Flour prepared from new wheat

Abstract

The purpose of the present invention is to provide a food product having improved stickiness and meltability in the mouth as compared with food products produced from GA-SX wheat flour.　The problem is solved by altering the allele at a high-molecular-weight glutenin gene Glu-D1 of GA-SX wheat (a wheat in which the enzymatic activity of GBSSI-A1 is not deficient, the enzymatic activities of GBSSI-B1 and GBSSI-D1 are deficient, and the enzymatic activities of any two of SSIIa-A1, SSIIa-B1 and SSIIa-D1 are deficient) to a d-type allele.

Description

[Technical Field]

[0001] The present invention relates to a novel wheat strain that can be used as a raw material for wheat flour compositions used in food production, and more specifically to wheat flour prepared from wheat strains that lack the enzymatic activity of two granule-bound starch synthase I (GBSSI) enzymes and two starch synthase type IIa (SSIIa) enzymes and that contain a d-type allele of the high-molecular-weight glutenin gene Glu-D1. [Background technology]

[0002] Wheat flour is widely used as an ingredient in various processed foods that are consumed daily as meals or snacks. Many of these processed foods are produced through a heating process, but changes in quality occur immediately after the heating process is completed. For example, bread has a very soft and moist texture immediately after baking. However, after a few hours or days, it becomes hard and dry. This phenomenon is generally called food aging, but in this invention it is referred to as deterioration. Since the progression of deterioration is directly related to the taste, reducing this progression is an important issue for the food industry.

[0003] To address this issue, a wheat flour (GA-SX wheat flour) was developed by milling wheat that lacked the enzyme activity of granule-bound starch synthase I (GBSSI)-A1, which is responsible for amylose synthesis, but lacked the enzyme activity of GBSSI-B1 and -D1, and lacked the enzyme activity of two of the enzymes starch synthase IIa (SSIIa)-A1, B1, and D1 involved in the elongation of amylopectin side chains (Patent Document 1: Japanese Patent No. 6226165). This wheat starch has a low amylose content and a structure in which the amylopectin side chains are shortened, resulting in a slow rate of starch retrogradation. As a result, the texture is soft, deterioration after production is slow, and the deliciousness can be maintained for a long time; these effects are particularly excellent in GA-SA wheat flour (wheat flour obtained by milling wheat that is not deficient in GBSSI-A1, but is deficient in the enzyme activities of GBSSI-B1 and D1, is not deficient in SSIIa-A1, and is deficient in SSIIa-B1 and D1).

[0004] However, foods made with GA-SX wheat flour suffer from the drawbacks of chewing and poor melt-in-the-mouth texture, particularly in baked goods such as white bread. These chewiness and poor melt-in-the-mouth texture are thought to be due to the starch composition described above. Wheat varieties with all three GBSSIs functioning contain approximately 25% amylose, whereas mutants lacking one contain 23–24% amylose, a 1–2% reduction. Mutants lacking two GBSSIs produce approximately 20% amylose, while those lacking all three produce waxy starch with no amylose. This difference in amylose content affects the gelatinization and retrogradation properties of starch. It is known that amylose retrogradates faster than amylopectin in gelatinized starch in the short term. Therefore, starch with a high amylose content is more likely to harden during retrogradation, while waxy starch lacking amylose maintains its soft state. For example, in bread, the texture is affected by the properties of starch, so the lower the amylose content, the softer the bread becomes. On the other hand, as the proportion of amylopectin increases, the bread becomes chewy and does not melt in the mouth. GA-SX wheat has an amylose content of 20% or less (Non-Patent Documents 1 and 2), which is thought to be why it has a chewy texture and poor melt-in-the-mouth feel.

[0005] On the other hand, glutenin and gliadin are the major proteins in wheat, and when they coexist and come into contact with water, they form viscoelastic gluten. Glutenin forms large polymers by forming disulfide bonds between its molecules, contributing to the elasticity (strength) of wheat flour dough. Gliadin exists in the form of monomers weakly bound by hydrogen bonds and other mechanisms, contributing to the extensibility of wheat flour dough. Glutenin is broadly divided into high-molecular-weight glutenin and low-molecular-weight glutenin. High-molecular-weight glutenin is encoded by the Glu-A1, B1, and D1 loci on the long arms of wheat chromosomes 1A, 1B, and 1D, while low-molecular-weight glutenin is encoded by the Glu-A3, B3, and D3 loci on the short arms of chromosomes 1A, 1B, and 1D. Each of these six loci has numerous alleles, and the molecular weight and expression level of the encoded subunits vary depending on the allele type, which is known to affect the secondary processing properties of wheat flour. Glu-D1 alleles include a (having a high-molecular-weight glutenin subunit pair "2+12"), c (4+12), d (5+10), and f (2.2+12) types (Non-Patent Document 3), and among these, Glu-D1d is known to be more effective in strengthening dough during breadmaking and increasing bread volume than other Glu-D1 alleles (Non-Patent Documents 4 and 5). Similarly, Glu-B3 alleles such as Glu-B3b, B3h, and B3i have been shown to be more effective in strengthening dough during breadmaking than other Glu-B3 alleles (c, j, ae) (Non-Patent Documents 4 and 6).

