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for invention","granted":false,"earliest_filing_date":"2007-05-11","application_expiry_date":"2012-10-02","has_disclaimer":false,"patent_status":"DISCONTINUED","publication_count":2,"has_spc":false,"has_grant_event":false,"has_entry_into_national_phase":false},"abstract":{"en":[{"text":"The present invention provides a polypeptide in which the amino acid at position 45 of an amino acid sequence represented by SEQ ID NO: 1 is substituted with a hydrophilic amino acid; or a polypeptide having deletion, insertion, substitution, or addition of one or several amino acids other than the amino acid at position 45 in the above amino acid sequence. This makes it possible to realize the production and selection of a cleistogamous plant which has a normal fertility and is suitable for practical use.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"}]},"abstract_lang":["en"],"has_abstract":true,"claim":{"en":[{"text":"A polypeptide which gives cleistogamy to a plant, wherein the polypeptide is: (A) a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 in the amino acid sequence of a class-B MADS-box protein with a hydrophilic amino acid; or (B) a polypeptide comprising an amino acid sequence having deletion, insertion, substitution, or addition of one or several amino acids other than the amino acid at position 45 in the amino acid sequence of the polypeptide (A).","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"The polypeptide according to claim 1, wherein the hydrophilic amino acid is threonine.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A polynucleotide encoding the polypeptide according to claim 1.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A plant expressing the polypeptide according to claim 1.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A plant expressing the polynucleotide according to claim 3.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A method for producing a plant to which cleistogamy is given, comprising any one of:\n (I) the step of inducing mutation in a gene that encodes a class-B MADS-box protein, wherein the mutation corresponds to an amino acid substitution at position 45 of the class-B MADS-box protein; \n (II) the step of introducing into a plant a gene being capable of inducing mutation in a gene that encodes a class-B MADS-box protein, wherein the mutation corresponds to an amino acid substitution at position 45 of the class-B MADS-box protein; and \n (III) the step of suppressing expression of genes encoding class-B MADS-box protein other than a polypeptide having an amino acid sequence represented by SEQ ID NO: 1.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A kit for producing a plant to which cleistogamy is given, wherein the kit comprises the polynucleotide according to claim 3.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A fragment of the polynucleotide according to claim 3, or an oligonucleotide comprising a complementary sequence of the base sequence of the fragment.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A method for screening a plant to which cleistogamy is given, comprising the step of hybridizing the oligonucleotide according to claim 8 with cDNA derived from a target plant.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A kit for screening a plant to which cleistogamy is given, wherein the kit comprises the oligonucleotide according to claim 8.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"An antibody being capable of binding specifically to the polypeptide according to claim 1.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A method for screening a plant to which cleistogamy is given, comprising the step of incubating the antibody according to claim 11 with a tissue extract derived from a target plant.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"},{"text":"A kit for screening a plant to which cleistogamy is given, wherein the kit comprises the antibody according to claim 11.","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"}]},"claim_lang":["en"],"has_claim":true,"description":{"en":{"text":"TECHNICAL FIELD The present invention relates to a method for producing a mother plant practically useful for production of genetically modified rice. In particular, the present invention relates to a protein which gives cleistogamy to a plant, a gene encoding the protein, and use of the protein and the gene. BACKGROUND ART Commercialization of genetically modified crops has been demanded because of their great potentiality. However, the commercialization of genetically modified crops has been difficult since producers and consumers have strong concerns about the gene flow caused by crossing with the related wild species or the neighboring cultivars for the reasons that effect of transgenes on an environment for long period and others is not evaluated. Until effect of transgenes on an environment for long period and others is evaluated, preventive measures against the gene flow have been required for safety ensuring, from the view point of precautionary principle. Generally, rice is a plant that performs self-pollination at a high frequency, but can perform cross-pollination at a low frequency since rice releases its pollen grains to the outside after its flowering. In outside cultivation, the ensuring of a distance between the genetically modified crops and the surrounding plants or the adjustment of a cultivation time of genetically modified crops (e.g. shift of their flowering stage) has been required to prevent genetically modified crops from being crossed with the surrounding wild plants or the related crops due to the release of pollen grains of the genetically modified crops. If these approaches are impossible, flower picking, removal of stamens, bagging of flowers, installation of wind-breaking nets, etc. are performed. However, they require enormous efforts, and the development of techniques for preventing the release of pollen grains of rice has been demanded. Techniques for preventing the gene flow caused by the release of pollen grains of the genetically modified plants have been proposed such as maternal inheritance by chloroplast transformation, apomixis, and male sterility. However, chloroplast transformation and apomixes of rice have not been practically used. In addition, male-sterile rice exists but is not practically used because it requires artificial pollination for harvesting of its seeds. As another technique for preventing the release of pollen grains, cleistogamy is known. As to barley, genetic analysis using cleistogamous cultivars has been made (see Patent Document 1 and Non-Patent Document 1, for example). It is revealed that cleistogamous cultivars exist according to the study on barley, and cleistogamy is controlled by small number of genes according to the genetic analysis. Some reports on cleistogamous rice have been made so far (see Non-Patent Documents 2 and 3). Further, as rice having a trait similar to cleistogamy, \"sheathed-panicle rice\" has been reported (see Non-Patent Document 4). Some other reports have been made on non-flower-opening rice caused by morphological change of floral organs (see Non-Patent Documents 5 and 6).\n [Patent Document 1] \nJapanese Unexamined Patent Publication No. 2005-229854 (published on September 2, 2005 ) [Non-Patent Document 1] \n Theor Appl Genet 109, 480-7(2004 ) [Non-Patent Document 2] \n J Fac Agric Hokkaido Univ. 53, 72-130(1963 ) [Non-Patent Document 3] \n Current Science 50, 419-420(1981 ) [Non-Patent Document 4] \n Rice Genetics Newsletter 3, 62-63(1986 ) [Non-Patent Document 5] \n Development 130, 705-718 (2003 ) [Non-Patent Document 6] \n Plant Cell Physiol.43, 130-135 (2002 ) DISCLOSURE OF INVENTION As cleistogamous rice that has been reported so far, 'Heieidaikoku (d7)' mutant is presumed to cause other growth disturbances (e.g. dwarfism and short grain) due to pleiotropic expressions of the causal gene. Meanwhile, 'Dhundhuni' is presumed to show a complex inheritance pattern for cleistogamy (there are many genes associated with cleistogamy, and maternal inheritance is also involved in cleistogamy). Thus, both the 'Heieidaikoku (d7)' mutant and the 'Dhundhuni' are difficult to be used as mother plants for crossing. \"Sheathed-panicle rice\", of which panicle is surrounded by sheath of flag leaf, is presumed to incur the risk of diseases after grain ripening and complicated harvesting. Therefore, \"sheathed-panicle rice\" is not suitable for practical use. As rice exhibiting cleistogamy caused by morphological change of floral organ \"lodicule\" (corresponding to petal of dicotyledon) that functions as a locomotive of flowering, superwoman1 (spw1) mutant, Act::OsMADS3 transgenic plants, and others have been reported. However, they have morphologically changed stamens and severe sterility and are not suitable for practical use. Thus, the development of cultivars having cleistogamy capable of preventing the release of pollen grains has been