds produced by the crosses were collected. Hybrid plants are grown from the aforementioned seeds, and cuttings taken from the aforementioned hybrid plants are treated with a chromosome doubling agent. The processed cuttings are allowed to grow, the resulting plants are self-pollinated, and the seeds are grown to produce plants. The DNA content and chromosome set of the obtained plants were measured, and allotetraploid plants were selected. Optionally, a step of further propagating the allotetraploid offspring of the allotetraploid plant. A grafted plant according to Embodiment 1, obtained by a method comprising the above.

[0183] 3. The aforementioned heterotetraploid rootstock Interspecific protoplast fusion was performed between the protoplast of a first Solanum lycopersicum variety and the protoplast of a stress-tolerant variety of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the seeds produced by the above cross were collected. We regenerate heterotetraploid tomato plants from fused protoplasts, self-pollinate the resulting plants, and grow heterotetraploid tomato plants from the resulting seeds. The DNA content and chromosome set of the obtained plants were measured, and further Optionally, a step of further propagating the allotetraploid offspring of the allotetraploid plant. A grafted plant according to Embodiment 1, obtainable by a method including the following.

[0184] 4. The plant according to any one of Embodiments 1 to 3, wherein the stress tolerance includes abiotic stress tolerance selected from the group consisting of cold tolerance, heat tolerance, drought tolerance, and salt tolerance.

[0185] 5. The plant according to Embodiment 4, wherein the stress-tolerant variety exhibits salt tolerance.

[0186] 6. The stress-tolerant varieties are LA0089, LA0130, LA0166, LA0247, LA0317, LA0376, LA0400, LA0407, LA0421, LA0426, LA0438, LA0443, LA0453, LA0462, LA0522, LA0716, LA0751, LA1221, LA1237, LA1257, LA1278, LA1294, LA1301, LA1306, LA1310, LA1316, LA1325, LA1331, LA1340, LA1351, LA1352, LA1357, LA1363, LA1367, LA1373, LA1383, LA1392, LA1401, LA1408, LA1421, LA1449, LA1572, LA1579, LA1589, LA1590, LA1609, LA1610, LA1629, LA1630, LA1646, LA1648, LA1674, LA1676, LA1694, LA1722, LA1777, LA1778, LA1809, LA1923, LA1926, LA1930, LA1932, LA1950, LA1958, LA1959, LA1961, LA1962, LA1969, LA1971, LA1972, LA1974, LA1986, LA2079, LA2080, LA2081, LA2149, LA2150, LA2157, LA2163, LA2182, LA2309, LA2310, LA2311, LA2327, LA2403, LA2408, LA2425, LA2560, LA2661, LA2662, LA2710, LA2711, LA2744, LA2747, LA2748, LA2755, LA2773, LA2876, LA2877, LA2878, LA2880, LA2884, LA2885, LA2931, LA2963, LA2964, LA3120, LA3153, LA3320, LA3465, LA3799, LA3847, LA4023, LA4105, LA4108, LA4133, LA4321, LA4324, LA4330, LA4335, Arkansas Traveler, Delicious, Edkawi, German Johnson Pink, Golden Princess, Heinz 1370, Homestead, New Yorker, RAF, Ramsi, and St.A plant according to Embodiment 4, selected from the group consisting of Pierre.

[0187] 7. The plant according to any one of Embodiments 1 to 3, wherein the stress tolerance includes biological stress tolerance selected from the group consisting of disease resistance, pest and disease resistance, bacterial resistance, fungal resistance, insect resistance, and nematode resistance.