[0006] The present inventors have been diligently studying how to obtain wheat flour of even higher quality and have unexpectedly found that the drawbacks of GA-SX wheat flour can be eliminated by converting GA-SX wheat flour to Glu-D1d allele, leading to the completion of the present invention. Furthermore, they have found that the drawbacks of GA-SX wheat flour can be further eliminated by converting GA-SX wheat flour to Glu-B3b or Glu-B3h allele, leading to the completion of the present invention. [Prior art documents] [Patent documents]

[0007] [License 1] Patent No. 6226165 [Non-licensed literature]

[0008] [Non-licensed Document 1] Inokuma et al., J. Agric. Food Chem. 2016, 64, 4, 941-947 [Non-licensed Document 2] Inokuma et al., J. Agric. Food Chem. 2021, 69, 7, 2271-2278 [Non-licensed Document 3] Payne and Lawrence, Cereal Research Communications, (1983) 11, 1, 29-35 [Non-licensed Document 4] Tatsuya Ikeda Journal of the Japanese Society of Food Chemical Engineering (2017) 64,3,171-176 [Non-licensed Document 5] Takata et al., Breeding Science (2000) 50, 303-308 [Non-licensed Document 6] Zhang et al., 2012, BMC Plant Biology, 12:243 [Non-licensed Document 7] Vrinten et al., Mol. Gen. Genet. (1999), 261: 463-471 [Non-licensed Document 8] Saito et al., Mol. Breeding, 2009, 23, 209-217 [Non-licensed Document 9] Shimbata et al., 2005, Theor. Appl. Genet., 111, 6, 1072-1079 [Non-licensed Document 10] Osakabe et al.,Proc. Natl. Acad. Sci. USA (2010) 107(26): 12034-12039 [Non-Patent Document 11] Ishikawa et al. (2005) Tohoku National Agricultural Research Center Research Report, 27-37 [Non-Patent Document 12] RB Gupta and KW Shepherd, Theor Appl. Genet. (1990) 80:65-74 [Non-Patent Document 13] Kojima et al. (2017) Report of the National Agriculture and Food Research Organization, Crop Development Center 1, 1-13 [Non-Patent Document 14] Wang et al., Theor. Appl. Genet.2009, 118:525-539 [Non-Patent Document 15] Fukuda et al. (2010) Breeding Research 12, 87-95 [Non-Patent Document 16] Inagaki, Masanori (2001) Journal of Agricultural, Forestry and Fisheries Technology Research, Vol. 24, No. 12, 44-49 [Non-Patent Document 17] Nakamura et al. Genome, 2002, 45, 1150-1156 Summary of the Invention [Problem to be solved by the invention]

[0009] The present invention aims to provide a food product that has improved chewiness and melt-in-the-mouth properties compared to foods made from GA-SX wheat flour. [Means for solving the problem]

[0010] As a result of extensive research to solve the above problems, the present inventors discovered that the defects of GA-SX wheat flour can be eliminated by converting the high-molecular-weight glutenin gene Glu-D1 of GA-SX wheat (wheat that does not lack the enzyme activity of GBSSI-A1, but lacks the enzyme activities of GBSSI-B1 and GBSSI-D1, and lacks the enzyme activity of any two of SSIIa-A1, SSIIa-B1, and SSIIa-D1) to a d-type allele, leading to the completion of the present invention. Furthermore, the present inventors discovered that the defects of GA-SX wheat flour can be further eliminated by converting the low-molecular-weight glutenin gene Glu-B3 to a b-type or h-type allele, leading to the completion of the present invention.

[0011] That is, the present invention includes the following aspects. [1] Wheat flour (GA-SX / GD1d wheat flour) obtained by milling the harvest of wheat that does not lack the enzyme activity of GBSSI-A1, lacks the enzyme activity of GBSSI-B1 and GBSSI-D1, lacks the enzyme activity of any two of SSIIa-A1, SSIIa-B1 and SSIIa-D1, and has a d-type allele of the high molecular weight glutenin gene Glu-D1. [2] The wheat flour according to [1], wherein the low molecular weight glutenin gene Glu-B3 is a b-type allele (GA-SX / GD1d / GB3b wheat flour). [3] The wheat flour according to [1], wherein the low molecular weight glutenin gene Glu-B3 is an h-type allele (GA-SX / GD1d / GB3h wheat flour). [4] A flour composition comprising the wheat flour described in [1] to [3]. [5] A method for producing food using the wheat flour described in [1] to [3]. [6] The method according to [5], wherein the food is a bakery food. [Effects of the Invention]