demanded. However, there exist no cleistogamous cultivars of rice for practical use in Japan. In addition, the existence of a gene giving cleistogamy and the mechanism to control cleistogamy were not revealed. The present invention has been attained in view of the above problems, and an object of the present invention is to provide a technique for producing and selecting a practical cleistogamous plant that has normal fertility but does not affect agronomic traits such as plant type, and heading. The inventors of the present invention obtained the sole cleistogamous rice mutant, superwoman1-cleistogamy (spw1-cls), from a mutant population mutagenized with chemical mutagen N-methyl-N-nitrosourea (MNU) by using japonica rice cultivar 'Taichung 65' as original cultivar. Specifically, the inventors of the present invention (1) selected from the above population mutant lines in which stamens were not exserted out of glume at flowering stage, and (2) selected a line that was normally fertile from the selected lines. Further, by map-based cloning of rice spw1-cls mutant, the inventors verified that cleistogamy was caused by mutation of a gene encoding SUPERWOMAN (SPW1) that is a class-B MADS-box protein, thereby accomplishing the present invention. That is, a polypeptide of the present invention is a polypeptide which gives cleistogamy to a plant, wherein the polypeptide is: (A) a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 in the amino acid sequence of a class-B MADS-box protein with a hydrophilic amino acid; or (B) a polypeptide comprising an amino acid sequence having deletion, insertion, substitution, or addition of one or several amino acids other than the amino acid at position 45 in an amino acid sequence of the polypeptide (A). A polypeptide of the present invention is capable of giving cleistogamy to a plant. Considering the fact that the superwoman1 (spw1) mutant has morphologically changed stamen and severe sterility (see Non-Patent Document 5), it could not be expected at all that substitution of one amino acid in SPW1 gave cleistogamy to a plant. A polypeptide of the present invention may be a polypeptide which gives cleistogamy to a plant, wherein the polypeptide is: (A) a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 in the amino acid sequence of a class-B MADS-box protein of monocotyledon with a hydrophilic amino acid; or (B) a polypeptide comprising an amino acid sequence having deletion, insertion, substitution, or addition of one or several amino acids other than the amino acid at position 45 in an amino acid sequence of the polypeptide (A). A polypeptide of the present invention may be a polypeptide which gives cleistogamy to a plant, wherein the polypeptide is: (A) a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 in an amino acid sequence represented by SEQ ID NO: 1, 6, 8, 10, 12, 14, 16, 18 or 20 with a hydrophilic amino acid; or (B) a polypeptide comprising an amino acid sequence having deletion, insertion, substitution, or addition of one or several amino acids other than the amino acid at position 45 in the amino acid sequence of the polypeptide (A). A polypeptide of the present invention may be a polypeptide which gives cleistogamy to a plant, wherein the polypeptide is: (A) a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 in an amino acid sequence represented by SEQ ID NO: 1, 6, or 8 with a hydrophilic amino acid; or (B) a polypeptide comprising an amino acid sequence having deletion, insertion, substitution, or addition of one or several amino acids other than the amino acid at position 45 in the amino acid sequence of the polypeptide (A). A polypeptide of the present invention is preferably such that the hydrophilic amino acid is threonine. A polynucleotide of the present invention encodes the above polypeptide. A polynucleotide of the present invention may be a polynucleotide that has a base sequence represented by SEQ ID NO: 5, or a polynucleotide that is capable of hybridizing under stringent conditions with the polynucleotide having a base sequence represented by SEQ ID NO: 5 and gives cleistogamy to a plant. A plant of the present invention expresses the above polypeptide. A plant of the present invention expresses the above polynucleotide. A plant producing method of the present invention is a method for producing a plant to which cleistogamy is given, comprising any one of:\n (I) the step of inducing mutation in a gene that encodes the class-B MADS-box protein, wherein the mutation corresponds to an amino acid substitution at position 45 of a class-B MADS-box protein; (II) the step of introducing into a plant a gene being capable of inducing mutation in a gene that encodes a class-B MADS-box protein, wherein the mutation corresponds to an amino acid substitution at position 45 of the class-B MADS-box protein; and (III) the step of suppressing expression of genes encoding class-B MADS-box protein other than SPW1 (polypeptide having an amino acid sequence represented by SEQ ID NO: 1). A plant production kit of the present invention is a kit for producing a plant to which cleistogamy is given, wherein the kit comprises the above polynucleotide. An oligonucleotide of the present invention is a fragment of the above polynucleotide, or an oligonucleotide comprising a complementary sequence of the base sequence of the fragment. An oligonucleotide of the present invention may be an oligonucleotide that has a base sequence represented by SEQ ID NO: 23. An oligonucleotide set of the present invention comprises a set of oligonucleotides including a fragment of the above polynucleotide, or an oligonucleotide comprising a complementary sequence of the base sequence of the fragment. An oligonucleotide set of the present invention comprises an oligonucleotide that has a base sequence represented by SEQ ID NO: 22 and oligonucleotide that has a base sequence represented by SEQ ID NO: 23. A screening method of the present invention is a method for screening a plant to which cleistogamy is given, comprising the step of hybridizing the above oligonucleotide with cDNA derived from a target plant. A screening method of the present invention is a method for screening a plant to which cleistogamy is given, comprising the step of hybridizing an oligonucleotide having a base sequence represented by SEQ ID NO: 23 with cDNA derived from a target plant. A screening method of the present invention preferably further comprises the step of hybridizing an oligonucleotide having a base sequence represented by SEQ ID NO: 22 with cDNA derived from a target plant. A screening kit of the present invention is a kit for screening a plant to which cleistogamy is given, wherein the kit comprises the above oligonucleotide. A screening kit of the present invention is a kit for screening a plant to which cleistogamy is given, wherein the kit comprises an oligonucleotide having a base sequence represented by SEQ ID NO: 23. A screening kit of the present invention preferably further comprises an oligonucleotide having a base sequence represented by SEQ ID NO: 22. An antibody of the present invention is capable of binding specifically to the above polypeptide. A screening method of the present invention is a method for screening a plant to which cleistogamy is given, comprising the step of incubating the above antibody with a tissue extract derived from a target plant. A screening kit of the present invention is a kit for screening a plant to which cleistogamy is given, wherein the kit comprises the above antibody. Additional objects, features, and strengths of the present invention will be made clear by the description below. Further, the advantages of the present invention will be evident from the following explanation in reference to the drawings. BRIEF DESCRIPTION OF DRAWINGS \n Fig. 1 is a view showing phenotypes of cleistogamous mutant spw1-cls. Fig. 2 is a view showing comparison in plant type, panicle, and rice grain between cleistogamous mutant spw1-cls and a wild type. Fig. 3 is a view showing structural change of floral organs in cleistogamous mutant spw1-cls and a causal gene of spw1-cls mutation. Fig. 4 is a view showing function complementation of cleistogamous mutantspw1-cls by a wild-type SPW gene. Fig. 5 is a view showing the analysis result of influences on OsMADS2 and OsMADS4 in terms of cleistogamy. BEST MODE FOR CARRYING OUT THE INVENTION [1: Polypeptide and polynucleotide] [1-1] Polypeptide The present invention provides a polypeptide which gives cleistogamy to a plant. A polypeptide of the present invention is a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 with a hydrophilic amino acid in the amino acid sequence of class-B MADS-box protein in a plant, or its mutant. The plant is preferably a monocotyledon, more preferably a Poaceae plant (e.g. rice, barley, wheat, rye, oat, proso millet, foxtail millet, or corn), most preferably rice, barley, or wheat. Class-B MADS-box proteins in plants are classified into AP3 proteins and PI proteins. The AP3 proteins are SPW 1 of