[0188] 8. The stress-tolerant variety is a plant according to Embodiment 7 selected from the group consisting of LA0490, LA0655, LA0656, LA1783, LA1791, LA1792, LA1800, LA1802, LA1964, LA1995, LA2009, LA2356, LA2369, LA2396, LA2443, LA2444, LA2445, LA2446, LA2447, LA2448, LA2449, LA2458, LA2530, LA2531A, LA2531B, LA2531C, LA2701, LA2818, LA2819, LA2820, LA2821, LA2822, LA2823, LA2824, LA2825, LA2826, LA2827, LA2828, LA2829, LA2830, LA2968, LA3025, LA3026, LA3027, LA3028, LA3029, LA3038, LA3039, LA3040, LA3041, LA3042, LA3043, LA3044, LA3045, LA3046, LA3047, LA3048, LA3049, LA3050, LA3051, LA3118, LA3129, LA3130, LA3145, LA3151, LA3152, LA3158, LA3159, LA3160, LA3161, LA3201, LA3202, LA3213, LA3214, LA3215, LA3216, LA3254, LA3258, LA3268, LA3269, LA3271, LA3273, LA3275, LA3276, LA3277, LA3292, LA3297, LA3309, LA3310, LA3313, LA3314, LA3341, LA3342, LA3428, LA3432, LA3433, LA3471, LA3472, LA3473, LA3526, LA3667, LA3839, LA3845, LA3846, LA3847, LA3856, LA3858, LA3859, LA3912, LA4025, LA4026, LA4285, LA4286, LA4441, LA4442, Arkansas Traveler, Delicious, Edkawi, German Johnson Pink, Golden Princess, Heinz 1370, Homestead, New Yorker, RAF, Ramsi, and St. Pierre.

[0189] 9. The plant according to any of the prior embodiments, wherein the stress-tolerant variety is selected from the group of (i) varieties having at least 80% homozygosity, and (ii) varieties that strictly or preferentially self-pollinate.

[0190] 10. The plant according to any of the prior embodiments, wherein the Solanum lycopersicum variety is a commercially available variety selected based on one or more of the following criteria: overall performance, robustness, high seed yield, vigor, and regional adaptability.

[0191] 11. A plant according to any of the prior embodiments, wherein the chromosome doubling agent is colchicine.

[0192] 12. Seeds for growing hybrid plants, wherein the seeds are (i) a. A first allotetraploid plant is formed from a first Solanum lycopersicum variety and a first stress-tolerant variety of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites. b. A second allotetraploid plant is crossed with a second stress-tolerant variety of a second Solanum lycopersicum variety, selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites. (The first stress-tolerant variety and the second stress-tolerant variety are different, and the first variety provides at least one tolerance to at least one stressor that is not provided by the second stress-tolerant variety.) (ii) Seeds for growing the hybrid plant, produced by collecting the seeds obtained by the aforementioned cross.

[0193] 13. Seeds according to Embodiment 12, wherein one of the stress-tolerant varieties is stress-tolerant to abiotic stress, and the other stress-tolerant variety is stress-tolerant to abiotic stress.

[0194] 14. The seed according to Embodiment 12 or 13, wherein the first and / or second heterotetraploid is established by the method described in Embodiment 2 or the method described in Embodiment 3.

[0195] 15. A hybrid plant or a part thereof grown from a seed according to any of embodiments 12 to 14.

[0196] 16. Non-natural grafted plants, The heterotetraploid hybrid plant described in Embodiment 15 as the rootstock, and At least one scion of a Solanum lycopersicum variety grafted onto the aforementioned rootstock The aforementioned non-natural grafted plant, consisting of the above.

[0197] 17. A method for producing grafted plants that have stress tolerance, wherein the method is We provide heterotetraploid rootstocks consisting of a first Solanum lycopersicum variety and stress-tolerant varieties of species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites. We provide cuttings of a second Solanum lycopersicum variety as scions. The step of grafting the scion onto the rootstock, thereby producing the grafted plant. The production method, including the above.