[0012] Foods made using GA-SX wheat flour show reduced hardening (deterioration) during storage for several days after production. However, these products can have undesirable textures, such as poor melt-in-the-mouth texture and a chewy texture. By using wheat flour prepared from the wheat of the present invention, it is possible to obtain products with improved texture while maintaining the same level of deterioration reduction as conventional GA-SX wheat flour. DETAILED DESCRIPTION OF THE INVENTION

[0013] The wheat used in the present invention is wheat that does not lack the enzymatic activity of GBSSI-A1, but lacks the enzymatic activities of GBSSI-B1 and GBSSI-D1, and lacks the enzymatic activity of any two of SSIIa-A1, SSIIa-B1, and SSIIa-D1, and has a d-type allele of the high-molecular-weight glutenin gene Glu-D1 (GA-SX / GD1d wheat).Preferably, it also has a b-type allele of the low-molecular-weight glutenin gene Glu-B3 (GA-SX / GD1d / GB3b wheat) or an h-type allele of the low-molecular-weight glutenin gene Glu-B3 (GA-SX / GD1d / GB3h wheat).

[0014] Common wheat is an allohexaploid, and its chromosomes consist of three homoeologous genomes, A, B, and D, numbered 1 to 7 (1A-7A, 1B-7B, and 1D-7D). "GBSSI" is a granule-bound starch synthase involved in the synthesis of amylose contained in wheat endosperm starch, also known as Waxy (Wx). GBSSI(Wx)-A1, GBSSI(Wx)-B1, and GBSSI(Wx)-D1 are encoded by genes located on chromosomes 7A, 4A, and 7D, respectively. Mutants lacking the enzyme function are known for each, and the amylose content varies depending on the combination of mutants. "SSIIa" is an enzyme involved in the elongation of the side chains (branch chains) of amylopectin in wheat endosperm starch. Like GBSSI, SSIIa-A1, B1, and D1 are functional in normal wheat, and defective mutants of each are also known. SSIIa-A1, SSIIa-B1, and SSIIa-D1 are encoded by genes located on chromosomes 7A, 7B, and 7D, respectively. Deficiency of one of the three enzymes results in slight shortening of the amylopectin side chains. Deficiency of two enzymes results in greater shortening, and deficiency of all three enzymes results in the greatest shortening of the side chains, with the secondary effect of producing high-amylose wheat with an amylose content of over 30%.

[0015] "Lack of enzymatic activity" means that a protein with normal enzymatic activity is not functioning in the wheat plant, preferably that a protein with normal enzymatic activity is not expressed. Specific examples include mutations in gene sequence (mutations such as substitution, deletion, insertion, inversion, or translocation of one or more bases, including deletion of the entire gene region), defects in mRNA transcription, defects in protein translation, and inhibition of enzymatic activity in the wheat plant. Any of these may occur as long as the enzymatic activity is reduced or eliminated to less than 10%, preferably less than 5%, and more preferably less than 1% of the wild-type enzymatic activity.

[0016] In this specification, "wheat that does not lack the enzyme activity of GBSSI-A1, lacks the enzyme activity of GBSSI-B1 and GBSSI-D1, and lacks the enzyme activity of any two of SSIIa-A1, SSIIa-B1, and SSIIa-D1" is referred to as "GA-SX wheat." GA-SX wheat is a combination of two types of SSIIa that lack enzyme activity, and includes the following wheats: GA-SA wheat: wheat that is not deficient in the enzyme activity of GBSSI-A1, but is deficient in the enzyme activities of GBSSI-B1 and GBSSI-D1, and is not deficient in the enzyme activity of SSIIa-A1, but is deficient in the enzyme activities of SSIIa-B1 and SSIIa-D1; GA-SB wheat: wheat that is not deficient in the enzyme activity of GBSSI-A1, but is deficient in the enzyme activities of GBSSI-B1 and GBSSI-D1, and is not deficient in the enzyme activity of SSIIa-B1, but is deficient in the enzyme activities of SSIIa-A1 and SSIIa-D1; GA-SC wheat: Wheat that is not deficient in GBSSI-A1 enzyme activity, but is deficient in GBSSI-B1 and GBSSI-D1 enzyme activity, and is not deficient in SSIIa-D1 enzyme activity, but is deficient in SSIIa-A1 and SSIIa-B1 enzyme activity. GA-SX wheat is wheat that can be produced according to known methods, for example, the method described in JP 2013-188206 A.