rice (SEQ ID NO: 1), HvAP3 of barley (SEQ ID NO: 10), TaMADS#51 of wheat (SEQ ID NO: 14), and TaMADS#82 of wheat (SEQ ID NO: 16). The PI proteins are OsMADS2 of rice (SEQ ID NO: 6), OsMADS4 of rice (SEQ ID NO: 8), HvPI of barley (SEQ ID NO: 12), WPI1 of wheat (SEQ ID NO: 18), and WPI2 of wheat (SEQ ID NO: 20). However, the class-B MADS-box proteins in plants are not limited to these proteins. Note that cDNA sequences of these proteins are as follows: SEQ ID NO: 2 (SPW of rice), SEQ ID NO: 11 (HvAP3 of barley), SEQ ID NO: 15 (TaMADS#51 of wheat), SEQ ID NO: 17 (TaMADS#82 of wheat), SEQ ID NO: 7 (OsMADS2 of rice), SEQ ID NO: 9 (OsMADS4 of rice), SEQ ID NO: 13 (HvPI of barley), SEQ ID NO: 19 (WPI1 of wheat), and SEQ ID NO: 21 (WPI2 of wheat). A polypeptide of the present invention comprises an amino acid sequence having substitution of an amino acid at position 45 in the amino acid sequence of such a class-B MADS-box protein with a hydrophilic amino acid. As to the plant produced by the inventors of the present invention, the amino acid at position 45, isoleucine, in SUPERWOMAN1 (SPW1) protein, which is a class-B MADS-box protein of rice, is substituted with threonine (SEQ ID NO: 4). However, the substitution of the amino acid at position 45 in the polypeptide of the present invention is not limited to this. Apart from threonine, the hydrophilic amino acid is glycine, serin, cysteine, aspartic acid, asparagine, glutamic acid, glutamine, lysine, arginine, or histidine. Considering the fact that the superwoman1 (spw1) mutant (see Non-Patent Document 5) has morphologically changed stamens and severe sterility, it cannot be anticipated that cleistogamy is given to a plant by substituting one amino acid of SPW1 by another. As used herein, the term \"polypeptide\" can refer to \"peptide\" or \"protein\". A \"fragment\" of the polypeptide means a partial fragment of the polypeptide. A polypeptide of the present invention may be isolated from natural resources or chemically synthesized. A polypeptide of the present invention may be a native polypeptide taken from natural environment, or a recombinant polypeptide that is expressed in a host cell and is substantially purified by any appropriate technique. Further, a polypeptide of the present invention may include additional polypeptide. Examples of the additional polypeptide include an epitope-labeled polypeptide which is well-known in the art, such as His, Myc, or Flag. As used herein in association with a polypeptide or a protein, the term \"mutant\" means a polypeptide having the activity of giving cleistogamy to a plant. Such a mutant is preferably a polypeptide having deletion, insertion, substitution, or addition of at least one amino acid in an amino acid sequence of the original polypeptide, more preferably a polypeptide having deletion, insertion, substitution, or addition of one or several amino acids in an amino acid sequence of the original polypeptide. Especially, \"neutral\" amino acid substitution is further preferable since it generally has little effect on the activity of the polypeptide. In the polypeptide of the present invention, a preferable mutant can have substitution, deletion, or addition of conservative or non-conservative amino acid, which does not change the activity of the polypeptide of the present invention of giving cleistogamy to a plant. It is well known in the art that some amino acids in the amino acid sequence constituting a polypeptide can be easily modified without significant effect on the structure or function of this polypeptide. In addition to the fact that amino acids can be artificially modified, it is also well known that in natural proteins there exists a mutant which does not significantly change the structure or function of the protein. A person skilled in the art can easily mutate one or several amino acids in an amino acid sequence of a polypeptide by using a well-known technique. According to the method described herein, it is possible to easily determine whether a produced mutant has a desired activity. Thus, a polypeptide of the present embodiment is a polypeptide having the activity of giving cleistogamy to a plant, and a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 in the amino acid sequence of a class-B MADS-box protein with a hydrophilic amino acid, or its mutant. In one embodiment, a polypeptide of the present embodiment can comprise (A) an amino acid sequence having substitution of an amino acid at position 45 in an amino acid sequence represented by SEQ ID NO: 1, 6, 8, 10, 12, 14, 16, 18 or 20 with a hydrophilic amino acid; or (B) an amino acid sequence having deletion, insertion, substitution or addition of one or several amino acids other than the amino acid at position 45 in the amino acid sequence (A). In another embodiment, a polypeptide of the present invention can comprise (C) an amino acid sequence having substitution of an amino acid at position 45 in an amino acid sequence represented by SEQ ID NO: 1, 6, or 8 with a hydrophilic amino acid, or (D) an amino acid sequence having deletion, insertion, substitution, or addition of one or several amino acids other than the amino acid at position 45 in the amino acid sequence (C). [1-2] Polynucleotide The present invention also provides a polynucleotide encoding a polypeptide that gives cleistogamy to a plant. In an embodiment, a polynucleotide of the present invention encodes a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 in the amino acid sequence of class-B MADS-box protein in a plant with a hydrophilic amino acid, or its mutant. As used herein, the term \"polynucleotide\" refers to \"gene\", \"nucleic acid\", or \"nucleic acid molecule\", and means a polymer of nucleotides. As used herein, the term \"base sequence\" refers to \"DNA sequence\", \"nucleic acid sequence\", or \"nucleotide sequence\". A polynucleotide of the present invention can exist in the form of DNA (e.g. cDNA or genome DNA) or RNA (e.g. mRNA). The DNA may be single stranded or double stranded. The single strand DNA or RNA may be a coding strand (also known as a sense strand) or a non-coding strand (also known as an anti-sense strand). In one aspect, a polynucleotide of the present embodiment can be a polynucleotide in which the base at position 348 or 349 in a base sequence represented by SEQ ID NO: 2 is substituted, or its mutant. The base at position 348 or 349 in the base sequence represented by SEQ ID NO: 2 is substituted, whereby the amino acid at position 45 of the encoded polypeptide, a hydrophobic amino acid, is substituted with a hydrophilic amino acid. Preferably, a polynucleotide of the present embodiment can be a polynucleotide having a base sequence represented by SEQ ID NO: 5, or its mutant. As used herein in association with DNA or polynucleotide, the term \"mutant\" means a polynucleotide encoding a polypeptide having the activity of giving cleistogamy to a plant. A \"polynucleotide mutant having a base sequence represented by SEQ ID NO: 5\" means a polynucleotide having deletion, insertion, substitution, or addition of one or several bases in a base sequence represented by SEQ ID NO: 5. Alternatively, the \"polynucleotide mutant having a base sequence represented by SEQ ID NO: 5\" means a polynucleotide that hybridizes under stringent conditions with a polynucleotide comprising a base sequence complementary to the base sequence represented by SEQ ID NO: 2. Hybridization can be performed by a well-known method, for example, according to the procedure described in Sambrook et al., Molecular Cloning, A Laboratory Manual, 2d Ed., Cold Spring Harbor Laboratory (1989 ). As a rule, the level of stringency increases (more difficult to hybridize) with increase in temperature and decrease in salt concentration. Therefore, it is possible to obtain more homologous polynucleotide. An appropriate hybridization temperature varies depending upon a base sequence and its length. For example, in a case where a DNA fragment consisting of 18 bases that encode 6 amino acids is used as a probe, the hybridization temperature is preferably not higher than 50 °C. As used herein, the \"stringent\" hybridization conditions mean overnight incubation at 42 °C in a hybridization solution (including 50% formamide, 5 × SSC (150mM NaCl, 15mM trisodium citrate), 50 mM sodium phosphate (pH7.6), 5 x Denhardt's solution, 10% dextran sulfate, and 20 µg/ml of denatured, sheared salmon sperm DNA), followed by washing filters in a 0.1 × SSC at approximately 65 °C. A polynucleotide which hybridizes with a \"portion\" of polynucleotide means a polynucleotide (either DNA or RNA) that hybridizes with at least approximately 15 nucleotides (nt), more preferably at least approximately 20 nt, still more preferably at least approximately 30 nt, even more preferably longer than approximately 30 nt, of the entire length of the reference polynucleotide. A polynucleotide (oligonucleotide) that hybridizes with such \"portion\" of polynucleotide is useful as a probe (including primer) discussed in