JP5954545B2 - Intergeneric hybrid plant and production method of Argyranthemum - Google Patents

Description

  [0001] The present invention relates to hybrid plants between the genus Argyranthemum and methods for increasing the production efficiency of hybrid plants. More specifically, the present invention relates to the production of tetraploid and aneu-tetraploid Argyranthemum plants, as well as Argyranthemum, which is a tetraploid or aneuploid tetraploid plant, and Ismeria. The invention relates to the production of intergeneric hybrid plants obtained from crossing with plants of the group of Ismeliaversiccolor and Grebionis species. All publications cited are hereby incorporated by reference.

  [0002] A particular plant feature is the ability to cross with other species occasionally, referred to as interspecies hybridization. Interspecies hybridization has been identified in several species, including Argyranthemum. For example, in Argyranthemum, many species have been reported to cross naturally when the geographical barrier to pollination is removed (Francisco-Ortega, J., Santos-Guerra, A., Mesa-Coello). , R., Gonzalez-Feria, E. and Crawford, D., Genetic resource conservation of the endogenous genus Argyranthaneum Sch. Bip. (Asteracee: Anthemide). Year)). With a wide range of flower colors available in commercially bred varieties of Argyranthemum, several species of Argyranthemum have been published by Cunnenen, T. et al. M.M. TheMarguerite Daisy (Argyranthemum spp): developing an underending for bleeding, Ph. D. It has been suggested that it was involved in the development of modern varieties reported by Thesis, University of Science Facility of Agriculture (1996). Thus, modern varieties are best described as Argyranthemum x Hybrid. All Argyranthemum species are found in Humphries, C.I. J. et al. A revision of the Macronesian genus Argyranthemum Webb ex Schults Bip. (Compositee-Anthideidee), Bulletinof the British Museum (Natural History), Boston, 5: 145-243 (1976) and Fjellheim, S .; HoltenJorgensen, M .; Kjos, M .; Borgen, L .; , A molecular study of hybridization and homoidhybrid speciation in Arg. 1 to 19 (2), which is reported in Argyranthemum (Asteraceae) on Ten9, theLand, Of diploid chromosomes.

  [0003] Over time, plants have been more accurately described and investigated by taxonologists, which have changed taxonomies and species names. In the genus Grebionis, Maberley, D. et al. J. et al. According to Mamberley's Plant Book, Cambridge University Press (2008), G. Coronaria and G. There are two types of segetum. However, these species have also been historically included in Chrysanthhemum and Xanthopthalum. Ismelia currently contains Only one versicolol is present. Historically, this species is known as Chrysanthem carinatum, Glebionis carinatum and Ismeria versicolol. In order to avoid confusion, in the present application, Grevionis belongs to G. Coronaria and G. Two species of segetam are included. The 2008 Maberley convention of inclusion of one of versicolol applies.

  [0004] Genetic complexity affects the choice of breeding methods. Backcross breeding is used to transfer one or several genes suitable for highly heritable traits to the desired variety. This technique is widely used for breeding varieties resistant to diseases. Various cyclic selection techniques are used to quantitatively improve the inheritance controlled by a number of genes. The use of circular selection in self-pollinated crops depends on the ease of pollination, the frequency of successful hybrids from each pollination, and the number of hybrid progeny from each successful cross.

  [0005] Backcross breeding has been used to transmit traits that follow simple Mendelian inheritance to desirable homozygous varieties or inbred lines that are circulating parents. The source of the transmitted trait is called the donor parent. The resulting plant is expected to have the attributes of the circulating parent (eg, variety) and the desired traits transmitted from the donor parent. After the first cross, individuals with the donor parent phenotype are selected and recrossed back to the circulating parent (backcross). The resulting plant is expected to have the attributes of the circulating parent (eg, variety) and the desired traits transmitted from the donor parent.

[0006] Line breeding is commonly used to improve self-pollinated crops. Two parents with suitable complementary traits are crossed to produce the F ₁ population. F ₂ population is produced by one or more F ₁ plants be selfed. Selection of the best individuals may begin in the F ₂ population. Then, the best individuals in the best families (families) are selected at the beginning of the F ₃ generation. The replicates of the family starting at F ₄ generation, can improve the effectiveness of the traits of selection with low heritability. Advanced stage of inbreeding (i.e., F ₆ and F ₇ generation), the best lines or phenotypes what a mix of strains that are similar to test for potential sales potential as new varieties.

  [0007] Line breeding methods and cyclic selection breeding methods are used to develop varieties from breeding populations. A breeding program incorporates desirable traits from two or more varieties or a wide variety of sources into a breeding pool and then develops the varieties by self-pollination and selection of the desired phenotype. Evaluate new varieties to determine which have commercially possible potential.

  [0008] Population selection and circular selection can be used to improve either self-pollinated or cross-pollinated crop populations. A genetically variable population of heterozygous individuals is identified or created by crossing several different parents. The best plants are selected on the basis of individual dominance, outstanding progeny, or excellent combinatorial capabilities. The selected plants are crossed to produce a new population and then continue the further selection cycle.

  [0009] A description of other breeding methods commonly used for different traits and crops can be found in one of several references (eg, Allard, RW, Principlesof Plant Breeding, John Wiley and). Sons Inc. (1960); Simmonds, N.W., Principles of Crop Improvment, Longman Group, New York, USA (1981)).

  [0010] Each breeding program should include a periodic objective assessment of the efficiency of the breeding procedure. Evaluation criteria will vary depending on purpose and goal, but appropriate standards, overall value of improved breeding lines, and success of production varieties per unit of input (eg per year, per expenditure, etc.) It should include profits per year from selection, based on a comparison of numbers.

  [0011] Promising improved breeding lines are thoroughly tested and compared to appropriate standards for over 3 years in an environment representative of the commercial target area (s). The best line is a candidate for a new commercial variety. Those that still lack some traits can be used as parents to produce new populations for further selection.

  [0012] These processes leading to the final stage of marketing and distribution require several steps from the time of the initial cross. Thus, the development of new varieties is a time consuming process that requires accurate future planning, efficient use of funds, and minimal turnaround.

  [0013] For most traits, the most difficult task is to identify genetically superior individuals, because the true value of the genotype is masked by other confounding plant traits or environmental factors is there. One way to identify a good plant is to observe its performance compared to other experimental plants and / or common varieties. If a conclusion is not reached in a single observation, the genetic value can be better estimated by repeating the observation.

  [0014] Interspecies hybridization has allowed the creation of new forms of plants and the transfer of desirable characteristics from one species to another, for example by gene transfer from wild species to related cultivated species. However, the ability of any two species to create viable interspecific hybrid seeds or plants is unpredictable and often proven to be impossible.

  [0015] Intergeneric hybridization, which is a cross between two plants between different genera, is less predictable than interspecific hybridization because the relative genetic distance is greater between genera than between species Less likely. Only a few successful intergeneric hybrids have been reported, but they are often only possible through the use of human intervention and embryo rescue. One form of embryo rescue is ovule culture, in which ovules are aseptically removed from seeds and placed on an artificial medium to germinate embryos and grow on plants. In Argylansem, the intergeneric hybrid is a female diploid A. Flutescens, and G. a male diploid. A. is a female diploid with and between carinatam (also known as I. versicolol). G. fullness and male diploid. These have been reported with coronaria, all of which have been developed by ovule culture (Ohtsuka, H. and Inaba, Z., Intergenerhybridization of marguerite (Argyranumumumranthrum frenscens) with anthracnum) Coronaria) using ocular culture, Plant Biotechnology, 25, 535-539 (2008); Ohtsuka, H. and Inaba, Z., Breedingof Argythenium by interspecificity. idization.1.Intergenerichybridization of Argyranthemum and Ismeria carinata (syn.Chrysanthemum carinatum), I.coronaria (syn.Chrysanthemumcoronaria) through ovule culture, Journal of the Japanese Society for HorticulturalScience, 72 (Suppl.1), 264 pp. (2003) Iwaki, Y., Ueda, Y. and Yamada, H., Studies on the acquisition method of an intergeneral hybridization of Ar yranthemum and Ismelia by ovule culture, HorticulturalResearch (Japan), 6 (Suppl.1), 212 pp. (2007)). However, the efficiency (number of pollinations performed vs. number of flowering plants produced) and the quality of the plants produced are very low.

  [0016] For example, Ohtsuka and Inaba (2008) Fullness x G. It was reported that only 16 embryos were obtained from 70 pollinations of carinatam (also known as I. versicolol), which were germinated by ovule culture, of which only 5 flowering plants developed. These five plants are G. It had a similar form to carinatam (also known as I. versicolol). However, after flowering, two died and the remaining three had light green leaves and showed fragile growth. Ohtsuka and Inaba (2008) describe A.S. Fullness x G. It was also reported that only 26 embryos were obtained from 61 pollinations of coronaria, which were germinated by ovule culture, of which only 16 flowering plants developed. These 16 plants were generally characterized by light green leaves with upright active growth with little branching and white or white / yellow tongue flower color. Ohtsuka and Inaba (2008) further explain that from this hybrid combination, "no new features could be found that could be useful in the production of flower beds and potted plants".

  [0017] The present invention can develop a commercially useful progeny from a cross combination using a female plant selected from the group of Ismeria versicolol and Grebionis sp. Hybridized with a male Argyranthemum heterotetraploid plant. It has made a surprising discovery that it can be done. These crosses surprisingly cause self-pollination of female parents in addition to success with Ismeria versicolol, Grebionis coronaria and Grebionis segetham. All of these species exhibit self-incompatibility (Anderson, NO, Liedl, BE, Ascher, PD, Widmer RE, and Desborough, SL (1988). , Evaluating self-incompatibility in Chrysanthemum, Sexual Plant Reproduction 1: pp. 173-181), the mechanism of stigma incompatibility has been overcome by the application of Argyrantemam pollen.