[0017] Examples of wild-type genes that do not lack enzyme activity include the sequences (genomic DNA and protein) of GBSSI-A1, B1, and D1, and SSIIa-A1, B1, and D1, which are registered in GenBank under the following accession numbers. These sequences are shown in the sequence listing in Table 1 below.

[0018] [Table 1]

[0019] These sequences are examples of wild-type sequences. Naturally occurring wheat (including improved wheat varieties) may contain enzyme proteins with equivalent activity but with partial differences in their nucleotide or amino acid sequences. In the present invention, the terms "GBSSI-A1 gene" and "GBSSI-A1 protein" encompass not only those with a completely identical nucleotide or amino acid sequence to the sequence listing, but also those with naturally occurring mutations that do not impair enzymatic activity. The same applies to other enzymes. Such naturally occurring mutant sequences typically share 90% or more, e.g., 95% or more, or 98% or more identity with the respective nucleotide or amino acid sequences listed in the sequence listing. In this specification, these GBSSI-A1, B1, D1, and SSIIa-A1, B1, and D1 genotypes are referred to as GBSSI-A1a, B1a, D1a, and SSIIa-A1a, B1a, and D1 alleles. As an example of a GBSSI-A1 mutant lacking enzyme activity compared to these wild-type alleles, a genetic mutation has been identified in which 23 base pairs are deleted at the junction between the first exon and the following intron in the wild-type GBSSI-A1 (Wx-A1) gene sequence, and a four-base difference sequence has been inserted, resulting in loss of expression of the encoded GBSSI-A1 protein (Non-Patent Document 7). In this specification, this mutant is referred to as the GBSSI-A1b allele. A mutant of GBSSI-B1 is known in which the entire gene region from the initiation codon to the termination codon of wild-type GBSSI-B1 (Wx-B1) is deleted (Non-Patent Document 8), and this mutant is referred to herein as the GBSSI-B1b allele. A mutant of GBSSI-D1 is known in which expression of GBSSI-D1 is lost due to a genetic mutation in which 588 bases around the stop codon in the wild-type GBSSI-D1 (Wx-D1) gene sequence are deleted and a different sequence of 12 bases is inserted (Non-Patent Document 7). In this specification, this mutant is referred to as the GBSSI-D1b allele. A mutant of SSIIa-A1 is known in which a 289-base region including the initiation codon is deleted from the wild-type SSIIa-A1 gene sequence and an 8-base sequence is inserted, resulting in loss of SSIIa-A1 expression (Non-Patent Document 9). In this specification, this mutant is referred to as the SSIIa-A1b allele. A known mutant of SSIIa-B1 has a 175-base insertion in exon 8 of the wild-type SSIIa-B1 gene sequence, resulting in a stop codon that disrupts normal expression of the SSIIa-B enzyme protein (Non-Patent Document 9). In this specification, this mutant is referred to as the SSIIa-B1b allele. A mutant of SSIIa-D1 is known in which the expression of normal SSIIa-D enzyme protein is lost due to a genetic mutation in which 63 bases around the junction region of exon 5 and the following intron in the wild-type SSIIa-D1 gene sequence are deleted (Non-Patent Document 9). In the present invention, this mutant is referred to as the SSIIa-D1b allele.

[0020] "Glutenin" is the main protein in wheat, and when it comes into contact with water in the presence of gliadin, it forms viscoelastic gluten. Glutenin forms large polymers by forming disulfide bonds between its molecules, and is involved in the elasticity (strength) of wheat flour dough. Gliadin exists in the form of a monomer weakly bonded by hydrogen bonds, etc., and is involved in the extensibility of wheat flour dough. Glutenin is broadly divided into high molecular weight glutenin and low molecular weight glutenin.

[0021] "High molecular weight glutenin" is encoded by the Glu-A1, B1, and D1 loci located on the long arms of wheat chromosomes 1A, 1B, and 1D, while "low molecular weight glutenin" is encoded by the Glu-A3, B3, and D3 loci located on the short arms of chromosomes 1A, 1B, and 1D. Numerous alleles (alleles) are known at each of these six loci, and the molecular weight and expression level of the encoded subunits vary depending on the type of allele, which is known to affect the secondary processability of wheat flour. The wheat used in the present invention is GA-SX wheat, in which the high-molecular-weight glutenin gene Glu-D1 is a d-type allele (GA-SX / GD1d). Preferably, the low-molecular-weight glutenin gene Glu-B3 is also a b-type allele (GA-SX / GD1d / GB3b wheat), or the low-molecular-weight glutenin gene Glu-B3 is also an h-type allele (GA-SX / GD1d / GB3h wheat). In another embodiment, the wheat used is GA-SX wheat, in which the high-molecular-weight glutenin gene Glu-D1 is a d-type allele, and preferably the low-molecular-weight glutenin gene Glu-B3 is also an i-type allele (GA-SX / GD1d / GB3i wheat).