detail herein. A polynucleotide of the present invention can be fused with a polynucleotide that encodes the foregoing tag label (tag sequence or marker sequence) at its 5'end or 3'end. Thus, a polynucleotide of the present embodiment is a polynucleotide that encodes a polypeptide having the activity of giving cleistogamy to a plant, and the polynucleotide encodes a polypeptide comprising an amino acid sequence having substitution of an amino acid at position 45 in the amino acid sequence of a class-B MADS-box protein in a plant with a hydrophilic amino acid, or its mutant. In one aspect, a polynucleotide of the present invention is a polynucleotide encoding a polypeptide that gives cleistogamy to a plant, and can be (A) a polynucleotide having a base sequence represented by SEQ ID NO: 5; (B) a polynucleotide having deletion, insertion, substitution, or addition of one or several bases in the base sequence in (A); or (C) a polynucleotide capable of hybridizing with the polynucleotide (A) under stringent conditions. Note that it can be easily understood by a person skilled in the art who reads the specification of the present application that although a polynucleotide of the present invention has been explained taking a base sequence represented by SEQ ID NO: 5 as an example, but a polynucleotide of the present invention is not limited to this. [2: Use of polypeptide according to the present invention] [2-1] Plant to which cleistogamy is given and method for producing the plant In one aspect, the present invention provides a plant that expresses a polypeptide of the present invention. In another aspect, the present invention provides a plant that expresses a polynucleotide of the present invention. In an embodiment, a plant of the present invention can be produced by introducing spw1-cls mutation into a plant that does not have spw1-cls mutation. Note that (i) an amino acid substitution of an amino acid at position 45 of a class-B MADS-box protein with a hydrophilic amino acid and (ii) a base substitution corresponding to the amino acid substitution in a gene that encodes the protein are herein referred to as \"spw1-cls mutation\" as necessary, and a plant to which cleistogamy is given by these substitutions is referred to as \"spw1-cls mutant\". Considering the fact that an amino acid near the amino acid at position 45 of the class-B MADS-box protein is mainly constituted by a hydrophobic amino acid, the scope of the present invention should encompass, as the spw1-cls mutant, a mutant obtained by substituting the hydrophobic amino acid other than the amino acid at position 45 with a hydrophilic amino acid. That is, a method for producing a plant according to the present embodiment comprises the step of expressing a polypeptide of the present invention in a plant. It is preferable that the step of expressing a polypeptide of the present invention in a plant is a step of introducing spw1-cls mutation into a plant. The step of introducing the spw1-cls mutation into a plant can be:\n (I) the step of inducing mutation in a gene that encodes the class-B MADS-box protein, wherein the mutation corresponds to an amino acid substitution at position 45 of the class-B MADS-box protein; or (II) the step of introducing into a plant a gene being capable of inducing mutation in a gene that encodes a class-B MADS-box protein, wherein the mutation corresponds to an amino acid substitution at position 45 of the class-B MADS-box protein. \nThat is, the step (I) is a step of generating a polynucleotide of the present invention in a plant, and the step (II) is a step of introducing a polynucleotide of the present invention into a plant. The step (I) includes (i) introduction of spw1-cls mutation by gene targeting, or (ii) introduction of spw1-cls mutation by the treatment with a mutagen. However, these are not only possibility. The step (II) includes (iii) introduction of spw1-cls mutant gene or (iv) introduction of spw1-cls mutation by crossing. However, these are not only possibility.\n (i) Introduction of spw1-cls mutation by gene targeting\n \nIt is possible to produce a cleistogamous plant by producing a class-B MADS-box protein mutant having deletion, insertion, substitution, or addition of one or several amino acids with respect to an amino acid at a target position or its neighboring position, by using a well-known technique in the art (e.g. gene targeting method ( Terada et al., (2002), Efficient gene targeting by homologous recombination in rice. Nat Biotechnol. 20(10): 1030-4 )).\n (ii) Introduction of spw1-cls mutation by a process using mutagen\n \nIt is possible to produce a cleistogamous plant by processing a target plant with a mutagen so that a class B MADS-box gene locus in the target plant is mutagenized. The inventors of the present invention processed Taichung 65 ( japonica rice cultivar) with a chemical mutagen N-methyl-N-nitrosourea (MNU) to obtain a mutant population, selected mutant lines in which stamens were not exserted out of glumes at flowering stage from the mutant population, and further selected a line that was normally fertile from the selected lines, so as to obtain a cleistogamous rice mutant, superwomanl-cleistogamy (spw1-cls). Further, mutation of SUPERWOMAN1 (SPW1) gene was confirmed by map-based cloning. It should be noted that a person skilled in the art can select a well-known mutagen in this field appropriately, if necessary.\n (iii) Introduction of spw1-cls mutant gene\n \nAs will be described later in Example, it is possible to produce a cleistogamous plant by introducing a genome DNA fragment having spw1-cls mutation into a loss-of-function mutant of a class-B MADS-box gene (e.g. spw1-1 or spw1-2). It should be noted that a person skilled in the art can introduce a gene into a plant successfully and express the gene stably or tissue-specifically, according to a technical knowledge in the art.\n (iv) Introduction of spw1-cls mutation by crossing\n \nIt is possible to produce a cleistogamous plant by crossing a cultivar to which cleistogamy is to be given with the spw1-cls mutant produced as described above and selecting a progeny having spw1-cls mutation homozygously from among their progenies.\n In another embodiment, a plant of the present invention can be produced by suppressing the expression of a class-B MADS-box gene other than SPW1 in a plant that does not have spw1-cls mutation. Specifically, as will be described later in Example, it is possible to produce a cleistogamous plant by suppressing the expression of a gene that specifically acts on a lodicule (e.g. OsMADS2) among class-B MADS-box genes other than SPW1. In this case, stamens are formed normally by other class-B MADS-box gene that functions redundantly. It should be noted that a person skilled in the art can successfully suppress the expression of a target gene according to a technical knowledge in the art (e.g. knockout method, antisense method, co-suppression method, and RNAi method). Thus, a person skilled in the art who reads the specification of the present application can easily produce a plant of the present invention by using a well-known technique in the art, and easily select a plant of the present invention by using an oligonucleotide and an antibody, both of which will be described in detail later. In order to screen a target plant (i.e. mutant having spw1-cls mutation), the target plant can be detected by TILLING method (a method for detecting mutation in a target gene locus by specific probe process and mismatched-site specific endonuclease process), etc. Further, it is possible to find out a plant that has amino acid substitution in an amino acid at a target position or its neighboring position or a plant having deletion, insertion, substitution or addition of one or several amino acids, by analyzing a base sequence of SPW1 gene in the obtained mutant. [2-2] Oligonucleotide (probe or primer) and use of the same The present invention provides an oligonucleotide consisting of a fragment of a polynucleotide of the present invention. As used herein, the term \"oligonucleotide\" refers to a molecule of several to several tens of nucleotides, and it is used interchangeably with \"polynucleotide\". The oligonucleotide is denoted by the number of nucleotides it contains. For example, the term dinucleotide (dimmer) or trinucleotide (trimer) is used to refer to oligonucleotides of short sequences, whereas long oligonucleotides are referred to as 30-mers or 100-mers. The oligonucleotide may be generated as a fragment of a longer polynucleotide or may be chemically synthesized. An oligonucleotide of the present invention is useful to select a plant to which cleistogamy is given. That is, by using an oligonucleotide of the present invention, it is possible to easily detect spw1-cls mutation. Therefore, with the present invention, it is possible to easily screen a plant having cleistogamy. Thus, it should be easily understood that the scope of the present invention for screening a plant having cleistogamy