[0198] 18. The aforementioned heterotetraploid rootstock Interspecific crosses were performed between a Solanum lycopersicum variety and stress-tolerant varieties of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the seeds produced by the crosses were collected. Hybrid plants are grown from the aforementioned seeds, and cuttings taken from the aforementioned hybrid plants are treated with a chromosome doubling agent. The processed cuttings are allowed to grow, the resulting plants are self-pollinated, and the seeds are grown to produce plants. The DNA content and chromosome set of the obtained plants were measured, and allotetraploid plants were selected. Optionally, a step of further propagating the allotetraploid offspring of the allotetraploid plant. The method according to Embodiment 17, produced by a method comprising:

[0199] 19. The aforementioned heterotetraploid rootstock Interspecific protoplast fusion was performed between protoplasts of a Solanum lycopersicum variety and protoplasts of a stress-tolerant variety of wild tomato selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the seeds produced by the aforementioned cross were collected. We regenerate tetraploid tomato plants from fused protoplasts, self-pollinate the resulting plants, and grow heterotetraploid tomato plants from the resulting seeds. The DNA content and chromosome set of the obtained plants were measured, and further Optionally, a step of further propagating the allotetraploid offspring of the allotetraploid plant. The method according to Embodiment 18, which can be obtained by a method including the following.

[0200] 20. The method according to Embodiment 19, wherein mitochondria and / or chloroplasts are provided only by the Solanum lycopersicum variety.

[0201] 21. The method according to Embodiment 19, wherein mitochondria and / or chloroplasts are provided only by the wild tomato variety.

[0202] 22. The method according to Embodiment 19, wherein the nucleus is provided only by the Solanum lycopersicum variety.

[0203] 23. The method according to Embodiment 19, wherein the kernels are provided only by the wild tomato variety.

[0204] 24. The method according to any one of embodiments 18 to 23, wherein the stress tolerance includes abiotic stress tolerance selected from the group consisting of cold tolerance, high temperature tolerance, drought tolerance, and salt tolerance.

[0205] 25. The method according to any one of embodiments 18 to 23, wherein the stress tolerance includes biological stress tolerance selected from the group consisting of disease resistance, fungal resistance, pest and disease resistance, bacterial resistance, insect resistance, and nematode resistance.

[0206] 26. The method according to Embodiment 18, wherein the chromosome doubling agent is colchicine.

[0207] 27. A method for producing seeds for producing hybrid plants, (i) a. A first allotetraploid plant comprising a first Solanum lycopersicum variety and a first stress-tolerant variety of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, b. A second allotetraploid plant consisting of a second Solanum lycopersicum variety and a second stress-tolerant variety of a species selected from the group consisting of S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites is crossed. (The first stress-tolerant variety and the second stress-tolerant variety are different, and the first variety provides at least one tolerance to at least one stressor that is not provided by the second stress-tolerant variety.) (ii) A step of collecting seeds from the cross product. The production method, including the above.

[0208] 28. A method for producing non-natural grafted plants, wherein the method is A rootstock obtainable by the method described in Embodiment 27 is provided. Provide at least one scion of the Solanum lycopersicum variety, The step of grafting the at least one scion onto the rootstock. The production method, including the above.

[0209] 29. A stress-tolerant allotetraploid rootstock containing chromosomes derived from Solanum lycopersicum, and at least one stress-tolerant species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and At least one scion of a Solanum lycopersicum variety grafted onto the aforementioned heterotetraploid rootstock. A complex tomato plant, including [this plant].

[0210] 30. A compound tomato plant comprising a scion grafted onto an allotetraploid rootstock, wherein the allotetraploid rootstock is obtained from the seeds or plant tissue of a tomato allotetraploid rootstock variety described herein.

[0211] 31. The composite tomato plant according to Embodiment 29 or 30, wherein the heterotetraploid rootstock has abiotic stress tolerance selected from the group consisting of cold tolerance, high temperature tolerance, drought tolerance, and salt tolerance.

[0212] 32. The composite tomato plant according to Embodiment 29 or 30, wherein the stress-tolerant allotetraploid rootstock has biological stress tolerance selected from the group consisting of disease resistance, fungal resistance, pest and disease resistance, bacterial resistance, insect resistance, and nematode resistance.

[0213] 33. A non-natural grafted plant according to Embodiment 1, or a compound tomato plant according to Embodiment 29 or 30, wherein the scion is a Solanum lycopersicum variety selected from the group consisting of Sugarino, Super Sweet, QualiT-99, QuailT-27, Mision, Dorma, Midelyce, Badiaa F1, Volantis, DR7024TS, Marvelance, Juanita, Ramsi, RAF, Edkawi, Golden Princess, and St. Pierre.