  [0018] The present invention was unexpected (Liedl, B and Anderson, N) because it is common to experience unilateral incompatibility under conditions of intergeneric hybridization. (1993) Reproductive barriers: Identification, uses and circulation. In: Plant Breeding Reviews, 11: 11-154). In other words, crossing works in one direction but not in the reverse direction. Reasons for this phenomenon include various types of incompatibility and differences in the number of endosperm balances. In addition to useful intergeneric hybrid progeny, female greviolis species and I. A number of progeny progenies were produced by the action of pollination of argyranthemum pollen on the flowers of versicolol. This phenomenon has not been previously reported within the genus described herein.

  [0019] The present invention provides another means for creating new intergeneric hybrid plants, increased head-inflorescence diameter, male sterility, female sterility, improved heat resistance, four seasons bloom, new flower color And development of features such as brighter flower colors. Prior to the present invention, previous reports of successful hybridization at any multiplicity level for plants derived from the group of Ismeria versicolol and Grebionis species used as female parents crossed with Argyranthemum plants as male parents. There was no.

  [0020] The foregoing examples of the related art and limitations related therewith are illustrative and not exclusive. Other limitations of the related art will be apparent to those skilled in the art upon reading this specification.

  [0021] The following embodiments and aspects thereof are described along with exemplary systems, means and methods that are not intended to limit the scope. In various embodiments, one or more of the problems described above have been reduced or eliminated, and other embodiments are directed to other improvements.

  [0022] One aspect of the present invention is the use of Grebionis sp. In a cross with a versicolol plant, the growth of an intergeneric hybrid plant is promoted by using an argyranthemum heterotetraploid plant as a male parent. As used herein, the term heteroploid refers to both tetraploid and heteroploid argyrancemum plants. The number of progeny obtained from the method of the present invention is unexpectedly high, and this progeny is based on having two types, (1) the result of self-pollination and (2) having both parental morphological features. Morphologically differentiated to be hybrid. Argyranthemum pollen is a product of Grevionis sp. It was thought that the self-incompatibility mechanism at the stigma of versicolol plants was facilitated, self-pollination was enabled, penetrated into the stigma, and pollination was performed. In addition to a number of in-house progenies, a small number of hybrid progenies were found. Further modifications could be used to reduce the number of autogenous progenies and increase the number of hybrid progeny. Several methods are well known in the art and include, but are not limited to, male sterile female parent development, application of gamatecide, male removal to leave only female lingual flowers, or For example, pollen can be removed by washing with water or blowing air. The technology of the present invention provides a means for developing new types of intergeneric hybrids as well as the Grevionis sp. Providing a means to develop private progenies from versicolol. Prior to the present invention, Grevionis sp. Hybridized with Argyranthemum as male parent or Previous reports of successful hybridization at any multiplicity level for versicolol female plants, or Grevionis sp. There were no previous reports on the development of autogenous progenies from versicolol. A further aspect of the present invention is to provide a tetraploid argylancemum plant as a female parent and I. as a male parent. It is to provide an intergeneric hybrid plant produced from a cross with a plant derived from the group consisting of Versicolol and Grebionis species.

  [0023] A further aspect of the present invention is that aneuploid tetraploid Argyranthemum plants as female parents and I. It is to provide an intergeneric hybrid plant produced from crossing with a group of plants consisting of Versicolol and Grebionis species.

  [0024] A further aspect of the present invention is that I. as a female parent. It is to provide a plant part of an intergeneric hybrid plant produced by crossing a plant derived from the group consisting of Versicolor and Grebionis species and an Argyranthemum plant as a male parent. A further aspect of the invention is to provide an intergeneric cross to produce a hybrid plant or part thereof that is clonally propagated.

  [0025] A further aspect of the present invention is that I. as a female parent. Crossing a plant derived from the group consisting of Versicolol and Grevionis species with an Argyranthemum plant as male parent, rescue of an embryo resulting from the cross, and obtaining an intergeneric hybrid plant grown therefrom It is to provide a method for producing an intergeneric hybrid plant.

  [0026] A further aspect of the present invention provides: (a) a first plant that is an aneuploid argyrancemum plant; Cultivating a second plant that is a plant derived from the group consisting of Versicolol and Grebionis species, (b) collecting pollen from the first plant, and (c) using the pollen, the second Pollinating the plant's head inflorescence; (d) isolating the embryo and germinating in vitro on a suitable medium; and (e) obtaining an intergeneric hybrid plant resulting from the growth of the embryo. It is to provide a method for producing an intergeneric hybrid plant.

  [0027] A further aspect of the present invention is a cross between (a) a plant derived from the group consisting of Ismeria versicolol, Grebionis coronaria and Grebionis segetham as a female parent, and a male tetraploid argyrancemum. A method for producing an intergeneric hybrid plant, comprising: obtaining a cutting of an argylancemum plant that is an intergeneric hybrid produced from the plant; and (b) cultivating the cutting and obtaining an argylancemum plant that is an intergeneric hybrid. Is to provide.

  [0028] A further aspect of the present invention is an intergeneric hybrid further defined by applying a plant hormone composition to the base of cuttings to induce root formation to produce an intergeneric hybrid plant It is to provide a method for producing plants.

  [0029] A further aspect of the present invention comprises: (a) cultivating an argyrancemum plant; (b) applying an antimitotic agent to the growth point of the plant; and (c) a treated growth point. Germinating shoots (shoots), (d) selecting the putative aneuploid tetraploid shoots thus developed, and (e) a chromosome set of said shoots by cytological karyotype analysis And (f) growing the shoots on the plant, and (g) examining the chromosomal stability, changing the number of chromosomes of the argyrancemum plant, and diploid 2n = 2x = 18 The aneuploid tetraploid 2n = 4x = 32, 33, 34, 35, 36, 37 to 38.

  [0030] A further aspect of the present invention is to provide a method for altering the chromosome number of an Argyranthemum plant, wherein the altered chromosome number is an aneuploid tetraploid for producing an aneuploid tetraploid plant. Defined as body. The plant parts of an aneuploid tetraploid Argyranthemum plant are flowers, cuttings, seeds, pollen, ovules or cells. The plant is then clonally propagated from the plant part.

  [0031] A further aspect of the present invention is that suitable genetic features useful for breeding Argyranthemum plants, preferably targeted traits (eg, male and female fertility, proper habits, speed of flowering). The size and color of the head-like inflorescence, the flowering period, the shape of the head-like inflorescence, etc.).

  [0032] A further aspect of the present invention is to develop an aneuploid tetraploid form of an Argyranthemum plant, preferably by using colchicine or other polyploid inducer (s).

  [0033] A further aspect of the present invention is preferably an increase in the overall head inflorescence diameter, an increase in the diameter of the tubular inflorescence of the head inflorescence, preferably by chromosome number and / or compared to diploid ancestry, Stabilizing and confirming aneuploid tetraploid plants by giving them morphological changes such as increased floral width, larger leaf size, and larger pollen diameter.

  [0034] A further aspect of the invention is that the tetraploid plants of the invention have a chromosome number ranging from 32, 33, 34, 35, 36, 37-38.

  [0035] A further aspect of the invention is that the intergeneric hybrid plant of the invention has a chromosome number of 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, or 27. It is to have.

[0036] A further aspect of the present invention is to provide argyrancemum plants that are teuploid. To plants crossed derived from the group consisting of Berushikororu and Gurebionisu species be collected pollen from F ₁ hybrids, and the pollen is applied to Arugiransemamu plants is different tetraploid, backcrossing the hybrid F ₂ plants It is to develop an allotriploid F ₁ hybrid plant by obtaining.

[0037] A further aspect of the present invention is the I.V. Berushikororu, producing F ₁ hybrid plants by crossing Gurebionisu-Segetamu and Gurebionisu-Koronaria, and selected the hybrid plants pollinated to Arugiransemamu plants is different tetraploid using pollen from this hybrid plants It is to produce a heteroploid hybrid plant.

  [0038] A further aspect of the invention is to infect a triploid argyrancemum hybrid plant with chrysanthemum B virus (CVB) and chrysanthemum dwarf viroid (CSVd).

  [0039] In addition to the exemplary aspects and embodiments described above, further aspects and embodiments will become apparent by studying the following description.

  [0040] In the following description and tables, a number of terms are used. In order to provide a clear and consistent understanding of the specification and claims, including the scope to be given such terms, the following definitions are presented.

[00412] allele. An allele is either one or more alternative forms of a gene with respect to a trait or characteristic. In a diploid cell or organism, two alleles of a given gene occupy corresponding loci on a pair of homologous chromosomes.

[0042] Amiprophos-methyl (APM) . As used herein, amiprophos-methyl (APM) refers to a compound used in plant breeding to induce chromosomal doubling.

[0043] The stamen group . The male part of the flower of a plant, collectively called stamens.

[0044] Anemone . A head-like inflorescence having a tubular flower having an elongated corolla tube. Anemone-type scaly inflorescences appear to be in the middle of normal (single) and double petal flowers. Anemone heads are different from double-headed heads, the latter being a substitute for stamens in tubular flowers.

[0045] Aneuploid tetraploid . As used herein, aneuploid tetraploid refers to tetraploid plants and any plant having a chromosome number approximately four times that of a haploid. For example, an aneuploid tetraploid Argyranthemum plant of the present invention has 32, 33, 34, 35, 36, 37 or 38 chromosomes.

[0046] Aneuploid triploid . As used herein, aneuploid triploid means triploid plants and any plant having a chromosome number approximately three times that of a haploid. For example, the aneuploid triploid plant of the present invention has 23, 24, 25, 26, 27, 28 or 29 chromosomes.

[0047] Anti-miotic agent . As used herein, anti-mitosis is used in plant breeding to induce chromosomal doubling, to disrupt cell growth by stopping mitosis (cell division) Refers to the compound or chemical used. Examples of anti-mitotic agents include, but are not limited to, colchicine, trifluralin, oryzalin and amiprophos-methyl (APM).

[0048] No mating reproduction . Alternative to standard sexual reproduction by asexual reproduction without fertilization. In flowering plants, the term matingless reproduction is generally used to identify asexual reproduction by seeds.