encompasses a screening method comprising the step of hybridizing an oligonucleotide of the present invention with a genome DNA or the like derived from a target plant, and a screening kit comprising an oligonucleotide of the present invention. That is, an oligonucleotide of the present invention includes a fragment of a polynucleotide of the present invention or an oligonucleotide comprising a complementary sequence of the base sequence of the fragment, and may be an oligonucleotide having a base sequence represented by SEQ ID NO: 23. An oligonucleotide set of the present invention includes at least one oligonucleotide described above, and may include an oligonucleotide having a base sequence represented by SEQ ID NO: 22, and an oligonucleotide having a base sequence represented by SEQ ID NO: 23. Further, a screening method of the present invention is a method for screening a plant to which cleistogamy is given, characterized by comprising the step of hybridizing an oligonucleotide of the present invention with cDNA derived from a target plant, and may comprise the step of hybridizing an oligonucleotide having a base sequence represented by SEQ ID NO: 23 with cDNA derived from a target plant. In this case, it is preferable to further comprises the step of hybridizing an oligonucleotide having a base sequence represented by SEQ ID NO: 22 with cDNA derived from a target plant. Still further, a screening kit of the present invention is a kit for screening a plant to which cleistogamy is given, the kit comprising an oligonucleotide of the present invention, and may include an oligonucleotide having a base sequence represented by SEQ ID NO: 23. In this case, it is preferable to further comprise an oligonucleotide having a base sequence represented by SEQ ID NO: 22. Examples of a method for identifying plant variety well known in the art include RFLP method, RAPD method, AFLP method, CAPS method, SSR method, ISSR method, and SNP method. A person skilled in the art who reads the specification of the present application can design a suitable oligonucleotide appropriately according to an adopted method. It is preferable to use CAPS method (especially dCAPS method), considering that sequence information of a target class-B MADS-box protein has been already known and that method to be used is merely required to detect whether the amino acid substitution at position 45 of the class-B MADS-box protein occurs. The CAPS method is a method for cutting a PCR product with a particular restriction enzyme to detect a base substitution (polymorphism) according to variations in length of the obtained fragments. In a case where there is no preferable restriction enzyme site, it is possible to newly provide a preferable restriction site by devising a primer (oligonucleotide) (dCAPS method). For example, a first oligonucleotide (5'-TGCAGAACTCGTGGTACTTGCCGGTGGAGGAGAACACCATG-3' (SEQ ID NO: 23)) has a partial base substitution introduced in a template for amplification (5'-TGCAGAACTCGTGGTACTTGCCGGTGGAGGAGAACATAATG-3 '), that is a complementary sequence of SPW1 DNA (SEQ ID NO: 2) or of SPW1 genome DNA (SEQ ID NO: 3). In order to forma pair of primers with the first oligonucleotide and be capable of amplifying a fragment containing a spw1-cls mutation site, a second oligonucleotide (e.g. 5'-CAGGGAGCTCACCGTGCTCTGCGACGCCCAGGTCGCCA-3' (SEQ ID NO: 22)) is designed. In a case where fragments amplified by PCR using the first and second oligonucleotides as a pair of primers are NcoI-processed, the amplified fragment is cut at a site (CCATGG) generated when there is spw1-cls mutation. On the other hand, the amplified fragment is not cut at a site (TCATGG) generated when there is no spw1-cls mutation. The above descriptions have explained the first oligonucleotide that is a dCAMPS marker, taking as an example an oligonucleotide having a base sequence represented by SEQ ID NO: 23. However, note that an oligonucleotide of the present invention is not limited to this. Further, the above descriptions have explained the second oligonucleotide that forms a pair of primers, taking as an example an oligonucleotide having a base sequence represented by SEQ ID NO: 22. However, an oligonucleotide of the present invention is not limited to this. Applicant of the present invention has developed a novel technique of identifying single base polymorphism (see Japanese Unexamined Patent Publication No. 2004-248635 (published on September 9, 2004 ). In an embodiment, an oligonucleotide of the present invention can be an oligonucleotide to be applied to the above technique. That is, an oligonucleotide of the present embodiment is a PCR primer for identifying a genotype of single base polymorphism, the first base from 3'end of the primer corresponds to the position of the single base polymorphism to be identified, and the third base from 3'end of the primer is substituted with a base other than one complementary to a template sequence annealed with the primer. The above substitution is preferably G to T substitution, A to C substitution, T to G substitution, or C to A substitution. A Tm value of an oligonucleotide of the present embodiment is not less than 50 °C, preferably not less than 60 °C, more preferably not less than 65 °C, most preferably 70 °C. By using an oligonucleotide of the present embodiment, it is possible to know existence of single base polymorphism (single base substitution) in a target gene from whether a target amplification product can be obtained. As used herein, the term \"primer\" means a single strand oligonucleotide that acts as an origin of template-directed DNA synthesis under appropriate conditions. A preferable length of the primer, which can be changed appropriately according to an intended use, can be generally a length corresponding to nucleotides with any number of bases in the range from 15 to 50 nucleotides, preferably 15 to 30 nucleotides. A sequence of the primer does not need to match a template sequence completely, but the sequence of the primer should be sufficiently complementary to the template sequence in order to hybridize the primer with the template. The term \"a pair of primers\" means a primer set including (i) 5' (upstream) primer which hybridizes with 5'-end of a nucleic acid sequence to be amplified and (ii) 3' (downstream) primer which hybridizes with a complement of 3'-end of the nucleic acid sequence to be amplified. The \"PCR (polymerase chain reaction)\" is described herein as an exemplary nucleic-acid amplification reaction. However, an oligonucleotide of the present embodiment can be applied to a nucleic-acid amplification reaction other than PCR. That is, an oligonucleotide of the present invention is characterized by being a fragment of an polynucleotide of the present invention or an oligonucleotide comprising a complementary sequence of the base sequence of the fragment, and applicable to a variety identification technique well known in the art. Therefore, a person skilled in the art who reads the specification of the present application easily understands that an oligonucleotide of the present invention is designed appropriately according to each technique adopted to detect spw1-cls mutation and is not limited to a particular oligonucleotide. [2-3] Antibody and use of the same The present invention provides an antibody being capable of binding specifically to a polypeptide of the present invention. An antibody of the present invention is not limited if it can bind specifically to a polypeptide of the present invention, and may be a polyclonal antibody or the like against the polypeptide. However, the antibody of the present invention is preferably a monoclonal antibody against the polypeptide of the present invention. The monoclonal antibody has the advantages of having uniform properties, being easily supplied, and being semipermanently preserved as a hybridoma, and other advantages. As used herein, the term \"antibody\" means immunoglobulin (IgA, IgD, IgE, IgG, IgM, and their Fab fragments, F(ab') 2 fragments, Fc fragments). Examples of the antibody include a polyclonal antibody, a monoclonal antibody, a single strand antibody, and an antiidiotype antibody. However, the antibody of the present invention is not limited to these antibodies. As used herein, the wording \"bind specifically to a polypeptide of the present invention\" means being capable of binding specifically to a polypeptide in which the amino acid at position 45 of the class-B MADS-box protein is substituted with a hydrophilic amino acid, but not bind to a polypeptide in which the amino acid at position 45 of the class-B MADS-box protein is not substituted. Therefore, by using the present invention, it is possible to easily screen a plant having cleistogamy. Thus, it should be easily understood that the scope of the present invention for screening a plant having cleistogamy encompasses a screening method comprising the step of incubating an antibody of the present invention with a tissue extract derived from a target plant, and