[0022] The wheat used in the present invention, which is a GA-SX wheat, can be produced by crossbreeding known wheat varieties lacking any combination of six enzymes, including those lacking GBSSI and SSIIa enzyme activity. Mutagenesis may be performed using radiation (e.g., gamma rays, beta rays, X-rays, neutrons), chemical treatment (e.g., ethyl methanesulfonate), or other mutagenic treatments, followed by selection of the desired enzyme-deficient mutants for use in crossbreeding. Various methods for producing monocotyledonous transformants are known, including genetic engineering techniques for disrupting the function of a target gene. For example, methods for inhibiting the expression of a target gene using RNAi or antisense technology are available. Furthermore, gene disruption methods for disrupting only the target gene in plants are also known (Non-Patent Document 10). Therefore, genetic engineering techniques can also be used to produce GA-SX wheat. In combination with the deficiency of two GBSSI and two SSIIa enzyme activities, the high-molecular-weight glutenin gene Glu-D1 allele type is the d type. Preferably, the allele type of the low-molecular-weight glutenin gene Glu-B3 is type b or type h. This may be achieved by first producing wheat lacking the enzymatic activities of two GBSSIs and two SSIIas, and then introducing the desired alleles of the high-molecular-weight glutenin gene Glu-D1, and preferably the glutenin gene Glu-B3, into the wheat. Alternatively, this may be achieved by selecting a wheat variety that has the desired alleles of the high-molecular-weight glutenin gene Glu-D1, and preferably the glutenin gene Glu-B3, during the process of producing wheat lacking the enzymatic activities of two GBSSIs and two SSIIas.

[0023] The Glu-D1d allele is based on the classification and nomenclature proposed in Non-Patent Document 3. The presence or absence of the Glu-D1d allele can be identified by detecting a unique nucleotide sequence in the Dx5 gene (GenBank accession number X12928: SEQ ID NO: 13), which encodes subunit 5 of the Glu-D1d allele. This identification can be achieved by PCR using the Dx_F, Dx5_F, and Dx_R primers (Dx_F: SEQ ID NO: 20, Dx5_F: SEQ ID NO: 21, Dx_R: SEQ ID NO: 22) described in Non-Patent Document 11. Alternatively, the region containing this unique mutation can be amplified using appropriately designed primers, and the gene sequence can be analyzed to confirm that it matches this sequence. Alternatively, a fraction containing high-molecular-weight glutenin protein can be extracted from wheat, separated by SDS-PAGE, and compared with known wheat samples to identify a band specific to the Glu-D1d allele (Non-Patent Document 3).

[0024] The Glu-B3b, B3h, and B3i alleles are based on the classification and nomenclature proposed in Non-Patent Document 12. These alleles can be identified by the following method. Whether or not a mutant has the Glu-B3b allele can be identified by examining the gene sequence described in Non-Patent Document 14 (GenBank Accession No. EU369719: SEQ ID NO: 14) for the presence of a sequence specific to Glu-B3b. This can be achieved by performing PCR using the SB2F and SB2R primers (SB2F: SEQ ID NO: 23, SB2R: SEQ ID NO: 24) described in Non-Patent Document 14 and examining whether an amplified fragment of the desired length is obtained. Alternatively, PCR can be performed using the LB1F and LB1R or LB4F and LB4R primers (LB1F: SEQ ID NO: 25, LB1R: SEQ ID NO: 26, LB4F: SEQ ID NO: 27, LB4R: SEQ ID NO: 28) described in Non-Patent Document 14, and analyzing the gene sequence of the resulting amplified fragment to determine whether it matches the Glu-B3b gene sequence (GenBank Accession No. EU369700: SEQ ID NO: 15 or EU369719: SEQ ID NO: 14). Alternatively, a fraction containing low-molecular-weight glutenin proteins can be extracted from wheat, separated by SDS-PAGE, and compared with known wheat samples to identify a band specific to the Glu-D1d allele (Non-Patent Document 12). Wheat varieties known to have the Glu-B3b allele include "Takunekomugi" and "Nanbukomugi" (Non-Patent Document 13).

[0025] The presence of the Glu-B3h allele can be identified by examining the gene sequence (GenBank Accession No. EU369717: SEQ ID NO: 16) described in Non-Patent Document 14 for the presence of a sequence specific to Glu-B3h. This can be achieved by performing PCR using the SB8F and SB8R primers (SB8F: SEQ ID NO: 29, SB8R: SEQ ID NO: 30) described in Non-Patent Document 14 and examining whether an amplified fragment of the desired length is obtained. Alternatively, PCR can be performed using the LB3F and LB3R primers (LB3F: SEQ ID NO: 31, LB3R: SEQ ID NO: 32) described in Non-Patent Document 14, analyzing the gene sequence of the amplified fragment, and examining whether it matches the Glu-B3h gene sequence (GenBank Accession No. 369717: SEQ ID NO: 18). Alternatively, similar to the Glu-B3b allele, SDS-PAGE can be used to confirm the presence of the Glu-B3h allele. Known wheat varieties that contain the Glu-B3h allele include "Horoshiri Komugi" and "Haruyutaka" (Non-Patent Document 13).