[0214] 34. A method for producing commercially available Solanum lycopersicum plant products, A Solanum lycopersicum variety was grafted onto a stress-tolerant allotetraploid rootstock containing chromosomes derived from Solanum lycopersicum and at least one stress-tolerant species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites to produce a compound tomato plant. The aforementioned compound tomato plant is grown, Harvesting commercially available Solanum lycopersicum plant products The production method, including the above.

[0215] 35. The method according to Embodiment 34, wherein the rootstock of the stress-tolerant heterotetraploid has stress tolerance selected from the group consisting of cold tolerance, high temperature tolerance, drought tolerance, salt tolerance, disease resistance, fungal resistance, insect resistance, and nematode resistance.

[0216] 36. The method according to Embodiment 34, wherein the stress-tolerant allotetraploid rootstock is obtained from the seeds or plant tissue of the tomato allotetraploid rootstock variety described herein.

[0217] 37. The method according to Embodiment 34, wherein the plant product is selected from tomato fruit, tomato seeds, and cuttings.

[0218] 38. Seeds of an allotetraploid tomato referred to as "MX20-06", wherein a sample of the allotetraploid tomato seeds has been deposited under NCMA accession number 202303001.

[0219] 39. Rootstock for stress-tolerant allotetraploid tomato containing chromosomes derived from Solanum lycopersicum, as well as at least one stress-tolerant species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites.

[0220] 40. A method for producing stress-tolerant heterotetraploid tomato plants, Interspecific hybrid seeds are produced by crossing Solanum lycopersicum varieties with stress-tolerant varieties selected from the Esculentum, Arcanum, Peruvianum, Hirsutum, Lycopersicoides sections, and their hybrid combinations. The aforementioned interspecific hybrid seeds are grown to produce interspecific hybrid plants. The interspecific hybrid plant or its cuttings are sprayed with a chromosome doubling agent to produce a chimeric interspecific hybrid. The chimeric interspecific hybrid is grown to produce tomato fruit. Seeds were collected from the aforementioned tomato fruits. The aforementioned seeds are allowed to grow, Selecting stress-tolerant heterotetraploid tomato plants. The production method, including the above.

[0221] 41. A method for producing stress-tolerant heterotetraploid tomato plants, Protoplasts isolated from S. lycopersicum were fused with another protoplast isolated from a stress-tolerant Solanaceae species. Select a heterokalion, To regenerate heterotetraploid tomato plants from the aforementioned heterokalion. The production method, including the above.

[0222] 42. The method according to Embodiment 40 or 41, wherein the stress-tolerant species is derived from the Esculentum group and selected from S. galapagense, S. cheesemaniae, S. lycopersicum, and S. pimpinellifolium.

[0223] 43. The method according to Embodiment 40 or 41, wherein the stress-tolerant species is derived from the Arcanum group and selected from S. neorickii, S. arcanum, and S. chmielewskii.

[0224] 44. The method according to Embodiment 40 or 41, wherein the stress-tolerant species is derived from the Peruvianum group and selected from S. huaylasense, S. peruvianum, S. corneliomulleri, and S. chilense.

[0225] 45. The method according to Embodiment 40 or 41, wherein the stress-tolerant species is derived from the Hirsutum group and selected from S. habrochaites and S. pennellii.

[0226] 46. ​​The method according to Embodiment 40 or 41, wherein the stress-tolerant species is derived from the section Lycopersicoides and selected from S. lycopersicoides and S. sitiens.

[0227] 47. The method according to Embodiment 40, wherein the chromosome doubling agent is an antimitotic agent selected from colchicine, trifluralin, oryzalin, and amiprophos-methyl.

[0228] 48. The method according to embodiment 40 or 41, wherein the stress-tolerant species has abiotic stress tolerance selected from the group consisting of cold tolerance, heat tolerance, drought tolerance, and salt tolerance.

[0229] 49. The method according to embodiment 40 or 41, wherein the stress-tolerant species has biotic stress tolerance selected from the group consisting of disease resistance, fungal resistance, pest and disease resistance, bacterial resistance, insect resistance, and nematode resistance.