a screening kit comprising an antibody of the present invention. An antibody of the present invention is characterized by being capable of binding specifically to a polypeptide of the present invention. A screening method of the present invention is a method for screening a plant to which cleistogamy is given, and characterized by comprising the step of incubating the above antibody with a tissue extract derived from a target plant. Further, a screening kit of the present invention is a kit for screening a plant to which cleistogamy is given and \ncharacterized by comprising the above antibody. As used herein, the wording \"antibody being capable of binding specifically to a polypeptide of the present invention\" means that the antibody includes complete antibody molecules and antibody fragments (e.g. Fab fragment and F(ab') 2 fragment) both of which can bind specifically to the polypeptide of the present invention. The Fab fragment and F(ab') 2 fragment lack Fc portion of a complete antibody, the Fc portion is removed more speedily through circulation, and the Fab fragment and F(ab') 2 fragment hardly have a character of non-specific tissue binding of a complete antibody ( Wahl et al., J. Nucl. Med. 24: 316-325 (1983 ) (herein incorporated by reference)) Therefore, these fragments are preferable. The \"antibody\" can be produced according to various publicly known methods (e.g. HarLow et al., Antibodies: A laboratory manual, Cold Spring Harbor Laboratory, New York (1988 ), and Iwasaki et al., Monoclonal antibody hybridoma and ELISA, Kodansha (1991 )). The monoclonal antibody can be produced by a well-known method in the art (for example, see hybridoma method ( Kohler, G. and Milstein, C., Nature 256, 495-497 (1975 )), trioma technique and human-B cell hybridoma technique ( Kozbor, Immunology Today 4,72 (1983 )) and EBV-hybridoma method ( Monoclonal Antibodies and Cancer Therapy, Alan R Liss, Inc., 77-96 (1985 ))). The peptide antibody can be also produced by a well-known method in the art (for example, see Chow, M. et al., Proc. Natl. Acad. Sci. USA 82: 910-914 ; and Bittle, F.J. et al., J. Gen. Virol. 66: 2347-2354 (1985 )). It is clear for a person skilled in the art that Fab fragments, F(ab') 2 fragments, and other fragments of an antibody of the present invention can be used according to a method disclosed herein. These fragments can be produced through cutting by proteolysis using a typical enzyme such as papain (which generates Fab fragments) or pepsin (which generates F(ab') 2 fragments). Alternatively, a fragment being capable of binding to a polypeptide of the present invention can be produced by use of recombinant DNA technique or chemical synthesis. Thus, an antibody of the present embodiment has only to include a fragment being capable of binding specifically to a polypeptide of the present invention (e.g. Fab fragment and F(ab') 2 fragment), and it should be noted that the present invention encompasses an immunoglobulin composed of (i) the fragment being capable of binding specifically to a polypeptide of the present invention and (ii) Fc fragment of another antibody. That is, an object of the present invention is to provide an antibody being capable of binding specifically to a polypeptide of the present invention and use of the same, without limitations to types of individual immunoglobulins specifically described herein (IgA, IgD, IgE, IgG, or IgM), peptide antigen producing method, and others. Therefore, it should be noted that the scope of the present invention encompasses an antibody obtained by a method other than the foregoing methods. Further, all of the academic documents and patent documents listed herein are incorporated by reference herein. [Example] [1: Methods] [1-1. Identification of cleistogamous mutant] To find cleistogamous mutants, M2 population of rice ( Oryza sativa L. subsp. japonica cv. Taichung 65) mutagenized with N-methyl-N-nitrosourea (MNU) were screened. The M2 population was cultivated under usual conditions. Wild-type rice flowers were opened at flowering stage and closed after 15 minutes. Some anthers remained outside the glumes even after the glumes were closed. Thus, first, plants in which anthers were not observed outside the glumes at flowering stage were selected. Subsequent examination reveled one fertile line in which the cleistogamy was inherited as a single recessive trait. To confirm stability of cleistogamy of this mutant in different genetic background, this mutant was crossed with cv. Kasalath(subsp. indica ) , and the segregation of cleitogamy in the F2 population were examined. [1-2. Paraffin sectioning] Flower buds of wild type and spw1-cls were fixed in FAA (formaldehyde : glacial acetic acid : ethanol = 1:1:18) for 24 hours at 4 °C, and then dehydrated in a graded ethanol series. Dehydrated samples in 100 % ethanol were replaced with xylene, and embedded in Paraplast plus (Oxford Labware, St. Louis, MO). Microtome sections at 8 µm thick were stained with Delafiled's hematoxyline and observed with a light microscope. [1-3. Scanning electron microscopy (SEM)] Fresh samples were observed directly through a scanning electron microscopy (VE-8000, Keyence, Osaka, Japan). [1-4. In situ hybridization] Paraffin sections were prepared as described above except that microtome sections at 8 mm thick were applied to slide glasses coated with APS (Matsunami Glasses, Osaka, Japan). Digoxigenin-labeled antisense and sense probes were prepared from the full-length or partial cDNAs of SPW1,OsMADS2 and OsMADS4. In situ hybridiation and immunological detection of the hybridization signals were performed as described in Kouchi, H. and Hata, S. Mol.Gen.Genet.238, 106-119 (1993 ). [1-5. Genetic Mapping] For map-based cloning, F2 population of the homozygous cleistogamous plant and cv.Kasalath (subsp. indica ) was used. Using CAPS markers, the causal gene was mapped on the long arm of chromosome 6. [1-6. Plant transformation] All the genomic DNA fragments or cDNAs (with or without mutations) were amplified by PCR reaction using PfuUltra DNA polymerase (Stratagene) and were verified by sequencing. The DNA fragments were cloned into pZH2B (gifted from Mr. M. Kuroda). For RNAi experiments, 312bp (SPW1), 300bp (OsMADS2) and 58bp (OsMADS4) of gene-specific DNA fragments were cloned in head-to-head orientation into pZH2Bi (gifted from Mr. M. Kuroda) with the rice polyubiquitin gene promoter and the third intron of rice aspartic protease gene. Vectors were introduced into rice scutellum-derived calli by Agrobacterium-mediated transformation under selection by Hygromycin according to a method described in Oikawa et al., Plant Mol. Biol. 55, 687-700 (2004 ). spw1-cls or spw1-1 homozygous calli were selected with allele-specific dCAPS markers (SEQ ID NO: 22 and 23). [1-7. Yeast two-hybrid assay] Coding sequences corresponding to M domain, I domain and K domain of the wild type or mutant (I45T or V45T) of SPW1, OsMADS2 and OsMADS4 were amplified using PfuUltra DNA polymerase (Stratagene), verified by sequencing and subcloned into the two-hybrid vectors, pAS2-1 and pACT2. All the yeast transformation and (β-galactosidase liquid assays were performed as described in Wang, K.L. et al., Nature 428, 945-950 (2004 ), except that the yeast cells were incubated at 20°C. [2: Phenotypes of spw1-cls having cleistogamy] In Fig. 1 , (a) shows appearance of a segregated line exhibiting a wild-type trait (left), and spw1-cls (right), for comparison in cleistogamy therebetween. In Fig. 1 , (b) shows development of a rice grain of spw1-cls in which part of glume is cut. An arrow indicates immature seed, and arrowhead indicates stamens. As is apparent from this, stamens were remained in a glume in spw1-cls. In Fig. 1 , (c) and (d) show structures of internal floral organs of wild type and spw1-cls mutant. A lodicule (indicated by an arrow) of a wild type was not elongated (c), but a lodicule (indicated by an arrow) of spw1-cls was elongated although other floral organs were normal (d). (e) shows comparison in shape of lodicules between wild type (left) and spw1-cls (right). In cleistogamous rice spw1-cls, a lodicule was transformed to be elongated, but other organs such as stamens were not changed. (f) shows transverse sections of wild type (left) and spw1-cls (right) florets. Arrows indicates lodicules. A lodicule of the wild type was enlarged. A lodicule of wild-type rice had many vascular bundles developed, while a lodicule of the spw1-cls mutant had immature vascular bundles and a flat structure. (g) and (h) show scanning electron micrographs of lodicules (lo) and marginal regions of paleae (ma) of the wild type and the spw1-cls mutant. In the wild-type rice, the lodicule had a surface composed of round and swollen cells (g). In spw1-cls, the lodicule had a surface composed of elongated and flat cells observed in the marginal regions of paleae (h). From these results, it is considered that spw1-cls could not flower since an organ to be developed as a lodicule was transformed to