Rice gene OsGLW3 and mutant and application thereof

Abstract

The invention discloses a rice gene OsGLW3 as well as a mutant and application thereof. The invention clones a novel dominant gene OsGLW3 for controlling the grain type, the plant type and the panicle type of rice, and the nucleotide sequence and the coded protein sequence of the dominant gene OsGLW3 are respectively shown as SEQ ID NO: 1 and SEQ ID NO: 3. In a mutant RE10498, a coding region 3119T of the gene is mutated into A, so that translation protein is terminated in advance, and slender grains are generated. The OsGLW3 encodes an unknown expression protein, the function of the gene is confirmed through complementary, knockout and overexpression experiments, and the OsGLW3RE is of a function obtaining type. The OsGLW3 gene has important significance on molecular mechanism research of rice grain shape formation, an excellent gene resource is provided for rice variety improvement and cross breeding, and the breeding process is greatly accelerated.

Description

Technical Field

[0001] The invention relates to the fields of genetic engineering and plant genetic breeding, and in particular to a rice gene OsGLW3 and a mutant thereof and application thereof. Background Art

[0002] Rice (Oryza sativa L) is a basic food source for human survival. It provides energy and nutrition for billions of people around the world and is a key factor in ensuring food security and maintaining social stability.

[0003] Grain shape is closely related to the thousand-grain weight of rice. Generally speaking, longer and wider grains tend to have larger volume and weight. Under the condition that the number of panicles and grains per panicle per unit area is relatively stable, appropriately increasing the parameters related to grain shape can significantly improve the yield potential of rice. For example, under ideal cultivation conditions, the thousand-grain weight of some large-grain rice varieties can be 20%-30% higher than that of ordinary varieties, thereby achieving an increase in yield. In the past few decades of rice breeding, breeders have continuously explored ways to increase yield by improving grain shape. From traditional hybrid breeding to modern molecular breeding technology, grain shape is used as one of the important selection indicators.

[0004] In addition, rice grain shape has an important impact on the appearance quality, processing quality and edible quality of rice. In terms of appearance quality, slender grains with low chalkiness are more popular in the market, and consumers often regard them as important characteristics of high-quality rice. Rice with uniform grain shape is easier to shell and grind during processing, which can reduce the broken rice rate and increase the rice yield, thereby improving the processing quality. As for edible quality, grain shape affects the steaming and cooking characteristics and taste of rice. Long-grain rice usually has good elasticity and ductility after steaming, and the rice tastes softer and more delicious, which is suitable for making a variety of foods such as fried rice; while short-grain rice is relatively sticky and suitable for making specific foods such as sushi. Different regions and consumer groups have different demands for rice quality. Therefore, it is particularly important to cultivate rice varieties with grain shapes that meet different market demands.

[0005] Numerous studies have shown that grain shape is regulated by multiple genes and is a complex quantitative trait. At present, many rice grain shape genes have been cloned. For example, GS3 is the first cloned and well-studied major gene controlling rice grain length. Its different alleles can significantly affect the change of grain length (Fan et al., 2006; Mao et al., 2010; Li Yang et al., 2016); GW2, GW5, etc. have an important regulatory effect on grain width (Liu et al., 2017; Song et al., 2007). They determine the width development of grains by affecting processes such as cell division and cell expansion; GL7, GW8, etc. can change rice quality while regulating grain shape (Wang et al., 2012; Yuexing et al., 2015).

[0006] The cloned grain type genes are all recessive mutations and semi-dominant mutations. So far, there has been no report on the application of dominant slender grain superior mutation genes in hybrid rice breeding. Therefore, creating dominant slender grain germplasm and cloning related genes, while analyzing their genetic mechanisms, can provide specific germplasm resources for hybrid rice quality improvement. Summary of the invention

[0007] The purpose of the present invention is to provide rice gene OsGLW3 and application thereof.

[0008] Another object of the present invention is to provide a mutant of the rice gene OsGLW3 RE and its applications.

[0009] In order to achieve the object of the present invention, in a first aspect, the present invention provides a rice gene OsGLW3, which is a gene encoding the following protein (A) or (B):

[0010] (A) a protein consisting of the amino acid sequence shown in SEQ ID NO: 3; or

[0011] (B) A protein derived from (A) with equivalent functions, wherein one or more amino acids are substituted, deleted or added to the sequence shown in SEQ ID NO: 3.

[0012] The nucleotide sequence of the gene OsGLW3 is:

[0013] I) the nucleotide sequence shown in SEQ ID NO: 1;

[0014] II) a nucleotide sequence in which one or more nucleotides of the nucleotide sequence shown in SEQ ID NO: 1 are replaced, deleted and / or added and the nucleotide sequence expresses a protein with the same function;

[0015] III) a nucleotide sequence that hybridizes with the sequence shown in SEQ ID NO: 1 under stringent conditions and expresses a protein with the same function, wherein the stringent conditions are hybridization at 65°C in a 0.1×SSPE solution containing 0.1% SDS or a 0.1×SSC solution containing 0.1% SDS, and the membrane is washed with the solution; or

[0016] IV) is a nucleotide sequence that has more than 90% homology with the nucleotide sequence of I), II) or III) and expresses a protein with the same function.