[0049] Argyrantemam . As used herein, Argyranthemum refers to the genus of plants of the family Asteraceae. The genus Argyranthemum includes, but is not limited to: Adactum, A. et al. Broussonetti, A.M. Calichrysum, A.M. Coronopifolium, A.I. Dissectorum, A.I. Escarrei, A.M. Fifofolium, A.I. Foeniclaceum, A. et al. Fullness, A. Gracile, A.M. Haemotoma, A. et al. Haouarytheum, A. et al. Hierence, A.H. Remsii, A.M. Lidii, A.I. Madaraense, A.M. Pinnatifidum, A. Sventenii, A.M. Sundingii, A.M. Thalassophilum, A. Tenerifae, A.M. Vincentii, A.M. Webbie and A.B. About 24 species are included, including winteri (Humphries, CJ, Arevion of the Macrossian genus Argyrantium, Nythium benth, Henrith, Bet., And Bet.). 145-243 (1976)).

[0050] Asexual reproduction / asexual reproduction . Asexual propagation or reproduction means all kinds of plant propagation except sexually produced seeds. Examples of asexual propagation include, but are not limited to, cuttings, grafting, lineage, no-gamete reproduction or regeneration in tissue culture.

[0051] Backcross . Backcrossing is a process in which a breeder repeatedly crosses hybrid progeny back to one of the parents. For example, the first generation hybrid F ₁ has _one of the parent genotypes of the F ₁ hybrid.

[0052] Bridging cross . A method of bypassing an incompatibility barrier between two genotypes or species by using a third genotype or species that partially matches each of the third genotypes or species in an intermediate cross.

[0053] Cranial inflorescence . A head-shaped inflorescence refers to an inflorescence in the form of an outer ring with a structure like a tubular flower (disc) at the center of an unpatterned flower called a disc flower and a petal-like structure called a tongue-like flower. Tubular flowers are generally complete and lingual flowers are generally incomplete. Plural forms of capitula are capitula.

[0054] Cells . As used herein, a cell includes a plant cell, whether isolated, tissue culture, or integrated into a plant or plant part.

[0055] Chimera . A chimeric or chimeric plant is a plant that consists of a group of two or more genetically different cells. Genetic differences usually arise from mutations.

[0056] Chromosome number . Number of chromosomes in plant cells.

[0057] Chromosome stability . As used herein, chromosomal stability refers to a chromosome that is not subject to sudden or extreme changes or fluctuations.

[0058] Chrysanthemum viroid (CSVd). It is a causative factor of “Chrysanthemum dwarf” disease and causes growth inhibition of growth in infected plants, such as reduced flower size and early flowering.

[0059] Chrysanthemum B virus (CVB). A virus that causes leaf spotting or vein permeability in infected plants.

[0060] Colchicine . Colchicine is a light yellow alkaloid, C ₂₂ H ₂₅ NO ₆ , obtained from canine saffron and is used in plant breeding to induce chromosomal doubling.

[0061] Cuttings . A part arising from a plant, such as a stem, leaf or root, removed from the plant to propagate a new plant through rooting or grafting.

[0062] diploid. A diploid (indicated by a somatic chromosome number 2n = 2x) has a pair of each type of chromosome (a homologous pair), and thus a basic (haploid) chromosome number (indicated by the symbol x). Is a somatic cell or plant that has doubled.

[0063] Tubular flowers . One of the small tubular radial florets that make up the central part of the head-like inflorescence of a Compositae or Asteraceae plant.

[0064] Dominant inheritance . Refers to a genetic system in which a particular feature or trait phenotype is determined by a dominant allele.

[0065] Dominant mutation . A dominant mutation phenotype is found in heterozygous genotypes.

[0066] Double petals . A head-shaped inflorescence having a tubular flower in which one or more petals replace stamens.

[0067] Male elimination . Removal of flower buds to prevent self-pollination.

[0068] Embryo . A young plant individual after fertilization or parthenogenesis when the pre-embryo has differentiated into an embryo and stalk.

[0069] Embryo culture . In vitro growth of isolated plant embryos in a suitable medium.

[0070] Embryo rescue . As used herein, embryo rescue is for a plant breeder to germinate embryos that are fragile and immature, or otherwise do not grow into mature, viable seeds on the parent plant. This is the method to use. For example, one form of embryo rescue is ovule culture, which allows ovules to be aseptically removed from seeds and placed on an artificial medium to germinate the embryos and grow into plants.

[0071] F ₂ . The symbol “F ₂ ” indicates a generation resulting from F ₁ self-pollination. The number “F” is a term commonly used in genetics and indicates a hybrid generation. The “F ₂ ” generation refers to offspring resulting from self-pollination or self-mating of members of the first generation, ie F ₁ generation.

[0072] Gametes . A cell or nucleus that can participate in sexual fusion and form a zygote.

[0073] Gene . As used herein, a gene refers to a segment of nucleic acid. Genes can be introduced into certain genomes, whether from different species or from the same species, using transformation or various breeding methods.

[0074] Genetic environment interaction / genotype environment interaction . Refers to the phenotypic effect of gene-environment interaction.

[0075] Genetic transformation . Refers to genetic mutations in cells resulting from uptake, genomic introduction, and expression of foreign genetic material.

[0076] Transformed gene (conversion) . A plant in which a gene has been converted (converted) refers to a plant that is grown by a plant breeding technique called backcrossing, in which case one or more genes introduced into the varieties by backcrossing techniques, genetic manipulation or mutation. In addition, essentially all of the desired morphological and physiological characteristics of the variant are restored.

[0077] Genotype . Refers to the genetic makeup of a cell or organism.

[0078] Grebionis species . As used herein, Grevionis species include, but are not limited to, Grevionis coronaria, also known as Crown Daisy, and Corn Marigold (Mabberley 2008). It refers to the genus of the Asteraceae plants including the known Grevionis segetham. Previous genera names included xantoptalum and chrysanthemum.

[0079] Pistil group . Plant ovule production department.

[0080] Haploid . A haploid is a cell nucleus containing only one representative of each chromosome of a chromosome set and is denoted by the symbol n. The haploid number (n) is the number of chromosomes in the haploid cell nucleus. Gametes are haploid cells.

[0081] Heterozygosity . Refers to a genetic configuration in which the corresponding alleles at a particular locus are different.

[0082] Higher growth temperatures . As used herein, a higher growth temperature refers to the ability of a progeny plant to grow at and withstand a temperature higher than that which the parent used to produce the progeny could withstand. Point to.

[0083] Homozygosity . Refers to a genetic configuration in which the corresponding alleles of a particular locus are identical.

[0084] Inbreeding . Defined as the production of offspring by the fusion of genetically closely related gametes.

[0085] Inbreeding . Inbreeding is a loss of fitness in a given population as a result of inbreeding or in plants that also result from self-pollination. This generally occurs in species that are usually outbred.

[0086] Inflorescence . A group or group of flowers arranged on a stem composed of an arrangement of main branches or branches.

[0087] Intergeneric hybridization . Intergeneric hybridization means sexual hybridization of two individuals of different genera. For example, an Argyranthemum plant crossed with an Ismeria Versicolor plant.

[0088] Intergeneric hybrids . Intergeneric hybrids refer to F ₁ generation plants resulting from intergeneric crosses or from crosses between two different genera.

[0089] Interspecific crossing . Interspecific crossing means sexual hybridization of two individuals of different species of the same genus. For example, a Grevionis coronaria plant crossed with a Grevionis segetam plant.

[0090] Interspecific hybrid . Interspecies hybrids refer to F ₁ generation plants resulting from interspecific crosses or from crosses between two different species.

[0091] I. Versicolor . As used herein, I.I. Versicolol refers to a genus of asteraceae plants, including but not limited to Ismeria versicolol, also known as TricolorDaisy (Mabberley 2008). Previous genus and species names used for Ismeria versicolol include Grevionis carinata, Grevionis carinatum and Chrysanthemum carinatum.

[0092] Karyotype analysis . As used herein, karyotype analysis means confirmation of chromosome number and organization by light microscopic analysis of stained metaphase chromosomes. Cells are collected and induced to differentiate and then arrested at metaphase (the stage of cell division where chromosomes condense and thus become visible). Chromosomes are stained with specific dyes that exhibit a pattern of light and dark bands. Large changes in chromosomes can be detected using karyotype analysis.

[0093] The locus . A locus confers one or more traits such as herbicide resistance, insect resistance, disease resistance, flower color, flower shape, plant height, and the like. A trait can be conferred by, for example, a naturally occurring gene that is introduced into the genome of a variant by backcrossing, natural or induced mutation, or a transgene that is introduced by gene conversion techniques. A locus can include one or more alleles integrated at a single chromosomal location.

[0094] M ₀ . M ₀ generation, a process has been generations in the mutagen. Subsequent generations are denoted as M ₁ , M ₂ , M _3, etc.

[0095] Monogenic inheritance . A particular feature or trait phenotype refers to a genetic system determined by a single gene.

[0096] haploid. The number of chromosomes in a haploid is the number of chromosomes in a single (non-homologous) set (x) and can differ from the number of haploids (n).

[0097] Mutation . Mutations are changes in the DNA sequence of the cell's genome, caused by mutagens such as radiation or chemicals, as well as by errors that occur during DNA replication.

[0098] Orizarin . As used herein, oryzalin refers to a compound used in plant breeding to induce chromosomal doubling.

[0099] Outbred mating . Also known as outcrossing, it is described as the production of offspring by the fusion of distant gametes. Crossbreeding is an antonym for inbreeding.

[0100] Ovule culture . In vitro culture of excised ovules in a suitable medium.

[0101] Phenotype . It refers to any observable feature or trait of a plant such as flower color, plant size.

[0102] Plants . As used herein, the term plant includes reference to all immature or mature plants, including plants from which seeds or pods have been removed. Seeds or embryos that produce plants are also considered plants.

[0103] A plant hormone composition . As used herein, a plant hormone composition refers to a chemical that controls plant growth. For example, indole-3-butyric acid, N ⁶ -benzyladenine and gibberellic acid.