[0026] Whether or not a gene has a Glu-B3i allele can be identified by examining the gene sequence (GenBank accession number EU369720: SEQ ID NO: 19) described in Non-Patent Document 14 for the presence of a sequence specific to Glu-B3i. This can be achieved by performing PCR using the SB9F and SB9R primers (SB9F: SEQ ID NO: 33, SB9R: SEQ ID NO: 34) described in Non-Patent Document 14 and examining whether an amplified fragment of the desired length is obtained. Alternatively, PCR can be performed using the LB3F and LB3R or LB4F and LB4R primers (LB3F: SEQ ID NO: 35, LB3R: SEQ ID NO: 36, LB4F: SEQ ID NO: 37, LB4R: SEQ ID NO: 38) described in Non-Patent Document 14, and analyzing the gene sequence of the resulting amplified fragment to determine whether it matches the Glu-B3i gene sequence (GenBank accession number EU369718: SEQ ID NO: 18, EU369720: SEQ ID NO: 17, or EU369714: SEQ ID NO: 19). Alternatively, it can be confirmed by SDS-PAGE, just like the Glu-B3b allele. Known wheat varieties with the Glu-B3i allele include Norin 61, Minami no Kaori, and Iwainodaichi (Non-Patent Document 15).

[0027] The gene sequences and primer sequences that can be used to identify each allele are shown in the sequence listings in Tables 2 and 3.

[0028] [Table 2]

[0029] [Table 3]

[0030] The wheat flour of the present invention is wheat flour (GA-SX / GD1d wheat flour) obtained by milling the harvested product of the aforementioned GA-SX / GD1d wheat. Preferably, wheat flour (GA-SX / GD1d / GB3b wheat flour) is obtained by milling the harvested product of GA-SX / GD1d / GB3b wheat, and more preferably wheat flour (GA-SX / GD1d / GB3h wheat flour) is obtained by milling the harvested product of GA-SX / GD1d / GB3h wheat. The milling method is not particularly limited, and any conventional milling method used to produce wheat flour from the harvested product (caryopsis or seeds) of conventional wheat varieties can be used. The form of the wheat flour is not particularly limited; for example, it may be wheat flour from which components such as "bran" have been removed through a conventional milling process, or it may be whole wheat flour without fractionation.

[0031] The wheat flour of the present invention can also be provided as a flour composition in which it is mixed with other wheat flours or flours other than wheat flour. Examples of other wheat flours include wheat flours such as strong flour, medium flour, and soft flour, as well as wheat-derived flours not classified into these categories. Examples of flours other than wheat flour include, but are not limited to, flours derived from grains such as rice, rye, barley, corn, buckwheat, soybean, barnyard millet, foxtail millet, and amaranth. In the flour composition of the present invention, the content of the wheat flour of the present invention is preferably 5 to 100% by mass, more preferably 25 to 100% by mass, even more preferably 50 to 100% by mass, and most preferably 100% by mass, based on the total amount of the flour composition.

[0032] The wheat flour and flour compositions of the present invention have a d-type allele for the high-molecular-weight glutenin gene Glu-D1 (GA-SX / GD1d), which reduces deterioration to the same extent as conventional GA-SX wheat flour, while improving the chewiness and melt-in-the-mouth problems that have been a problem in foods, particularly bakery foods, produced from conventional GA-SX wheat flour and flour compositions containing GA-SX wheat flour, thereby producing products with improved texture.Furthermore, the low-molecular-weight glutenin gene Glu-B3 has a b-type or h-type allele, which allows for products with even improved texture.

[0033] The flour and flour composition of the present invention can be used to produce various foods that use flour or flour-containing flour compositions. Examples of such foods include bakery foods such as breads, cakes, baked goods, and pizza; noodles such as udon and Chinese noodles; fried foods such as tempura and fries; baked foods such as gyoza, spring rolls, and shumai (wheat-flour-containing skins); and fish paste foods such as kamaboko and chikuwa. Preferably, the foods are bakery foods. These foods can be produced by commonly employed production methods, except for using the flour and flour composition of the present invention.