[0230] 50. The method according to embodiment 40, wherein the S. lycopersicum is the acceptor.

[0231] 51. The method according to embodiment 40, wherein the Solanaceae species is sexually incompatible with S. lycopersicum.

[0232] 52. The method according to embodiment 40, wherein the protoplast fusion is asymmetric.

[0233] 53. A method for producing stress-tolerant allopolyploid hybrid plants or seeds, comprising: crossing a Solanum lycopersicum variety with a stress-tolerant species selected from Esculentum, Arcanum, Peruvianum, Hirsutum, and their hybrid combinations to produce interspecific hybrid seeds; growing the interspecific hybrid seeds to produce an interspecific hybrid plant; subjecting the interspecific hybrid plant or its cutting to a chemical treatment for chromosome doubling to generate a chimeric interspecific hybrid; growing the chimeric interspecific hybrid to produce tomato fruits; collecting seeds from the tomato fruits; growing the seeds and selecting stress-tolerant allopolyploid tomato plants; crossing the allopolyploid tomato plant with itself or with a second allopolyploid plant to produce hybrid allopolyploid tomato seeds. The aforementioned hybrid heterotetraploid tomato seeds were harvested, and further The method involves optionally growing the aforementioned hybrid allotetraploid tomato seeds to produce stress-tolerant hybrid allotetraploid plants. The production method, including the above.

[0234] 54. A chimeric plant tissue comprising a first plant cell and a second plant cell, wherein the first plant cell is an allotetraploid comprising chromosomes derived from Solanum lycopersicum and chromosomes derived from at least one species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the second plant cell is a diploid Solanum lycopersicum.

[0235] 55. A method for conferring desirable traits derived from wild species of tomato to cultivated Solanum lycopersicum varieties, The conferral method comprises grafting a cultivated Solanum lycopersicum variety onto an allotetraploid rootstock having a desired trait, wherein the allotetraploid rootstock contains chromosomes derived from Solanum lycopersicum and at least one stress-tolerant species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites.

[0236] 56. The method according to Embodiment 55, wherein the desired trait is tolerance to biological or abiotic stress.

[0237] 57. Seeds of an allotetraploid tomato referred to as "MX20-15," wherein a sample of the allotetraploid tomato seeds has been deposited under NCMA accession number 202401032.

[0238] 58. A method for producing unnaturally stress-tolerant complex tomato plants, We provide stress-tolerant heterotetraploid tomato plants as rootstock. We provide the Solanum lycopersicum variety as the scion. The scion is grafted onto the rootstock, thereby producing a non-naturally stress-tolerant complex tomato plant. The production method, including the above.

[0239] 59. The method according to Embodiment 58, wherein the rootstock comprises chromosomes derived from Solanum lycopersicum and chromosomes derived from at least one species selected from the Esculentum group, Arcanum group, Pervianum group, Hirsutum group, Lycopersicoides section, and hybrid combinations thereof.

[0240] 60. The method according to Embodiment 58 or 59, wherein the scion is a commercially available variety.

[0241] 61. The stress-tolerant allotetraploid plant, A plant derived from a Solanum lycopersicum variety and a stress-tolerant Solanaceae species, wherein the Solanum lycopersicum variety and the plant derived from the stress-tolerant Solanaceae species are essentially homozygous, and the stress-tolerant Solanaceae species is selected from the Esculentum group, Arcanum group, Peruvianum group, Hirsutum group, Lycopersicoides section, and hybrid combinations thereof, and interspecific hybrid seeds are produced by crossing the Solanum lycopersicum variety and the plant derived from the stress-tolerant Solanaceae species. The aforementioned interspecific hybrid seeds are grown to produce interspecific hybrid plants. The interspecific hybrid plant or its cuttings are sprayed with a chromosome doubling agent to produce a chimeric interspecific hybrid. The chimeric interspecific hybrid is grown to produce tomato fruit. Seeds were collected from the aforementioned tomato fruits. To grow the aforementioned seeds and produce stress-tolerant allotetraploid tomato plants. The method according to any one of embodiments 58 to 60, produced by...