[0104] Plant parts . As used herein, the term plant part includes, but is not limited to, protoplast, leaf, stem, root, root tip, cocoon, pistil, seed, embryo, pollen, ovule, cotyledon. , Hypocotyls, head-like inflorescences, lingual petals / florets, tubular flower petals / florets, shoots, tissues, petiole, cells, meristem cells, and the like.

[0105] Pollination . Pollination is a method that enables fertilization and sexual reproduction by transferring pollen to plants.

[0106] Progeny . As used herein, progenies include Argyranthemum plants and I.I. It includes F ₁ plants produced from a cross between the group of plants consisting of Berushikororu and Gurebionisu species. Progenies include, but are not limited to, subsequent F ₂ , F ₃ , F ₄ , F ₅ , F ₆ , F ₇ , F ₈ , F ₉ and F ₁₀ generations, Further included is a cross between progeny.

[0107] Protoplast fusion / somatic cell fusion . Refers to a plant breeding method in which protoplasts from two different plants (ie plant cells without cell walls) are fused together to form a new hybrid plant with both characteristics.

[0108] Quantitative trait locus (QTL) . Quantitative trait loci (QTL) refer to loci that control to some extent traits that can be represented numerically, usually distributed continuously.

[0109] Tongue flowers . Tongue or sublingual flowers are one of the outer irregular florets in the headed inflorescence of some Compositae or Asteraceae plants. In some Asteraceae or Compositae plants, the tongues of the lingual flowers are called petals.

[0110] Recessive inheritance . Refers to a genetic system in which a particular feature or trait phenotype is determined by a recessive allele.

[0111] Recessive mutation . The recessive mutation phenotype is found only in homozygous genotypes.

[0112] Playback . Regeneration refers to plant development from tissue culture.

[0113] Sexual reproduction / sexual reproduction . Refers to the propagation of plants from seeds.

[0114] Somatic cells . Any cell of the plant other than spores, gametes or their precursors.

[0115] tetraploid. As used herein, tetraploid refers to a cell or plant having a chromosome number four times that of a haploid chromosome. Tetraploid Argyranthemum has 36 chromosomes and is represented by 2n = 4x in somatic cells.

[0116] Trifluralin . As used herein, trifluralin refers to a compound used in plant breeding to induce chromosome doubling.

[0117] triploid. As used herein, triploid refers to a cell or plant having a chromosome number that is three times that of a haploid chromosome. The number of chromosomes in the triploid is 27 and is represented by 2n = 3x in somatic cells.

  [0118] The following embodiments and aspects thereof are described along with exemplary systems, means and methods that are not intended to limit the scope. In various embodiments, one or more of the problems described above have been reduced or eliminated, and other embodiments are directed to other improvements.

  [0119] In one aspect of the invention, new plants have been produced that have unique characteristics that are desirable for use as ornamental plants. These new plants of the present invention are previously unknown intergeneric hybrids and were created at Yellow Rock, New South Wales, Australia. This plant is a strain of I. as a female parent. It was produced from an unexpected finding that an intergeneric hybrid plant can be efficiently produced from a cross between a plant derived from the group of Versicolor and Grebionis species and an Argyranthemum plant as a male parent.

  [0120] The present invention provides another means for developing new intergeneric hybrid plants using Argyranthemum. Previously, I. crossed with any Argyranthemum. There were no reports of successful hybridization at any fold level for Versicolol and Grebionis species.

  [0121] A further aspect of the invention consists of a female parent, Ismeria versicolol, also known as Tricolor Daisy, Grevionis coronaria, also known as Shungiku, and Grevionis segetham, also known as Alageshungiku. It is to provide an intergeneric hybrid plant produced from a cross between a plant derived from a group and an Argyranthemum plant as a male parent.

  [0122] A further aspect of the present invention is derived from the group consisting of Ismeria versicolol, also known as tricolor daisy, Grevionis coronaria, also known as Sengiku, and Grevionis segetam, also known as Alageshungiku, as male parents. It is to provide an intergeneric hybrid plant produced from a cross between a plant and an Argyranthemum plant as a female parent.

  [0123] A further aspect of the present invention is a method for the treatment of I.I. It is to provide a plant part of an intergeneric hybrid hybrid haploid plant produced by crossing a plant derived from the group consisting of Versicolol and Grebionis species and an Argyranthemum plant as a male parent.

  [0124] One aspect of the invention is to provide an intergeneric cross to produce a hybrid plant or portion thereof that is clonally propagated.

  [0125] A further aspect of the present invention is a method for the treatment of I.V. Crossing a plant derived from the group consisting of Versicolol and Grebionis species with an Argyranthemum plant as a male parent, rescuing an embryo resulting from the cross, and obtaining an intergeneric hybrid plant grown therefrom It is to provide a method for producing intergeneric hybrid hybrid triploid plants.

  [0126] A further aspect of the present invention is a method for the treatment of I.M. Crossing a plant derived from the group consisting of Versicolol and Grevionis species with an Argyranthemum plant as a female parent, rescuing an embryo resulting from the cross, and obtaining an intergeneric hybrid plant grown therefrom It is to provide a method for producing intergeneric hybrid hybrid triploid plants.

  [0127] Further embodiments of the invention include (a) I.I. Cultivating a first plant that is a plant derived from the group consisting of Versicolol and Grebionis species and a second plant that is an Argyranthemum plant; (b) collecting pollen from the second plant; c) pollinating the first plant's head inflorescence with this pollen; (d) isolating the embryo and germinating in vitro on a suitable medium; and (e) resulting from the growth of this embryo. Obtaining an intergeneric hybrid plant, comprising the step of producing an intergeneric hybrid plant.

  [0128] A further aspect of the present invention is a genus produced from a cross between (a) a plant derived from the group consisting of Ismeria versicolol, Grebionis coronaria and Grebionis segetham as a female parent and Argyranthemum, a male parent. It is to provide a method for producing an intergeneric hybrid plant, comprising the steps of obtaining an interhybrid plant cutting and (b) cultivating the cutting to obtain an intergeneric hybrid plant.

  [0129] A further aspect of the invention is the production of an intergeneric hybrid plant, further defined by applying a plant hormone composition to the base of the cuttings to induce root formation to produce an intergeneric hybrid plant Is to provide a method.

  [0130] A further aspect of the present invention comprises: (a) cultivating an argylancemum plant; (b) applying an antimitotic agent to the growth point of said plant; and (c) from the treated growth point. Germinating shoots; (d) selecting putative aneuploid tetraploid shoots thus developed; (e) evaluating a chromosomal set of said shoots by cytological karyotype analysis; And (f) growing the shoots on the plant, and (g) examining the chromosome stability, and changing the number of chromosomes of the argyrancemum plant, from diploid 2n = 2x = 18, It is to provide a method for increasing the number of somatic chromosomes to 2n = 4x = 32, 33, 34, 35, 36, 37-38 in sex tetraploid.

  [0131] A further aspect of the present invention is to provide a method for altering the chromosome number of an Argyranthemum plant, wherein the altered chromosome number is an aneuploid tetraploid for producing an aneuploid tetraploid plant. Defined as body. The plant parts of an aneuploid tetraploid Argyranthemum plant are flowers, cuttings, seeds, pollen, ovules or cells. The plant is then clonally propagated from the plant part.

  [0132] A further aspect of the present invention is to provide suitable genetic features useful for breeding Argyranthemum plants, preferably targeted traits (eg, male and female fertility, proper habits, speed of flowering). The size and color of the head-like inflorescence, the flowering period, the shape of the flower, etc.).

  [0133] A further aspect of the present invention is to develop aneuploid tetraploid forms of Argyranthemum plants, preferably by using colchicine or other polyploid inducer (s).

  [0134] A further aspect of the invention is preferably an increase in the overall head inflorescence diameter, an increase in the diameter of the tubular inflorescence of the head inflorescence, preferably by chromosome number and / or compared to diploid ancestry, Stabilizing and confirming aneuploid tetraploid plants by giving them morphological changes such as increased floral width, large leaf size, and large pollen diameter.

  [0135] A further aspect of the invention is that the aneuploid tetraploid plants of the invention have a chromosome number in the range of 32, 33, 34, 35, 36, 37-38.

  [0136] A further aspect of the invention is that the intergeneric hybrid plant of the invention has a chromosome number of 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, or 27. It is to have.

[0137] Further embodiments of the invention include Argyransemam, which is a heteroploid, To crossing a plant derived from the group consisting of Berushikororu and Gurebionisu species be collected pollen from F ₁ hybrids, and by applying the pollen to Arugiransemamu plants is different tetraploid, backcrossing the hybrid F ₂ plants To develop an adiploid F ₁ hybrid.

[0138] A further aspect of the present invention is the I.V. Berushikororu, producing F ₁ hybrid plants by crossing Gurebionisu-Segetamu and Gurebionisu-Koronaria, select hybrid plants pollinated to Arugiransemamu plants is different tetraploid using pollen from this hybrid plants It is to produce a heteroploid hybrid plant.

  [0139] Another further aspect of the present invention is the introduction of a virus (Chrysanthemum B virus, CVB) and viroid (Chrysanthemum dwarf viroid, CSVd) into an allitriploid Argyranthemum hybrid plant. Possibly altering its potential as a commercially viable product by changing its phenotype.

[0140] In addition to the exemplary aspects and embodiments described above, further aspects and embodiments will become apparent by studying the following description.
I. Development of aneuploid tetraploid Argyranthemum species

  [0141] The present invention provides a method for producing aneuploid tetraploid Argyranthemum plants as defined herein having a chromosome number ranging from 32, 33, 34, 35, 36, 37-38. To do.