a lodicule-glume mosaic organ, and as a result, cells of the organ lost the swelling capability. In Fig. 1 , (i) shows F2 individual expressing wild type (chasmogamy) obtained by crossing indica rice cultivar, Kasalath, with spw1-cls mutant, and F2 individual expressing spw1-cls (cleistogamy) on the right side. From F2 population obtained by crossing of spw1-cls with indica rice cultivar \"Kasalath\", 25 % of cleistogamous individuals were obtained. It was found that the cleistogamy trait was expressed stably independent of genetic backgrounds, i.e. cleistogamy was expressed stably even with the change in genetic backgrounds. In Fig. 1 , (j) shows a post-fertilization flower of the spw1-cls mutant into which 35S promoter:: GFP gene was introduced as a foreign gene. In (j), part of a glume is removed so that the inside of the glume can be observed. It is apparent that green fluorescence caused by high expression of the GFP gene and cleistogamy were observed, and ovary was developed with stamens remaining inside the caryopsis. Thus, it was confirmed that cleistogamy was expressed stably even when spw1-cls mutant was used as a mother plant for genetic recombination. The d7 mutant known as \"Heieidaikoku\" is cleistogamous rice reported so far. In the d7 mutant, the proximal parts of lemma and palea are fused tightly while having normal lodicules, resulting in the non-opened flowers at flowering stage. The d7 mutant, however, is presumed to cause other growth disturbances (e.g. dwarf grain and short grain) due to pleiotropic expressions of causal genes, thus is not suitable for practical use. In view of this, plant shape and panicle shape were compared between spw1-cls mutant having cleistogamy and a wild type ( Fig. 2 ). (a) shows plant shape of a segregant expressing wild-type traits, and (b) shows plant shape of spw1-cls. (c) shows comparison between panicles of wild type (left) and those of spw1-cls (right). (d) shows comparison in rice grain appearance between wild type (left) and spw1-cls mutant (right). It is apparent that the appearance of spw1-cls is similar to that of wild type, except that spw1-cls has cleistogamy. In addition, spw1-cls has seed fertility rate equal to that of wild-type rice, panicles are exserted normally, and there was no significant differences in other traits (plant type, heading date, etc.) between spwl-cls and wild-type rice (Table 1).\n TABLE-tabl0001 [Table 1] Agronomic traits of spw1-cls Heading date Maximal plant height No. of panicles Maximal panicle length No. of grains Fertility Grain weight (days) (cm) (per plant) (cm) (per panicle) (%) (mg) Wild type 116.1±2.5 92.30±2.56 14.6±1.9 22.64±1.26 113.1±14.8 87.55±5.69 20.99±0.65 Spw1-cls 116.8±2.0 90.53±3.53 13.0±2.4 23.70±1.11 127.9±16.7 72.97±11.33 19.58±0.82 As described above, spw1-cls is considered to have influence specifically on the properties of lodicules, and spw1-cls mutant is suitable for practical use. [3: Mechanism of cleistogamy in spw1-cls] In order to reveal mechanism in which cleistogamy is brought by mutation of SPW1 gene, flowers of SPW1-gene-related transformed plants were analyzed ( Fig. 3). Fig. 3 shows (a) the structures of floral organs of a loss-of-function mutant spw1-1 of SPW1 gene, (c) the structures of floral organs of a line in which expression of SPW1 gene was severely suppressed by RNAi method, and (d) the structures of floral organs of a line in which expression of SPW1 gene was mildly suppressed by RNAi method. As shown in Fig. 3 , when the amount of expression of SPW1 gene was reduced by RNAi method, the followings were observed: (a) spw1-1 and spw1-2 (not shown) that are allele of loss-of-function mutation of SPW1, (c) the structures having an elongated lodicule and pistil-like stamens, and (d) the structures having an elongated lodicule as in spw1-cls, but normal stamen appearance. \nIn a case where the expression of SPW1 gene was suppressed severely, an elongated lodicule and pistil-like stamens were observed as in the case of the loss-of-function mutant spw1-1. In a case where the expression of SPW1 gene was suppressed mildly, only an elongated lodicule was observed as in the spw1-cls mutant. However, these lines had transformed pollens and were completely sterile. In Fig. 3 , (e) shows function complementation of a loss-of-function mutant spw1-1 by a mutated SPW1 gene. (e) of Fig. 3 shows that introduction of the mutated SPW1 gene having I45T mutation (gSPW1 I45T ) into spw1-1 results in spw1-cls type flower having an elongated lodicule and normal stamens. As a result of this, it was found that appropriately weakening the function of the SPW1 gene induced morphological change similar to that of spw1-cls, and also it was found difficult to secure pollen fertility and seed fertility merely by reducing the amount of expression of SPW gene. In the present invention, it was verified by map-based cloning that cleistogamy was induced by mutation of SUPERWOMAN1 (SPW1) gene. SPW1 is classified as class-B of MADS-box protein transcriptional activator, and class-B MADS-box proteins are further classified into PISTILLATA (PI) sub-family and APETALLA3 (AP3) sub-family. Further, proteins that belong to the subfamilies form heterodimer, and the heterodimer binds to DNA having a particular sequence called CArG box so that transcription of genes is activated. In \"ABC model\" for formation of floral organs, which model has been proposed for Arabidopsis and other plants, it is known that in dicotyledons, the class-B MADS-box proteins are required for formation of their petals and stamens, and in mocotyledons, the class-B MADS-box proteins are required for formation of their lodicules and stamens. It is considered that SPW 1, which is the sole AP3-type class-B MADS-box protein of rice, forms a heterodimer with PI-type proteins OsMADS2 and OsMADS4, and thus controls the formation of lodicules and stamens. In Fig. 3 , (b) is a view showing the structure of SPW1 gene. Boxes represent exons, thin lines represent introns, ATG indicates translation initiation codon, and TGA indicates translation stop codon. spw1-1and spw1-2 bring loss-of-function mutation (superwoman mutation) due to splicing defect, resulting in transformation of lodicules to glume-like organs and transformation of stamens to pistils. In spw1-cls mutant, the amino acid at position 45 (isoleucine(I)) is substituted with threonine (T). In Fig. 3 , (f) shows comparison between class-B MADS-box proteins ( Arabidopsis (PI, AP3) and rice (SPW1, OsMADS2, OsMADS4)) in amino acid sequences of MADS domains. In (f) of Fig. 3 , reversed characters indicate that amino acids are conserved in three or more proteins out of the five proteins. Shaded areas indicate that amino acids having similar properties are conserved. In wild-type class-B MADS-box proteins, all of the amino acids at position 45 (indicated by an arrow) are hydrophobic amino acids (leucine (L), valine (V), isoleucine (I)), which are highly conserved. However, in spw1-cls, the amino acid at position 45 is substituted with a hydrophilic amino acid, threonine (T), suggesting that the structure of the spw1-cls protein is changed slightly from the structure of the wild-type proteins. Considering that an amino acid near the amino acid at position 45 of the class-B MADS-box protein is mainly a hydrophobic amino acid, the scope of the present invention should encompass that spw1-cls mutant is obtained by substituting the hydrophobic amino acid other than the amino acid at position 45 with a hydrophilic amino acid. Next, it was confirmed that the spw1-cls mutation was caused by amino acid substitution (mutation of the 45th isoleucine to threonine (I45T)) in MADS-box domain (DNA-binding protein domain) ( Fig. 4 ). (a) of Fig. 4 shows function complementation of spw1-cls mutant by a wild-type SPW1 gene. Introduction of wild-type SPW1 gene (gSPW1 wt ) into a spw1-cls mutant resulted in a wild-type flower (in which lodicule (indicated by an arrow) was not elongated). On the other hand, even when a mutant SPW1 gene (gSPW1 I45T ) was introduced into a spw1-cls mutant, spw1-cls flower having a lodicule remained elongated and having normal stamens was obtained (b). From these results, it was found that I45T mutation was the cause of spw1-cls mutation. dCAPS markers (SEQ ID NO: 22 and 23) produced by using this mutation enables cleistogamy-determination on young plants. With the use of the DNA markers, it is possible to successfully select and effectively breed cultivars into which clestogamy is introduced by crossing. Flowering of many Poaceae plants such as rice is considered to occur due to the swelling of lodicules. According to the present invention, similar mutation can be given to a class-B MADS-box gene so as to properly decrease the heterodimer-forming ability, whereby it is possible to produce or select cleistogamous plants in many Poaceae plants. [4: Interaction of class-B MADS-box proteins] As described previously, SPW1 identified as a causal gene of cleistogamy is one of \"class-B