[0242] 62. The stress-tolerant heterotetraploid tomato plant described above, Protoplasts isolated from Solanum lycopersicum were fused with other protoplasts isolated from stress-tolerant Solanaceae species. We isolated the heterokaryon, To regenerate heterotetraploid tomato plants from the aforementioned heterokalion. The method according to any one of embodiments 58 to 60, produced by...

[0243] 63. The method according to Embodiment 58 or 62, wherein the stress-tolerant Solanaceae species is selected from the Esculentum group, Arcanum group, Peruvianum group, Hirsutum group, Lycopersicoides section, and hybrid combinations thereof.

[0244] 64. The method according to Embodiment 58 or 62, wherein the stress-tolerant Solanaceae species is derived from the Esculentum group and selected from S. galapagense, S. cheesemaniae, S. lycopersicum, S. pimpinellifolium, and hybrid combinations thereof.

[0245] 65. The method according to embodiment 58 or 62, wherein the stress-tolerant Solanaceae species is derived from the Arcanum group and is selected from S. neorickii, S. arcanum, S. chmielewskii, and their hybrid combinations.

[0246] 66. The method according to embodiment 58 or 62, wherein the stress-tolerant Solanaceae species is derived from the Peruvianum group and is selected from S. huaylasense, S. peruvianum, S. corneliomulleri, S. chilense, and their hybrid combinations.

[0247] 67. The method according to embodiment 58 or 62, wherein the stress-tolerant Solanaceae species is derived from the Hirsutum group and is selected from S. habrochaites, S. pennellii, and their hybrid combinations.

[0248] 68. The method according to embodiment 58 or 62, wherein the stress-tolerant Solanaceae species is derived from the Lycopersicoides section and is selected from S. lycopersicoides, S. sitiens, and their hybrid combinations.

[0249] 69. The method according to embodiment 58 or 62, wherein the stress-tolerant Solanaceae species has abiotic stress tolerance selected from the group consisting of cold tolerance, heat tolerance, drought tolerance, and salt tolerance.

[0250] 70. The method according to embodiment 58 or 62, wherein the stress-tolerant Solanaceae species has biotic stress tolerance selected from the group consisting of disease resistance, fungal resistance, pest and disease resistance, bacterial resistance, insect resistance, and nematode resistance.

[0251] 71. The method according to any one of embodiments 62 to 70, wherein the protoplast fusion is asymmetric and the S. lycopersicum protoplast is the acceptor.

[0252] 72. A non-natural compound tomato plant produced by the method described in any one of Embodiments 58 to 71.

[0253] 73. The non-natural compound tomato plant according to Embodiment 72, wherein the heterotetraploid rootstock contains at least one allele derived from a scion variety of Solanum lycopersicum.

[0254] 74. A method for producing hybrid allotetraploid tomato seeds for producing hybrid allotetraploid plants, wherein the method is A first stress-tolerant allotetraploid tomato plant is crossed with a second stress-tolerant allotetraploid tomato plant containing chromosomes derived from at least one species different from the first allotetraploid tomato plant. Steps to collect the aforementioned hybrid heterotetraploid tomato seeds. The production method comprising the first stress-tolerant allotetraploid tomato plant and the second stress-tolerant allotetraploid tomato plant being essentially homozygous.

[0255] 75. The method according to Embodiment 74, wherein the first or second stress-tolerant allotetraploid tomato plant comprises chromosomes derived from Solanum lycopersicum, as well as at least one species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, S. habrochaites, and their hybrids.

[0256] 76. The method according to Embodiment 74 or 75, wherein the first allotetraploid tomato plant provides at least one tolerance to at least one stressor that is not provided by the second allotetraploid tomato plant, and the stress tolerances provided by the first allotetraploid plant and the second allotetraploid plant are complementary.

[0257] 77. The method according to any one of embodiments 74 to 76, wherein the first stress-tolerant allotetraploid tomato plant has abiotic stress tolerance selected from the group consisting of cold tolerance, heat tolerance, drought tolerance, and salt tolerance.