  [0142] Argyranthemum plants are derived from any one of the following species or hybrids of the following species: A. Adactum, A.E. Browsonity, A.B. Calicrisum, A.M. Coronopifolium, A. Dissector, A. Escalai, A. Filifolium, A. Phoenicaceum, A. Fullness, A. Grachle, A. Haemoto, A. Hauriateum, A. Hierence, A. Remsii, A. Redy, A. Madalens, A.M. Pinnafifidam, A. Sventeni, A. Sundingui, A. Thalassofilum, A. Tenerifae, A.M. Vincenti, A.M. Webby and A.C. Wintery. An aneuploid tetraploid form of an argyrancemum plant is developed, preferably by using an anti-mitotic agent. Examples of anti-mitotic agents include, but are not limited to, colchicine, trifluralin, oryzalin, amiprophos-methyl, and other polyploid inducer (s). The tetraploid may occur naturally or can be induced using a spindle inhibitor such as colchicine. The technique of polyploidization induced with colchicine has been used since the 1930s. Colchicine inhibits the incorporation of the tubulin subunit into the spindle, resulting in no chromosomal transfer and accumulation of cells at the metaphase stage of mitosis. If the chromatids separate but are not divided into separate cells by the spindle, the number of chromosomes doubles to create a homoploid. When creating polyploids for breeding purposes, the apical meristem layer that produces gametophyte tissue must be doubled. In order to optimize the likelihood of doubling success, a large number of actively growing small meristems are processed. Typically, colchicine is used at a concentration of 0.1% to 2.0% depending on the tissue and species. Methods for treating seeds and plant parts with colchicine or other spindle inhibitor are described in Poehlman, J. et al. M.M. Breeding Field Crops, University of Missouri, Holt, Rinehardt and Winston Inc. (1966); Watts, L .; , Flowerand Vegetable Plant Breeding, Grower Books (1980); Callaway D. et al. J. et al. And Callaway M. et al. B. , Breeding Original Plants, Timber Press Inc. (2000) is well known in the art.

[0143] Ploidy changes affect crossability, fertility, cell size and heterozygosity. These factors provide limitations as well as potential benefits in plant breeding. Ploidy manipulation was used for germplasm introgression between different ploidy taxa. For example, tetraploids were induced using colchicine to overcome the F ₁ sterility of interspecific lily hybrids. A tetraploid level interspecific cross was performed between four lily species hybrid hybrids. Van Tuyl, J.A. And van Holsteijn, H .; See Lily breathing research in the Netherlands Acta Horticulture, 414: 35-45 (1996). Naturally produced a diploid Buddleja globosa tetraploid plant using colchicine treatment and introduced Budreja globosa in order to introduce yellow flowers into Buddleja davidii. Crossed with the tetraploid Buddeleya Davidi. Rose, J. et al. Kubba, J .; And Tobutt, K .; See Induction of tetraproids for bleeding hard ornaments, Acta Horticulture, 560: 109-112 (2001). The all-yellow cyclamen persicum varieties are diploid and do not have “eyes” on the petals. Using colchicine treatment, tetraploid yellow flower cyclamen were induced. After crossing with the tetraploid “meat” varieties, separation was performed so that the “mesh” selection of yellow flowers was not maintained by the seeds. Takamura, T .; Sugimura, T .; Tanaka, M .; And Kage, T .; See Breeding of the yellow-flowed tetracyclide with “eye”, Acta Horticulture, 454: 119-126 (1998).

[0144] The present invention comprises: (a) cultivating an argylancemum plant; (b) applying an anti-mitotic agent to the growth point of the plant; and (c) shoots from the treated growth point of the plant. Budding; (d) selecting a putative aneuploid tetraploid shoot thus developed from the plant; and (e) a chromosome of the aneuploid tetraploid shoot by cytological karyotype analysis. Changing the number of chromosomes in the Argyranthemum plant, comprising: evaluating the set; (f) growing aneuploid tetraploid shoots on the plant; and (g) examining chromosome stability. Methods are provided for developing aneuploid tetraploid plants having chromosome numbers ranging from 32, 33, 34, 35, 36, 37-38. One skilled in the art can induce tetraploid induction to produce plants with more or less chromosome numbers than the expected tetraploid number, and such aneuploid plants are referred to herein as aneuploid tetraploids. It will be understood that this includes plants with chromosome numbers ranging from 32, 33, 34, 35, 36, 37-38.
II. Intergeneric hybrid plant production

  [0145] The flowering industry is committed to developing new and different varieties of flowering plants. One effective way to create such new varieties is by flower color manipulation. Flower color is mainly attributed to two types of pigments: flavonoids and carotenoids. Flavonoids contribute to colors ranging from yellow to red to even blue. Carotenoids impart a reddish-orange or yellow shade and are generally the only pigments of yellow or orange flowers. The flavonoid molecule that contributes greatly to flower color is anthocyanin, which is a glycosylated derivative of cyanidin, delphinidin, petunidin, peonidin, malvidin and pelargonidin, localized in the vacuole. A variety of anthocyanins gives a noticeable color difference. Flower color is also affected by co-pigmentation with colorless flavonoids, metal complexation, glycosylation, acylation, methylation and vacuolar pH. Forkman, G .; See Flavonoids flow pigments: the formation of the natural spectrum and its extension by genetic engineering, Plant Breeding 106: 1-26 (1991).

  [0146] The present invention unexpectedly gave rise to intergeneric hybrid plants with new flower colors and other useful attributes such as improved heat resistance, larger head-like inflorescences. The method of the present invention uses a plant derived from Ismeria versicolol, Grebionis coronaria or Grebionis segetham as a female parent in an intergeneric cross with an Argyranthemum plant as a male parent. Collecting pollen from the Argyranthemum plant; (b) using this pollen Pollinating the inflorescences of female plants of the versicolol or grebionis species, (c) isolating embryos resulting from pollination by embryo rescue, and (d) culturing the embryos in vitro on a nutrient medium of agar And (e) obtaining an intergeneric hybrid plantlet resulting from the growth of the embryo, and (f) transferring the plantlet to a growth medium in a greenhouse where the mature intergeneral hybrid plant is developed. It was out. By using the heteroploid male Argyranthemum, the parent hybrid progeny was produced, but the majority of the progeny were the result of self-pollination by a smaller number of true hybrids.

  [0147] A further aspect of the invention is the step of: (a) cross-breeding a group of plants consisting of Ismeria versicolol, Grevionis coronaria or Grevionis segetham in any combination to produce an embryo from said cross; Providing a method comprising: using embryo rescue for said embryo; and (c) obtaining an interspecies hybrid plant that can then be used as a female parent in a cross with a male parent, Argyrantemam.

  [0148] A further aspect of the invention is that the intergeneric hybrid plant of the invention has a chromosome number of 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, or 27. A diploid to haploid triploid as defined herein.

  [0149] A further aspect of the invention was produced from the cross of (a) a plant derived from the group consisting of Ismeria versicolol, Grebionis coronaria and Grebionis segetham as female parent and Argyranthemum plant as male parent. An intergeneric hybrid plant is propagated, including the step of obtaining an intergeneric hybrid plant cutting and (b) cultivating the cutting to obtain an intergeneric hybrid plant.

[0150] A further aspect of the present invention is the production of an intergeneric hybrid plant, further defined by applying a plant hormone composition to the base of cuttings to induce root formation to produce an intergeneric hybrid plant Is to provide a method.
III. Grevionis sp. XI. As male parent in cross-linking Production of intergeneric hybrid plants using versicolol hybrids

[0151] It is common in plant breeding that some plants cross and other plants do not cross. For example, plant A crosses very well with plant B, but rarely crosses with plant C. However, plants B and C are easily crossed. In order to introduce a gene derived from plant C into a hybrid with plant A, cross-linking hybridization can be performed. Under such circumstances, plants B and C are crossed to develop an F ₁ hybrid plant, pollen is collected from the F ₁ hybrid plant, applied to the stigma of plant A, and a new hybrid plant is developed. As a result, the hybrid is Ax (BxC). Using this method, the following combinations were developed: heteroploid intergeneric hybrids of argyransemam x (all possible combinations of Grevionis sp. XI. Versicolol).

[0152] The present invention surprisingly found that one species is Argyranthemum and the other two species are G. cerevisiae. Segetam, G. Coronaria and I.I. It allowed the development of intertriploid hybrid plants with characteristics derived from at least three separate species selected from the group of versicolol.
IV. Backcross of a heteroploid hybrid

[0153] Breeding ornamental plants includes the development of new types of plants with new characteristics. Under some circumstances, new plants that have developed cannot be used for further breeding operations due to incompatibilities such as incompatibility, various incompatibility, non-uniform chromosome numbers and other unknown factors. An example of this problem was found for intertriploid hybrids developed by crossing the heteroploid algillan semem with Ismeria versicolol. Almost all plants that developed were male sterile (no pollen). Surprisingly, several plants with some pollen were produced. Hybridization has been described previously, using pollen collected from an allotriploid F ₁ hybrid plant and applying that pollen to the arboretum mudhead inflorescence to develop backcross progeny. Performed according to standard procedures.
V. Infection of hybrid plants with chrysanthemum B virus (CVB) and chrysanthemum dwarf viroid (CSVd)

  [0154] A further aspect of the invention is to infect a triploid argyrancemum hybrid plant with chrysanthemum B virus (CVB) and chrysanthemum dwarf viroid (CSVd). This involves grafting an infected plant to an uninfected plant, healing the graft, then growing a newly infected plant, and testing it by PCR and / or ELISA based methods to confirm the infection status Can be achieved.

  [0155] In addition to the exemplary aspects and embodiments described above, further aspects and embodiments will become apparent by studying the following examples.

[0156] The following examples are presented to further illustrate the present invention. These examples should not be construed to limit the scope of the invention in any manner beyond the limitations set forth in the appended claims. Many changes and modifications may be made within the spirit and scope of the invention.
Example 1-Development of an aneuploid tetraploid Argyranthemum species

  [0157] The present invention provides a new method of altering the number of chromosomes in an Argyranthemum plant to double the number of somatic chromosomes from diploid to aneuploid tetraploid. The method for changing the number of chromosomes of the present invention was started by first cultivating an argylan semam plant, and then an anti-mitotic agent such as colchicine, trifluralin, oryzalin or amiprophos-methyl (APM) was applied to the growth point of the argylan semam plant. Next, aneuploid tetraploid shoots were budded from the treated growth points of the Argyranthemum plant, and putative aneuploid tetraploid shoots developed from the growing point of the Argyranthemum plant were selected. The aneuploid tetraploid shoot chromosome set was then evaluated by cytological karyotype analysis, and then the analyzed aneuploid tetraploid shoots were grown in plants. New argylansemam plants were examined for chromosomal stability and new aneuploid tetraploid argylansemema plants were selected and maintained. Pollen is collected from the inflorescences of Argyranthemum aneuploid tetraploid plants and then receptive Grevionis sp. It was applied to the inflorescences of versicolol plants. The embryos were then rescued from the cross, and the rescued embryos were then isolated by embryo rescue in tissue culture and intergeneric hybrid plants were produced from the rescued embryonic tissues.