[0034] Examples of bakery foods include breads such as white bread, French bread, rolls, and sweet rolls; fried breads such as yeast donuts; steamed breads; pizzas such as pizza pies; cakes such as sponge cakes; and baked goods such as cookies and biscuits. The bakery foods of the present invention can be produced, for example, by kneading the wheat flour or cereal flour composition of the present invention with various auxiliary ingredients commonly used in the production of bakery foods, such as chemical leavening agents such as baking soda, yeast, yeast food, salt, sugars, oils and fats, eggs, dairy products, and water, to form a dough, which is then left to rise by fermentation or baked or deep-fried as is. Additives such as vitamins and minerals can be added as needed. Conventional production methods may be used to produce the bakery foods of the present invention. [Example]

[0035] The present invention will be described in more detail below based on examples, but the present invention is not limited to these examples.

[0036] <1. Wheat Development> To develop the wheat of the present invention, wheat having the alleles shown in the table below was used. TIFF0007807770000004.tif91158

[0037] (1) Development of SK wheat SK wheat was produced by introducing the GBSSI-D1b, SSIIa-B1b, and D1b alleles from wheat (ii) into wheat (i) through successive backcrossing. Backcrossing is a breeding method in which one parent variety (the recurrent parent) is crossed again with the progeny obtained from a cross between two varieties. Repeated crosses with the recurrent parent are called successive backcrossings. This successive backcrossing is often used to efficiently introduce genes (e.g., disease resistance) that a particular superior variety lacks while maintaining many of its characteristics. The parent used in the recurrent cross is called the recurrent parent, and the parent that donates the desired gene is called the nociceptive parent. To produce SK wheat, successive backcrossing was performed using wheat (i) as the recurrent parent and wheat (ii) as the nociceptive parent. The individuals obtained in each successive backcross cycle were identified for alleles at the GBSSI-D1, SSIIa-B1, and D1 loci, and individuals heterozygous for the a and b alleles at all three loci were selected for the next backcross. After backcrossing, the selected individuals were selfed to obtain progeny, and individuals homozygous for each of the eight loci listed in Table 1 were selected.

[0038] (2) Production of HK wheat Successive backcrossings were performed using wheat (iii) as the recurrent parent and wheat (iv) as the nodal parent, and wheat (iii)-A, in which the GBSSI-A1b and GBSSI-D1b alleles were introduced into wheat (iii), was selected. Similarly, successive backcrossings were performed using wheat (v) as the nodal parent with wheat (iii), and wheat (iii)-B, in which the SSIIa-A1b, B1b, and D1b alleles were introduced into wheat (iii), was selected. Wheat (iii)-A and (iii)-B were crossed, and genetic fixation was performed using the haploid breeding method (Non-Patent Document 16). From the resulting population (DH population), HK wheat, which possesses homozygous alleles for each of the eight loci listed in Table 1, was selected.

[0039] (3) Development of N1-1 wheat Wheat (vii) was crossed with wheat (viii), and the resulting F1 generation individuals were backcrossed with wheat (vi) as the recurrent parent. From this progeny, several individuals were selected that were homozygous for GBSSI-A1a, B1b, and SSIIa-A1a alleles and heterozygous for GBSSI-D1, SSIIa-B1, and D1. A next-generation population was obtained from these individuals, and then genetic fixation was performed using haploid breeding to obtain a DH population. From this DH population, N1-1 wheat was obtained, which is homozygous for each allele of the eight loci listed in Table 1.

[0040] In the above crossing and selection process, the alleles at each locus were determined according to the following method. Allele determination for the GBSSI-A1 and D1 loci was performed according to the method in Non-Patent Document 17. Allele determination for the GBSSI-B1 locus was performed according to the method in Non-Patent Document 8. Allele determination of the SSIIa-A1, B1, and D1 loci was performed according to the method described in Non-Patent Document 9. The Glu-D1d allele was determined according to the method described in Non-Patent Document 11. The Glu-B3b and B3h alleles were determined according to the method described in Non-Patent Document 14. The Glu-B3i allele was identified by sequencing the gene sequence of the fragment amplified using the LB3F and LB3R primers in Non-Patent Document 14 under the PCR conditions described in the same document, and examining the homology with the sequence of the Glu-B3i allele (GenBank accession number EU369718, sequence number 18). DFBI wheat (MK 5-5 NIL wheat described in Non-Patent Document 2), which is a conventional GA-SX wheat, was used as a comparison example. The genotypes of each wheat are as shown in Table 1 below.

[0041] Table 1. Genotypes (alleles) of each wheat variety TIFF0007807770000005.tif65156

[0042] <2. Preparation of flour> The harvested wheat raw material was ground using a Buhler test mill. Water was added to the wheat raw material to bring the moisture content to 14%, and the wheat raw material was left to stand overnight. The wheat raw material was then placed in the test mill and ground to produce flour with a milling yield of 60%.