[0258] 78. The method according to any one of embodiments 74 to 77, wherein the second stress-tolerant heterotetraploid tomato plant has biological stress tolerance selected from the group consisting of disease resistance, fungal resistance, pest and disease resistance, bacterial resistance, insect resistance, and nematode resistance.

[0259] 79. A hybrid allotetraploid tomato plant produced by growing a hybrid allotetraploid tomato seed according to any one of Embodiments 74 to 78.

[0260] 80. A non-natural compound tomato plant comprising a Solanum lycopersicum scion grafted onto a hybrid allotetraploid rootstock, wherein the hybrid allotetraploid rootstock is produced from the plant described in Embodiment 79.

[0261] 81. The non-natural compound tomato plant according to Embodiment 80, wherein the hybrid heterotetraploid rootstock contains at least one allele derived from a scion variety of Solanum lycopersicum.

[0262] 82. A hybrid allotetraploid tomato plant comprising chromosomes derived from Solanum lycopersicum, and chromosomes derived from at least one species selected from S. pimpinellifolium, S. cheesemaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, S. habrochaites, and their hybrids, wherein the hybrid allotetraploid tomato plant provides at least one tolerance to at least one stressor that is not provided by one of its parent lines.

Claims

1. A method for producing unnaturally stress-tolerant complex tomato plants, We provide stress-tolerant heterotetraploid tomato plants as rootstock. We provide the Solanum lycopersicum variety as the scion. The scion is grafted onto the rootstock, thereby producing a non-naturally stress-tolerant complex tomato plant. The production method, including the above.

2. The method according to claim 1, wherein the rootstock comprises a chromosome derived from Solanum lycopersicum and a chromosome derived from at least one species selected from the Esculentum group, Arcanum group, Pervianum group, Histutum group, Lycopersicoides section, and hybrid combinations thereof.

3. The method according to claim 1, wherein the scion is a commercially available variety.

4. The aforementioned stress-tolerant allotetraploid plant, A plant derived from a Solanum lycopersicum variety and a stress-tolerant Solanaceae species, wherein the plant derived from the Solanum lycopersicum variety and the stress-tolerant Solanaceae species is essentially homozygous, and the stress-tolerant Solanaceae species is selected from the Esculentum group, Arcanum group, Peruvianum group, Hirsutum group, Lycopersicoides section, and hybrid combinations thereof, and interspecific hybrid seeds are produced by crossing the plant derived from the Solanum lycopersicum variety and the stress-tolerant Solanaceae species. The aforementioned interspecific hybrid seeds are grown to produce interspecific hybrid plants. The interspecific hybrid plant or its cuttings are sprayed with a chromosome doubling agent to produce a chimeric interspecific hybrid. The chimeric interspecific hybrid is grown to produce tomato fruit. Seeds were collected from the aforementioned tomato fruits. To grow the aforementioned seeds and produce stress-tolerant allotetraploid tomato plants. The method according to claim 1, produced by

5. The aforementioned stress-tolerant heterotetraploid tomato plant, Protoplasts isolated from Solanum lycopersicum were fused with other protoplasts isolated from stress-tolerant Solanaceae species. We isolated the heterokaryon, The method according to claim 1, which is produced by regenerating heterotetraploid tomato plants from the heterokaryon.

6. The method according to claim 4, wherein the stress-tolerant Solanaceae species is selected from the Esculentum group, Arcanum group, Peruvianum group, Hirsutum group, Lycopersicoides section, and hybrid combinations thereof.

7. The method according to claim 4, wherein the stress-tolerant Solanaceae species is derived from the Esculentum group and selected from S. galapagense, S. cheesmaniae, S. lycopersicum, S. pimpinellifolium, and hybrid combinations thereof.

8. The method according to claim 4, wherein the stress-tolerant Solanaceae species is derived from the Arcanum group and selected from S. neorickii, S. arcanum, S. chmielewskii, and hybrid combinations thereof.

9. The method according to claim 4, wherein the stress-tolerant Solanaceae species is derived from the Peruvianum group and selected from S. huaylasense, S. peruvianum, S. corneliomulleri, S. chilense, and hybrid combinations thereof.