  [0158] One aspect of the present invention includes, but is not limited to, male and female fertility, appropriate habits, speed of flowering, size and color of head-like inflorescence, flowering time, and head-like inflorescence Obtaining a plant of the genus Argyranthemum having appropriate genetic characteristics useful for breeding the target trait, including After the Argyranthemum plant was obtained, an aneuploid tetraploid Argyranthemum plant was developed using the following method. 100 cuttings were collected from a vegetatively growing stockplant, the cut base was dipped in 2000 ppm indole-butyric acid powder and then planted in a breeding wedge-shaped trademark Oasis® . The cuttings were then placed under intermittent spray in a breeding house maintained at about 20 ° C. After 3 weeks, the cuttings were acclimated to greenhouse conditions and then one plant was planted in each of 100 10 cm diameter pots filled with a standard nursery potting mix. Plants planted in pots were placed in a greenhouse with a minimum temperature of 15 ° C. After about 2 weeks, the plants were ready for colchicine application. Colchicine was prepared as a 0.1% to 2.0% weight / volume paste using a water-soluble gel. By treating a plant with a range of colchicine concentrations, it was possible to determine which concentration yielded optimal results for each line treated.

  [0159] The paste was applied thoroughly to the shoots and washed off after 24 hours using a fine water spray. By counting the number of plants and the applied buds during application, the production efficiency could be measured at the end.

  [0160] Plants were observed and maintained in accordance with best practices and growth procedures for several weeks thereafter (eg, Hamrick, D. (Ed), BallRed Book, Crop Production Volume 2, Ball Publishing (2003)). New shoots were evaluated for signs of heteroploidy. These signs include the following shoots with larger leaves than normal leaves and wide petiole diameter, large stigma florets, large inflorescences with central tubular florets, long floral lengths and widths As well as florets of large pollen diameter. The shoots that looked diploid were cut from the plant. Selected putative aneuploid tetraploid shoots were labeled and trimmed to promote further growth.

[0161] Once new putative aneuploid tetraploid shoots were produced, cuttings were harvested and propagated. These second generation plants were continuously evaluated for quasi-ploidy stability and uniformity based on phenotype. The putative aneuploid tetraploid was then confirmed or discarded by performing karyotype analysis. After confirmation, the plants were propagated at least twice more so that they became stable aneuploid tetraploids. Plants were periodically examined visually for tetraploid morphological characteristics. Plant morphological changes include, but are not limited to, an increase in the diameter of the overall head inflorescence, an increase in the diameter of the tubular inflorescence, Increasing width, large leaf size, large stigma size, and large pollen diameter were included. Chromosomes were counted for selected strains of Argyranthemum, Ismeria and Grebionis, and intergeneric hybrids.
Example 2-Chromosome numbers of Argyranthemum, Ismeria and Grebionis strains

実施例３−発育したアルギランセマム異四倍体株の起源及び説明 [0162] Table 1 below shows the number of chromosomes in the Argyranthemum, Ismeria and Grebionis strains. The first column shows the plant identification number, the second column shows the genus and strain, the third column shows the range of the number of chromosomes, and the fourth column shows the confirmed ploidy level.

Example 3 Origin and Description of Growing Argyranthemum Heteraploid Strains

実施例４−表４〜１０に示した育種交雑で使用したグレビオニス種及びＩ．ベルシコロール変種及びハイブリッド [0163] Table 2 below provides the origin and description of the argyrantemam heterotetraploid plants that were produced and then used in the breeding crosses shown in Tables 4-10. A diploid parent strain (shown in column 1 and phenotype in column 2) was treated with various amounts of colchicine, a chemical mutagen that can cause chromosome doubling. Heteroploid plants selected for further breeding and analysis (indicated by their identification numbers) are shown in the third column. The phenotype of the heterotetraploid strain is shown in the fourth column.

Example 4-Grevionis species used in the breeding crosses shown in Tables 4-10 and Versicolor variety and hybrid

実施例５−同じ遺伝子型から得られた二倍体アルギランセマム雌性親及び異四倍体アルギランセマム雌性親ｘグレビオニス種及びＩ．ベルシコロールを用いる属間ハイブリッド植物の生産効率の比較 [0164] Table 3 below shows the Grevionis species used in the breeding crosses shown in Tables 4-10 and The plant names and descriptions of versicolol varieties and hybrids are presented. The plant name is shown in the first column, the plant identification number is shown in the second column, and the shape and color of the head-like inflorescence are shown in the third column.

Example 5-Diploid Argyransemema female parent and Heteroploid Argyransemema female parent x Grevionis species obtained from the same genotype and Comparison of production efficiency of intergeneric hybrid plants using versicolol

[0165] A comparative cross was performed to determine the production efficiency between the diploid and adiploid argyrancemum female parents used. Four diploid Argyranthemum female parents were used. These same accessions of heterotetraploid form were similarly developed and used. A diploid and heterotetraploid Argyranthemum plant as the female parent and Grebionis sp. Crossing was carried out with versicolol plants. The results were compared. Table 4A shows the number of headed inflorescences pollinated for each hybrid combination, Table 4B shows the number of rescued embryos, and Table 4C shows the number of plants developed.

[0166] Tables 4A, B, and C show I.V. It shows the improved production efficiency that can be obtained when using a heteroploid argylancemum plant as a female parent in a hybrid combination using versicolol and Grebionis species.
Example 6 Interspecific Cross-Grebionis sp. Methods for hybridization of versicolol

  [0167] In a further aspect of the invention, the Grevionis coronaria, Grevionis segetham and Ismeria versicolol species are combined in any possible pair combination according to the same procedure as previously described for other crossings performed. Crossbreeding. Each pair combination produced several viable progeny and grew into flowering plants.

[0168] Male parents to evaluate flowering F ₁ plants and cross with those with desirable characteristics (high pollen fertility, specific flower color or flower color pattern, crowding habits, etc.) with a tetraploid Argyranthemum female parent Used as. The goal was to further increase the diversity of allotriploid progeny and to incorporate new genes and gene combinations into the progeny. For example, Grevionis segetham does not produce many progeny when crossed with an allotetraploid Argyranthemum female parent. The Gurebionisu-Segetamu crossed with Isumeria-Berushikororu, allowed to develop a F ₁ hybrid plants, by using the F ₁ hybrid plants as male parents in crosses with different tetraploid Arugiransemamu female parent derived from Gurebionisu-Segetamu gene Was incorporated into the progeny of the triploid. This type of hybridization is called cross-linking.

[0169] The F ₁ hybrid between Ismeria Versicolol x Grebionis Coronaria and Grebionis Coronaria x Ismeria Versicolol is Chaudhuri B. K. Chaudhuri S .; K. Basak, S.M. L. And Dana, S, Cytogenetics of a crossbetween specials of annual Chrysanthhemum, Cytologia 41: 111-121 (1976). These authors also reported also F ₂ hybrid growth due to interbreeding the F ₁ hybrid. Bose, M, Miller, R, and Deloles, S, Chrysanthhemum system, genetics and breeding, Plant Breeding Reviews no. 14, 321-361, John Wiley and Sons (1997). Versicolol and G. It is described that coronaria can hybridize. Dowrick, G.M. And El-Bayoumi, A, Nucleic acid content and chromatography in Chrysanthemum, Genetic Research Cambridge 13: 241-250 (1969). Segetam and G. The DNA content of the hybrid with coronaria was investigated. Tables 5A, 5B, and 5C below show the results of interspecific crossings that took place from 2000 to 2012 at Yellow Rock, New South Wales, Australia. The female parent strain is shown in the first column, and the male parent strain is listed by its plant identification number in the top row of each table. Embryos were rescued 14-21 days after pollination. Table 5A shows the number of headed inflorescences pollinated for each cross. Table 5B shows the number of rescued embryos. Table 5C shows the number of embryos germinated and planted in greenhouse pots. DU indicates that data is not available.

[0170] As shown in Table 5A above, for example, in a cross between a female Grebionis segetham plant 08-89 and a male Ismeria versicolol plant 08-80, two headed inflorescences were pollinated (line 16). Eyes, 6th column).

[0171] As shown in Table 5B above, 14 embryos were rescued from two pollinations of the female Grebionis segetham plant 08-89 and the male Ismeria versicolol plant 08-80 (line 16, 6). Column).

[0172] As shown in Table 5C above, out of 14 embryos rescued from two pollinations of female Grebionis segetham plant 08-89 and male Ismeria versicolol plant 08-80 (Table 5B), Ten embryos germinated and grew into plants in the greenhouse (16th row, 6th row).
Example 7-Heteroploid Argyranthemum as a female parent and various male Grebionis species xI. Cross-linking with versicolol hybrid

[0173] I. Following the development of an interspecific hybrid progeny derived from the cross between Versicolol and Grebionis species, the heterotetraploid Argyranthemum as a female parent and various Grebionis species xI. Crosses were carried out with versicolol hybrid male parents. Heteroploid female parents were classified based on the diploid parent from which they were obtained, and 5 headed inflorescences were pollinated within each group for each male parent listed. Two male parents were selected for each male parental cross combination. In Tables 6A, B and C below, the left column represents the heteroploid argyrantemam female parent and the remaining 8 columns represent various combinations of various I.D. Represents versicolol x Grevionis hybrid male parent. Table 6A shows the number of pollenate inflorescences, Table 6B shows the number of embryos rescued from cross-pollination, and Table 6C shows the number of hybrid cross-bred hybrid plants that have developed.

[0174] As shown in Table 6B, the total number of embryos rescued from the cross-pollination performed was 221.