MADS-box genes\" known as genes controlling the formation of morphology of petals (lodicules) and stamens in a wide range of higher plants. It is known that class-B MADS-box proteins, products of these genes, are classified into two sub-class proteins, AP3-type proteins and PI-type proteins, and heterodimer formed with a AP3-type subclass protein and a PI-type subclass protein binds to DNA to activate the transcription of gene cluster required for the formation of morphology of organs. SPW1 is the sole AP3-type protein in rice and considered to form heterodimer with PI-type proteins, OsMADS2 and OsMADS4. In view of this, it was analyzed by yeast two-hybrid method using I45T SPW1 proteins (SPW1 I45T ) whether formations of heterodimers to be formed between SPW1 proteins and two PI-type proteins can be effected by spw1-cls mutation. In Fig. 5 , (a) shows the result by yeast two-hybrid method, analyzing protein-to-protein interaction between wild-type or I45T mutant-type SPW1 protein and OsMADS2 protein or OsMADS4 protein. By using intensity of a reporter gene (LacZ gene) expression as an index was measured the interaction between wild-type SPW1 protein (SPW1(wt): white column) or mutant-type SPW1 protein (SPW1(I45T): black column) and wild-type protein or V45T mutant-type protein of OsMADS2, and the interaction between wild-type SPW1 protein (SPW1(wt)) or mutant-type SPW1 protein (SPW1 (I45T)) and wild-type protein or V45T mutant-type protein of OsMADS4. As a result, it was found that the heterodimer-forming ability of the I45T SPW1 protein (SPW1 I45T ) decreased. In (a), the more tightly two proteins are bound to each other, the higher values are. It is clear that wild-type SPW1 wt is bound to wild-type OsMADS2 and OsMADS4 while mutant-type SPW1 I45T is little bound to OsMADS4, and the degree of binding of mutant-type SPW1 I45T with OsMADS2 is in the range from 1/2 to 1/10 the degree of binding of wild-type SPW1 wt with OsMADS2. Further, V45T mutants of OsMADS2 and OsMADS4 were produced to confirm the binding of the V45T mutants of OsMADS2 and OsMADS4 with SPW1 wt . As a result, it was found that binding of OsMADS2V45T with SPW1 wt was about half binding of OsMADS2wt with SPW1 wt , and OsMADS4V45T is little bound to SPW1 wt . In addition, it was found that OsMADS2V45T and OsMADS4V45T were not bound to SPW1 I45T at all. In Fig. 5 , (b) and (c) show expression sites of OsMADS2 gene and OsMADS4 gene using in situ hybridization. As a result of analysis by in situ hybridization, it was found that OsMADS2 expressed in primordia of lodicule and stamen (b), and OsMADS4 expressed only in primordia of stamen (c). These results indicate that in the spw1-cls mutant, in lodicule primordia, the formation of lodicule was inhibited due to weak binding of SPW1 I45T with OsMADS2, and in stamen primordia, while SPW1 I45T is not bound to OsMADS4 at all, a sufficient activity for the formation of stamens is maintained although binding of SPW1 I45T to OsMADS2 is weak. Since OsMADS4 does not express in lodicule primordia, the interaction between SPW and OsMADS2 is considered to be important for the formation of lodicules. This is supported by the fact that a low level of heterodimer formation between SPW1 I45T and OsMADS2 is sufficient to form stamens, but insufficient to establish complete properties of lodicules. To verify this idea, expressions of OsMADS2 gene and OsMADS4 gene were suppressed by RNAi method. In Fig. 5 , (d) and (e) show the structures of floral organs in lines in which the expressions of the OsMADS2 gene and the OsMADS4 gene were suppressed by RNAi method. In the case of a plant in which the expression of the OsMADS2 gene was suppressed, normal stamens and elongated glume-like lodicule were exhibited ((d) in Fig. 5 ) since OsMADS4 mRNA did not exist in the second whorl, but existed in the third whorl. On the contrary, in the case of a plant in which the expression of the OsMADS4 gene was suppressed, phenotypes of stamens and lodicules were not exhibited since OsMADS2 existed in both the second whorl and the third whorl. From the above results, it is considered that in spw1-cls, the heterodimer-forming ability of the SPW1 protein, which is a class-B MADS-box protein, with OsMADS2 or OsMADS decreased, and as a result, class-B activity decreased, so that morphological change was caused only in lodicules, but not in stamens. With the present invention, it is possible to easily produce a plant to which cleistogamy is given. Further, with the present invention, it is possible to easily screen plants having cleistogamy. Specific embodiments or examples implemented in BEST MODE FOR CARRYING OUT THE INVENTION only show technical features of the present invention and are not intended to limit the scope of the invention. Variations can be effected within the spirit of the present invention and the scope of the following claims. INDUSTRIAL APPLICABILITY Cultivars having chasmogamy release their pollens and easily receive pollens of other varieties. Therefore, there have been concerns about crossing and/or gene flow of the cultivars. Self-pollination is impossible without retaining of the function of stamens. The present invention provides cleistogamous cultivars that have been desired, and makes a significant contribution to efficiency of breeding. Conventionally, in order to determine whether a plant has cleistogamy, it was necessary to observe flowering states of individual plants and visually check whether anthers were out of glume at flowering stage. In addition, it was difficult to select individuals having cleistogamy and retaining the function of stamens. The present invention provides a genetic marker by which a plant having cleistogamy can be successfully selected and makes contribution to considerably efficient plant breeding. Thus, the present invention has various applications in the agricultural field, and can also be effectively used in the food industry. In addition, the present invention can be used in a basic research in the biological field.\n \n \n \n \n \n \n \n \n \n \n \n \n \n \n \n","lang":"en","source":"EPO_FULLTEXT","data_format":"ORIGINAL"}},"description_lang":["en"],"has_description":true,"has_docdb":true,"has_inpadoc":true,"has_full_text":true,"biblio_lang":"en"},"jurisdiction":"EP","collections":[],"usersTags":[],"lensId":"197-885-109-383-240","publicationKey":"EP_2019137_A1","displayKey":"EP 2019137 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(I) the step of inducing mutation in a gene that encodes a class-B MADS-box protein, wherein the mutation corresponds to an amino acid substitution at position 45 of the class-B MADS-box protein;
(II) the step of introducing into a plant a gene being capable of inducing mutation in a gene that encodes a class-B MADS-box protein, wherein the mutation corresponds to an amino acid substitution at position 45 of the class-B MADS-box protein; and
(III) the step of suppressing expression of genes encoding class-B MADS-box protein other than a polypeptide having an amino acid sequence represented by SEQ ID NO: 1."],"number":6,"annotation":false,"title":false,"claim":true},{"lines":["A kit for producing a plant to which cleistogamy is given, wherein the kit comprises the polynucleotide according to claim 3."],"number":7,"annotation":false,"title":false,"claim":true},{"lines":["A fragment of the polynucleotide according to claim 3, or an oligonucleotide comprising a complementary sequence of the base sequence of the fragment."],"number":8,"annotation":false,"title":false,"claim":true},{"lines":["A method for screening a plant to which cleistogamy is given, comprising the step of hybridizing the oligonucleotide according to claim 8 with cDNA derived from a target plant."],"number":9,"annotation":false,"title":false,"claim":true},{"lines":["A kit for screening a plant to which cleistogamy is given, wherein the kit comprises the oligonucleotide according to claim 8."],"number":10,"annotation":false,"title":false,"claim":true},{"lines":["An antibody being capable of binding specifically to the polypeptide according to claim 1."],"number":11,"annotation":false,"title":false,"claim":true},{"lines":["A method for screening a plant to which cleistogamy is given, comprising the step of incubating the antibody according to claim 11 with a tissue extract derived from a target plant."],"number":12,"annotation":false,"title":false,"claim":true},{"lines":["A kit for screening a plant to which cleistogamy is given, wherein the kit comprises the antibody according to claim 11."],"number":13,"annotation":false,"title":false,"claim":true}]}},"filters":{"npl":[],"notNpl":[],"applicant":[],"notApplicant":[],"inventor":[],"notInventor":[],"owner":[],"notOwner":[],"tags":[],"dates":[],"types":[],"notTypes":[],"j":[],"notJ":[],"fj":[],"notFj":[],"classIpcr":[],"notClassIpcr":[],"classNat":[],"notClassNat":[],"classCpc":[],"notClassCpc":[],"so":[],"notSo":[],"sat":[]},"sequenceFilters":{"s":"SEQIDNO","d":"ASCENDING","p":0,"n":10,"sp":[],"si":[],"len":[],"t":[],"loc":[]}}