[0043] <3. Bread making test> Two loaves of bread were made using the above wheat flour using a 70% standard dough method. 70 parts by weight of 100 parts by weight of wheat flour were added to 2.3 parts by weight of yeast, 0.1 parts by weight of yeast food, and 40 parts by weight of water. The dough was mixed at low speed for 2 minutes and medium speed for 2 minutes using a commercially available bread mixer (SK Mixer Co., Ltd., product name: SK200) to obtain a dough. This dough was fermented for 4 hours at a temperature of 27°C and a relative humidity of 75%. To this dough, 30 parts by weight of wheat flour, 5 parts by weight of sugar, 2 parts by weight of salt, 5 parts by weight of shortening, 2 parts by weight of skim milk powder, and an appropriate amount of water were added. The amount of water added was determined in advance to ensure that the dough properties (elasticity and stickiness) were equivalent for each test group. After mixing at low speed for 2 minutes, medium speed for 3 minutes, and high speed for 1 minute, the dough was scraped off and further mixed at low speed for 1 minute, medium speed for 3 minutes, and high speed for 7 minutes. The dough was fermented for 20 minutes, then divided into 460-gram portions, shaped, and placed in baking molds. The dough was then proofed at 38°C and 85% humidity, and baked (reel oven, 210°C, 25 minutes) when the fermented dough reached 1 cm above the mold. After baking, the loaves were removed from the molds and allowed to cool at room temperature for 1 hour. Then, they were sealed in polyethylene bags, and one loaf was left at room temperature until the next day, while the other was left until the third day. Before tasting, each slice was 12.5 mm thick, and 10 panelists evaluated it according to the criteria shown in the evaluation criteria table (Table 2). The evaluation the next day was based on the next day's evaluation of DFBI, with the score of 3.0. For the evaluation on the third day, DFBI slices that had been frozen the day after baking and thawed at room temperature for 4 hours on the day of evaluation were used as the standard (3.0).

[0044] Table 2 Evaluation criteria table TIFF0007807770000006.tif89153

[0045] <Production example> Bread was made using the wheat flour prepared from SK, HK, N1-1, and DFBI, and a sensory evaluation was performed on the day after baking and on the third day. The results are shown in Table 3.

[0046] Table 3. Sensory evaluation results TIFF0007807770000007.tif62153

[0047] In terms of softness, SK, HK, and N1-1 were equivalent to DFBI. On the other hand, in terms of crunchiness and melt-in-the-mouth feel, all three samples (SK, HK, and N1-1) scored higher than DFBI on both the day after baking and the third day after baking. Among the three samples, HK was rated higher than N1-1 on both the day after baking and the third day after baking, with SK receiving an even higher rating. From the above, the presence of Glu-D1d in GA-SX wheat significantly improved the crunchiness and melt-in-the-mouth feel of bread. Furthermore, the effect was greatest when it was also present with Glu-B3h, followed by Glu-B3b and B3i, in that order.

Claims

1. GA-SX / GD1d wheat flour is obtained by milling a harvest of wheat that does not lack the enzyme activity of GBSSI-A1, but lacks the enzyme activities of GBSSI-B1 and GBSSI-D1, and lacks the enzyme activity of any two of SSIIa-A1, SSIIa-B1, and SSIIa-D1, and that has a d-type allele of the high-molecular-weight glutenin gene Glu-D1.

2. The wheat flour according to claim 1, further comprising a b-type allele of the low-molecular-weight glutenin gene Glu-B3 (GA-SX / GD1d / GB3b wheat flour).

3. The wheat flour according to claim 1, further comprising an h-type allele of the low-molecular-weight glutenin gene Glu-B3 (GA-SX / GD1d / GB3h wheat flour).

4. A flour composition comprising the wheat flour according to any one of claims 1 to 3.

5. A grain flour composition for bakery foods, comprising the wheat flour according to any one of claims 1 to 3.

6. A method for producing food, using the wheat flour according to any one of claims 1 to 3.

7. The method of claim 6, wherein the food product is a bakery food product.

8. A grain flour composition for bakery foods comprising the wheat flour described in any one of claims 1 to 3, wherein the grain flour composition for bakery foods is used to obtain foods having an improved texture compared to bakery foods produced using wheat flour (GA-SX wheat flour) obtained by milling a harvest of wheat that does not lack the enzyme activity of GBSSI-A1, lacks the enzyme activity of GBSSI-B1 and GBSSI-D1, and lacks the enzyme activity of any two of SSIIa-A1, SSIIa-B1 and SSIIa-D1.

9. A method for producing bakery foods having an improved texture compared to bakery foods produced using wheat flour (GA-SX wheat flour) obtained by milling a harvest of wheat that does not lack the enzyme activity of GBSSI-A1, lacks the enzyme activity of GBSSI-B1 and GBSSI-D1, and lacks the enzyme activity of any two of SSIIa-A1, SSIIa-B1, and SSIIa-D1, comprising using wheat flour described in any one of claims 1 to 3.

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