  [0175] As shown in Table 6C, the total number of hybrid cross-bred plants that developed was 132.

[0176] Plants obtained from the work carried out in Tables 6A, B and C include Argyranthemum, Grebionis sp. It allowed the incorporation of several combined features derived from versicolol. One skilled in the art can appreciate that further iterations of crossing and fine tuning of the parental combination can increase the number of progeny and plants in any crossing combination.
Example 8-Hybridization method between Ismeria versicolol plant and Argyranthemum heterotetraploid plant

[0177] Another aspect of the present invention involved crossing a plant derived from the group consisting of Ismeria versicolol and an arbiranecem genus tetraploid plant. I. Versicolor female plants were selected for desired traits such as flower color, flower stripes, ease of growth and reproduction, high branching, speed of flowering and male and female fertility. The pollen was then removed from the selected tetraploid Argyranthemum plant used as the male parent. The pollen If the Versicolor roll head is receptive, a small brush is used. It was applied on the inflorescences of versicolol female plants. I. No male velocicolous inflorescence was performed. Next, pollen derived from an adiploid argylancemum plant was pollinated. The head-like inflorescence derived from the Versicolor plant was harvested after 2-3 weeks, and the florets were taken out. Aseptic technique was then applied to each floret. Preferably, the florets were placed in a container with 1% sodium hypochlorite added to coat the material, followed by a drop of Tween 20 surfactant. The container was closed with a lid and shaken once every minute for 5 minutes. The vessel was then emptied and the florets were rinsed 3 times with autoclaved distilled water. The florets were then removed and examined individually. Any developing seed coat and ovary wall tissue was then removed to expose the ovule. The embryos were then removed from the ovule and placed on an appropriate embryo rescue medium in a Petri dish. The medium is preferably half of the recommended ratio (ie 2.21 g / L) of Murashige and Skoog (1962) salt (Murashige, T. and Skoog, F. Physiologia Plantarum, 15: 473-497 (1962)), containing 1% activated carbon, 20 g / L sucrose and 0.7% agar. The medium was adjusted to pH 5.8 and then autoclaved at 121 ° C. for 17 minutes at 1 kg / m ² . Petri dishes containing the scrutinized embryos were sealed with paraffin and placed in a growth chamber maintained under fluorescent light for 16 hours a day at a continuous temperature of 25 ° C. After 2 to 4 weeks, the germinated embryos are transferred to a greenhouse environment where they follow the conventional nursery practice suitable for the growth of Argyranthemum plants (eg Hamrick, D. (Ed), (2003)) to develop the embryos. Promoted to mature plants. For example, germinated embryos were transferred to a seed growing mixture containing fertilizer, kept moist and placed in the shade. The germinated embryos were then transferred to larger pots with conventional potting medium and high light intensity, grown and flowered. Then, were selected F ₁ hybrid plants on desired estimation was collected cuttings.
Example 9-Hybridization method between Grevionis coronaria plant and heterotetraploid Argyranthemum plant

[0178] Another aspect of the present invention comprises crossing a plant derived from the group consisting of Grebionis coronaria with a tetraploid plant of the genus Argyranthemum. G. Coronaria female plants were selected for desired traits such as flower color, flower stripes, ease of growth and reproduction, high branching, speed of flowering and male and female fertility. Pollen was removed from selected tetraploid Argyranthemum plants to be used as male parents. The pollen is treated with G. If the coronal inflorescence is receptive, use a small brush to It was applied on the inflorescences of coronaria female plants. G. The males of coronal inflorescences were performed. Next, pollen derived from a heteroploid argyrancemum plant was pollinated. Head-like inflorescences derived from coronaria plants were harvested after 2-3 weeks and florets were removed. Aseptic technique was then applied to each floret. Preferably, the florets were placed in a container with 1% sodium hypochlorite added to coat the material, followed by a drop of Tween 20 surfactant. The container was closed with a lid and shaken once every minute for 5 minutes. The vessel was then emptied and the florets were rinsed 3 times with autoclaved distilled water. The florets were then removed and examined individually. Any developing seed coat and ovary wall tissue was then removed to expose the ovule. The embryos were then removed from the ovule and placed on an appropriate embryo rescue medium in a Petri dish. The medium preferably contained half of the recommended ratio (ie 2.21 g / L) Murashige and Skoog (1962) salt, 1% activated carbon, 20 g / L sucrose and 0.7% agar. The medium was adjusted to pH 5.8 and then autoclaved at 121 ° C. for 17 minutes at 1 kg / m ² . Petri dishes containing the scrutinized embryos were sealed with paraffin and placed in a growth chamber maintained under fluorescent light for 16 hours a day at a continuous temperature of 25 ° C. Two to four weeks later, the germinated embryos are transferred to a greenhouse environment where they follow the conventional nursery practice suitable for the growth of Argyranthemum plants (eg Hamrick, D. (Ed), (2003)) to allow embryo development. Promoted to mature plants. For example, germinated embryos were transferred to a seed growing mixture containing fertilizer, kept moist and placed in the shade. The germinated embryos were then transferred to larger pots with conventional potting medium and high light intensity, grown and flowered. Then, were selected F ₁ hybrid plants on desired estimation was collected cuttings.
Example 10—Hybridization method between Grevionis segetham plant and heterotetraploid Argyranthemum plant

[0179] Another aspect of the present invention included crossing a plant derived from the group consisting of Grevionis segetham with an arbiranecemum heteroploid plant. G. Segetam female plants were selected for desired traits such as flower color, ease of growth and reproduction, high branching, fast flowering and male and female fertility. Pollen was removed from selected tetraploid Argyranthemum plants to be used as male parents. The pollen is treated with G. If the Segetam's head is receptive, use a small brush to It was applied on the inflorescences of Segetam female plants. G. Segatum's head inflorescence was not eliminated. Next, pollen derived from a heteroploid argyrancemum plant was pollinated. Head-like inflorescences derived from Segetam plants were harvested after 2-3 weeks and florets were removed. Aseptic technique was then applied to each floret. Preferably, the florets were placed in a container with 1% sodium hypochlorite added to coat the material, followed by a drop of Tween 20 surfactant. The container was closed with a lid and shaken once every minute for 5 minutes. The vessel was then emptied and the florets were rinsed 3 times with autoclaved distilled water. The florets were then removed and examined individually. Any developing seed coat and ovary wall tissue was then removed to expose the ovule. The embryos were then removed from the ovule and placed on an appropriate embryo rescue medium in a Petri dish. The medium preferably contained half of the recommended ratio (ie 2.21 g / L) Murashige and Skoog (1962) salt, 1% activated carbon, 20 g / L sucrose and 0.7% agar. The medium was adjusted to pH 5.8 and then autoclaved at 121 ° C. for 17 minutes at 1 kg / m ² . Petri dishes containing the scrutinized embryos were sealed with paraffin and placed in a growth chamber maintained under fluorescent light for 16 hours a day at a continuous temperature of 25 ° C. Two to four weeks later, the germinated embryos are transferred to a greenhouse environment where they follow the conventional nursery practice suitable for the growth of Argyranthemum plants (eg Hamrick, D. (Ed), (2003)) to allow embryo development. Promoted to mature plants. For example, germinated embryos were transferred to a seed growing mixture containing fertilizer, kept moist and placed in the shade. The germinated embryos were then transferred to larger pots with conventional potting medium and high light intensity, grown and flowered. Then, were selected F ₁ hybrid plants on desired estimation was collected cuttings.
Example 11-I. as a female parent Intergeneric crosses using versicolol and Grevionis species and Argyranthemum teuploid as male parents

[0180] I. as a female parent according to previously described methods. Crosses were performed using versicolol and Grevionis species and Argyranthemum terploid as male parents. Table 7A shows I.V. as a female parent. The number of head-like inflorescences pollinated using versicolol and grebionis species and argylancemum tetraploid as male parents is shown.

  [0181] As shown in Table 7A, I. A total of 61 head-like inflorescences were pollinated using versicolol and Grebionis species and Argyranthemum terploid as male parent.

[0182] Table 7B shows I.V. The number of embryos rescued from pollination pollinated with versicolol and grebionis species and argylancemum tetraploid as male parents is shown.

  [0183] As shown in FIG. A total of 341 embryos were rescued from pollination pollinated with versicolol and Grevionis species and Argyranthemum terploid as male parent.

[0184] Table 7C shows I.D. The number of hybrids and self-pollinated plants developed from pollination using versicolol and Grevionis species and Argyranthemum terploid as male parents is shown.

  [0185] As shown in Table 7C, I.I. A total of 37 hybrids and 166 self-breeding individuals were developed from pollination using versicolol and Grevionis species and Argyranthemum teuploid as male parents.

  [0186] The work performed in Tables 7A, B, and C was performed as I.V. as a female parent crossed with a male diploid argyrancemum plant. It has been shown that the use of plants derived from the group of Versicolol and Grebionis species provides hybrid progeny and self-pollinated progeny.

[0187] Table 7D shows the chromosome counts of the selected intergeneric hybrid plants shown in Table 7C.

[0188] As shown in Table 7D, for all hybrid plants that developed, the chromosome count was expected to be approximately 2n = 27, but surprisingly there was a wide variation in the observed number .
Example 12-I. as a female parent Intergeneric crosses using versicolol and Grevionis species and argyrancemum diploid as male parent

[0189] The results of the cross shown in Example 11 were promising, so I. It was decided to try similar crosses using versicolol and Grevionis species and Argyranthemum diploid as male parents. Table 8A shows I.V. as a female parent. The number of head-like inflorescences pollinated with versicolol and Grevionis species and three argyrancemum diploid parents is shown.

  [0190] As shown in Table 8A, I. A total of 26 scaly inflorescences were pollinated using versicolol and Grevionis species and three argyrancemum diploid parents.

[0191] Table 8B shows I.V. The number of embryos rescued from crosses using versicolol and Grebionis species and three argyrancemum diploid parents is shown.

  [0192] As shown in Table 8B, I.I. A total of 604 embryos were rescued from crosses using versicolol and Grebionis species and three argyrancemum diploid parents.