10. The method according to claim 4, wherein the stress-tolerant Solanaceae species is derived from the Hirsutum group and selected from S. habrochaites, S. pennellii, and hybrid combinations thereof.

11. The method according to claim 4, wherein the stress-tolerant Solanaceae species is derived from the Lycopersicoides section and selected from S. lycopersicoides, S. sitiens, and hybrid combinations thereof.

12. The method according to claim 4, wherein the stress-tolerant Solanaceae species has abiotic stress tolerance selected from the group consisting of cold tolerance, high temperature tolerance, drought tolerance, and salt tolerance.

13. The method according to claim 4, wherein the stress-tolerant Solanaceae species has biological stress tolerance selected from the group consisting of disease resistance, fungal resistance, pest and disease resistance, bacterial resistance, insect resistance, and nematode resistance.

14. The method according to claim 5, wherein the protoplast fusion is asymmetric and the S. lycopersicum protoplast is an acceptor.

15. A non-natural compound tomato plant produced by the method described in claim 1.

16. The non-natural composite tomato plant according to claim 15, wherein the heterotetraploid rootstock contains at least one allele derived from a scion variety of Solanum lycopersicum.

17. A method for producing hybrid allotetraploid tomato seeds for producing hybrid allotetraploid plants, wherein the method is A first stress-tolerant allotetraploid tomato plant is crossed with a second stress-tolerant allotetraploid tomato plant containing chromosomes derived from at least one species different from the first allotetraploid tomato plant. Steps to collect the aforementioned hybrid heterotetraploid tomato seeds. Includes, The production method wherein the first stress-tolerant allotetraploid tomato plant and the second stress-tolerant allotetraploid tomato plant are essentially homozygous.

18. The method according to claim 17, wherein the first or second stress-tolerant allotetraploid tomato plant comprises chromosomes derived from Solanum lycopersicum and at least one species selected from S. pimpinellifolium, S. cheesmaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, S. habrochaites, and hybrids thereof.

19. The method according to claim 17, wherein the first allotetraploid tomato plant provides at least one tolerance to at least one stressor that is not provided by the second allotetraploid tomato plant, and the stress tolerances provided by the first allotetraploid plant and the second allotetraploid plant are complementary.

20. The method according to claim 17, wherein the first stress-tolerant heterotetraploid tomato plant has abiotic stress tolerance selected from the group consisting of cold tolerance, high temperature tolerance, drought tolerance, and salt tolerance.

21. The method according to claim 17, wherein the second stress-tolerant heterotetraploid tomato plant has biological stress tolerance selected from the group consisting of disease resistance, fungal resistance, pest and disease resistance, bacterial resistance, insect resistance, and nematode resistance.

22. A hybrid allotetraploid tomato plant produced by growing the hybrid allotetraploid tomato seeds described in claim 17.

23. A non-natural compound tomato plant comprising a Solanum lycopersicum scion grafted onto a hybrid allotetraploid rootstock, wherein the hybrid allotetraploid rootstock is produced from the plant described in claim 22.

24. The non-natural composite tomato plant according to claim 23, wherein the hybrid heterotetraploid rootstock contains at least one allele derived from a Solanum lycopersicum scion variety.

25. A chimeric plant tissue comprising a first plant cell and a second plant cell, wherein the first plant cell is an allotetraploid comprising chromosomes derived from Solanum lycopersicum and chromosomes derived from at least one species selected from S. pimpinellifolium, S. cheesmaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, and S. habrochaites, and the second plant cell is a diploid Solanum lycopersicum.

26. A hybrid allotetraploid tomato plant comprising chromosomes derived from Solanum lycopersicum, and chromosomes derived from at least one species selected from S. pimpinellifolium, S. cheesmaniae, S. galapagense, S. pennellii, S. peruvianum, S. chilense, S. chmielewskii, S. corneliomulleri, S. sitiens, S. habrochaites, and hybrids thereof, wherein the hybrid allotetraploid tomato plant provides at least one tolerance to at least one stressor that is not provided by one of its parent lines.

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