[0193] Table 8C shows I.V. The number of self-pollinated plants and intergeneric hybrid plants developed from crosses using versicolol and Grevionis species and three argyrancemum diploid parents is shown.

  [0194] As shown in Table 8C, I.I. A total of 196 self-pollinated plants and 13 intergeneric hybrid plants were developed from crosses using versicolol and Grevionis species and 3 argyrancemum diploid parents.

[0195] The work performed in Tables 8A, B, and C was performed as I.V. as a female parent crossed with a male diploid Argyranthemum plant. It has been shown that the use of plants derived from the group of Versicolol and Grebionis species has yielded hybrid progeny and self-pollinated progeny.
Example 13-Use of a triploid hybrid for backcrossing

実施例１４−植物表現型を改変するためのウイルス及びウイロイドの導入 [0196] Almost all triploid hybrids developed from heterotetraploid Argyranthemum x ismeria versicolol hybrids were male sterile (no pollen). Occasionally, however, plants with some pollen were produced. Hybridization was performed according to previously described standard procedures, and 2 hybrid plants were developed from 18 pollinated heads as shown in Table 9. This result suggests that some heteroploid intergeneric hybrid plants have the ability as male parents to contribute to further breeding tasks that are important for the further development of new varieties. It was amazing.

Example 14-Introduction of virus and viroid to modify plant phenotype

  [0197] Chrysanthemum B virus (CVB) and chrysanthemum dwarf viroid (CSVd) have been reported to infect Argyranthemum and many other species of Asteraceae. Rahman, M .; Infection with both CVB and CSVd alone or in combination with CVB and CSVd alone, in Argyranthemum, as reported by Virus and viroid studies in the marguerite Daisy, PhD thesis, University of Science 2007. It brought about morphological changes. Morphological changes induced by these infectious agents ranged from changes in flower color and speed of flowering to changes in plant habits. Several adiploid hybrid plants (lines: heterotetraploid argyransemema x ismeria versicolol and heteroploid argylansemema x grebionis coronaria) infected with CSVd and CVB were developed by the method described previously. Selected to graft on Argyranthemum plants. Argyranthemum plants infected with these two organisms were tested twice (after 6 months interval) and confirmed to be infected with CSVd and CVB based on standard ELISA and PCR based techniques at both time points. . After planting the roots of a triploid plant to the roots of a CSVd / CVB positive algillansemmum plant, using standard potting media and cultivation practices (eg, Hamrick, D., 2003) Planted in pots of 10 cm diameter. One month later, the grafted plant was planted in a larger pot and an Argyranthemum plant was excised from the haploid hybrid using a scalpel. Two more months later, the grafted plants were tested for CSVd and CVB infection. All grafted plants tested were positive for both CSVd and CVB.

[0198] To compare the morphology of infected and uninfected allotriploid hybrids, experiments were performed by breeding and growing several accessions using standard growth procedures. Observations were made during the growth test and at its completion. The results are shown in Table 10.

  [0199] As shown in Table 10, the results show that the heteroploid intergeneric hybrids infected with CSVd and CVB can show morphological changes in characteristics such as flower color, behavior and speed of flowering, Many of these changes have shown benefits for commercial production.

  [0200] Although several exemplary aspects and embodiments have been discussed above, those skilled in the art will recognize these specific modifications, substitutions, additions and sub-combinations. Accordingly, the following appended claims and any claims subsequently introduced are intended to encompass all such modifications, substitutions, additions and subcombinations as included within their true spirit and scope. It is intended to be interpreted.

[0201] The use of the terms "a", "an" and "the" and similar indicating objects in the context of describing the present invention (especially in the context of the following claims) is indicated elsewhere herein. Should be construed to include both the singular and the plural unless it is otherwise inconsistent with the context. The terms “comprising”, “having”, “including” and “containing” are intended to be open-ended terms unless otherwise specified (ie It should be construed as "not including but including"). The description of a range of values herein is intended only to serve as an abbreviation for individually referring to each distinct value included in the range, unless otherwise specified herein. Is incorporated herein as if it were individually described herein. For example, when ranges 10-15 are disclosed, 11, 12, 13, and 14 are also disclosed. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples or exemplary terms (e.g., "such as") presented herein are merely for the purpose of clarifying the invention and, unless otherwise claimed, delimit the scope of the invention. It is not limited. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

Claims (25)

  An intergeneric hybrid plant produced from a cross between a tetraploid argylancemum plant as a male parent and a female parent plant selected from the group consisting of Ismeria versicolol and Grevionis species.   The plant part of the intergeneric hybrid plant according to claim 1.   From protoplasts, leaves, stems, roots, root tips, cocoons, pistils, seeds, embryos, pollen, ovules, cotyledons, hypocotyls, heads, inflorescences, tubular flowers, shoots, shoots, tissues, petiole, cells and meristem cells The plant part according to claim 2, which is selected from the group consisting of:   An intergeneric hybrid plant or plant part thereof that is clonally propagated from the plant according to claim 1. a. Crossing a tetraploid argylancemum plant as a male parent with a plant selected from the group consisting of Ismeria versicolol and Grebionis species as a female parent;
b. Producing an embryo from the cross;
c. Using embryo rescue for said embryo;
d. Obtaining an intergeneric hybrid plant grown from the embryo.   Crossing includes sub-steps of collecting pollen from the male parent and sub-step of pollinating the flower on the female parent using the pollen, and rescues embryos resulting from the pollination in tissue culture; The method of claim 5. Using an embryo rescue for an embryo produced from a cross with a tetraploid Argyranthemum plant as a male parent and a plant selected from the group consisting of Ismeria versicolol and Grebionis species as a female parent, An intergeneric hybrid plant or a part thereof produced by growing a plant. Collecting pollen from a tetraploid Argyranthemum plant as a male parent and using this pollen to cross pollinate a flower selected from the group consisting of Ismeria versicolol and Grebionis species as a female parent An intergeneric hybrid plant or a part thereof produced by growing the embryo using embryo rescue to the produced embryo . a. Obtaining a cutting of an intergeneric hybrid plant produced from a cross between a tetraploid argylancemum plant as a male parent and a plant selected from the group consisting of Ismeria versicolol and Grebionis species as a female parent;
b. Growing the cuttings to obtain an intergeneric hybrid plant.   10. The method of claim 9, further comprising applying a plant hormone composition to the base of the cutting to induce root formation. Produced by growing cuttings of intergeneric hybrid plants produced from crosses of heterotetraploid argylancemum plants as male parents and plants selected from the group consisting of Ismeria versicolol and Grebionis species as female parents Intergeneric hybrid plants or parts thereof.   6. The method of claim 5, further comprising the step of producing a tetraploid argylan semam plant by increasing the chromosome number of the argylan semam plant prior to step a. Increasing the number of chromosomes in the Argyranthemum plant to produce a tetraploid,
a ′ . A sub-step of growing an Argyranthemum plant;
b ′ . Applying an anti-mitotic agent to the plant;
c ′ . A substep of budding shoots from the plant;
d ′ . A sub-step of selecting shoots of heterotetraploid,
e ' . A sub-step of evaluating the shoot chromosome set;
f ′ . A sub-step of growing the shoots to produce a tetraploid argylancemum plant;
g ′ . And a sub-step of examining chromosomal stability of said tetraploid argylancemum plant. A heteroploid argylancemum plant produced by increasing the number of chromosomes of an argylancemum plant , wherein the teuploid argylancemam plant is selected from the group consisting of Ismeria versicolol and Grebionis species as female parents. A tetraploid argyrancemum plant that is used as a male parent in a cross with a plant that uses embryo rescue for embryos produced from the cross to obtain an intergeneric hybrid plant .   From protoplasts, leaves, stems, roots, root tips, cocoons, pistils, seeds, embryos, pollen, ovules, cotyledons, hypocotyls, heads, inflorescences, tubular flowers, shoots, shoots, tissues, petiole, cells and meristem cells 15. A plant part of an aneuploid argylancemum plant according to claim 14, selected from the group consisting of:   An Argyranthemum plant or part thereof that is clonally propagated from the plant part according to claim 15.   15. A heteroploid plant according to claim 14, having a chromosome number of 32, 33, 34, 35, 36, 37, 38 or 39.   The intergeneric hybrid plant according to claim 1 having a chromosome number of 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26 or 27.   The intergeneric hybrid plant according to claim 1, which is infected with chrysanthemum dwarf viroid (CSVd) and chrysanthemum B virus (CVB). They are infected with chrysanthemum stunt viroid (CSVd) and chrysanthemum B virus (CVB), a intergeneric hybrid plants, the Arugiransemamu plant different tetraploid as the female parent, I. Intergeneric hybrid plants produced by growing embryos using embryo rescue on embryos produced from crosses with hybrid male parents obtained from cross-breeding of plants derived from the group of Versicolol and Grebionis species .   An intergeneric hybrid plant produced from a cross between a diploid argyranthemum plant as a male parent and a female parent plant selected from the group consisting of Ismeria versicolol and Grebionis species. a. Crossing a diploid Argyranthemum plant as a male parent with a plant selected from the group consisting of Ismeria versicolol and Grebionis species as a female parent;
b. Producing an embryo from the cross;
c. Using embryo rescue for said embryo;
d. Obtaining an intergeneric hybrid plant grown from the embryo. Crossing includes sub-steps of collecting pollen from the male parent and sub-steps of pollinating flowers on the female parent using the pollen, and rescues embryos resulting from the pollination in tissue culture; The method of claim 22 . Growing the diploid Argyranthemum plant as a male parent using embryo rescue to an embryo produced from a cross with a plant selected from the group consisting of Ismeria versicolol and Grebionis species as a female parent intergeneric hybrid plant or parts thereof produced by. Produced from a cross , including collecting pollen from a diploid Argyranthemum plant as a male parent and using this pollen to pollinate a flower selected from the group consisting of Ismeria versicolol and Grevionis species as a female parent. An intergeneric hybrid plant or a part thereof produced by growing the embryo using embryo rescue on the developed embryo.