[0017] Its cDNA sequence is:

[0018] i) the nucleotide sequence shown in SEQ ID NO: 2;

[0019] ii) a nucleotide sequence in which one or more nucleotides are substituted, deleted and / or added to the nucleotide sequence shown in SEQ ID NO: 2 and the nucleotide sequence expresses a protein with the same function;

[0020] iii) a nucleotide sequence that hybridizes with the sequence shown in SEQ ID NO: 2 under stringent conditions and expresses a protein with the same function, wherein the stringent conditions are hybridization in a 0.1×SSPE solution containing 0.1% SDS or a 0.1×SSC solution containing 0.1% SDS at 65°C, and the membrane is washed with the solution; or

[0021] iv) a nucleotide sequence that has more than 90% homology with the nucleotide sequence of i), ii) or iii) and expresses a protein with the same function.

[0022] In a second aspect, the present invention provides a biological material containing the gene OsGLW3, wherein the biological material includes but is not limited to recombinant DNA, an expression cassette, a transposon, a plasmid vector, a viral vector, an engineered bacterium or a non-renewable plant part.

[0023] In a third aspect, the present invention provides any of the following uses of the gene OsGLW3 or a biological material containing the gene:

[0024] (1) Regulate plant grain shape, plant shape and ear shape;

[0025] (2) Used for plant variety improvement;

[0026] (3) Used for preparing transgenic plants.

[0027] In the present invention, the rice grain type includes grain length, grain width and grain length-to-width ratio; the plant type includes plant height; and the ear type includes ear length.

[0028] In the present invention, the plant is a grass plant, preferably rice.

[0029] In a fourth aspect, the present invention provides a rice OsGLW3 gene mutant OsGLW3 RE , which is a gene encoding the following protein (a) or (b):

[0030] (a) a protein consisting of the amino acid sequence shown in SEQ ID NO: 4; or

[0031] (b) A protein derived from (a) with equivalent functions, wherein one or more amino acids are substituted, deleted or added to the sequence shown in SEQ ID NO: 4.

[0032] Gene mutant OsGLW3 RE The 3119th base in the coding region of the rice OsGLW3 gene mutated from T to A, resulting in premature termination of protein translation.

[0033] In a fifth aspect, the present invention provides a method comprising a gene mutant OsGLW3 RE The biological material includes, but is not limited to, recombinant DNA, expression cassettes, transposons, plasmid vectors, viral vectors, engineered bacteria or non-renewable plant parts.

[0034] In a sixth aspect, the present invention provides the gene mutant OsGLW3 RE Or any of the following applications of biological materials containing the gene mutant:

[0035] (1) Regulate plant grain shape, plant shape and ear shape;

[0036] (2) Used for plant variety improvement;

[0037] (3) Used for preparing transgenic plants.

[0038] In a seventh aspect, the present invention provides a method for making rice grains longer, narrower, with an increased length-to-width ratio, a reduced grain weight, an increased plant height, a thinner flag leaf, and an increased panicle length, the method comprising: using genetic engineering means to overexpress a gene mutant OsGLW3 in rice; RE .

[0039] The overexpression method may be selected from the following 1) to 5), or an optional combination:

[0040] 1) by introducing a plasmid carrying the gene;

[0041] 2) by increasing the copy number of the gene on the plant chromosome;

[0042] 3) by changing the promoter sequence of the gene on the plant chromosome;

[0043] 4) by operably linking a strong promoter to the gene;

[0044] 5) By introducing enhancers.

[0045] The expression vector carrying the target gene can be introduced into plant cells by conventional biotechnology methods such as Ti plasmid, plant virus vector, direct DNA transformation, microinjection, electroporation, etc. (Weissbach, 1998, Method for Plant Molecular Biology VIII, Academy Press, New York, pp. 411-463; Geiserson and Corey, 1998, Plant Molecular Biology, 2nd Edition).

[0046] In an eighth aspect, the present invention provides the use of the transgenic rice obtained according to the above method in plant breeding.

[0047] The breeding methods include, but are not limited to, transgenic, hybridization, backcrossing, selfing or asexual reproduction.

[0048] By means of the above technical solution, the present invention has at least the following advantages and beneficial effects:

[0049] (I) The present invention provides a rice gene OsGLW3. The present invention cloned a new gene OsGLW3 that controls rice grain shape. The coding region 3119T of the gene mutated to A, which showed a gain of function, specifically in that the grain length of the plant increased, and the grain width and weight decreased. The OsGLW3 gene is of great significance for the study of the molecular mechanism of rice grain shape formation. The present invention provides a new gene resource for rice grain shape improvement breeding, which greatly promotes the rice breeding process.

[0050] (ii) The grain shape and appearance quality of hybrid rice can be improved by hybridizing mutants carrying OsGCW3 with sterile lines.

[0051] (III) By building OsGLW3 RE The grain length of the transgenic plants transformed with wild-type Zhonghui 8015 after overexpression of the vector increased, which can be used to improve the grain length of rice to meet market demand. BRIEF DESCRIPTION OF THE DRAWINGS

[0052] Figure 1 The mature grains of ZH8015 and RE10498 in the preferred embodiment of the present invention. The scale is 5 mm. CF: grain width (C), grain length (D), length-to-width ratio (E), and thousand-grain weight (F) of ZH8015 and RE10498. Data: mean ± standard deviation (n = 20). P value was obtained by two-tailed t test.

[0053] Figure 2The agronomic traits of ZH8015 and RE10498 in the preferred embodiment of the present invention are compared. A, B: comparison of plant type and ear type of ZH8015 and RE10498, scale: A, 10 cm; B, 5 cm. CM: ear length (C), number of primary branches (D), number of secondary branches (E), sword leaf length (F), sword leaf width (G), plant height (H), number of effective ears (I), total number of grains per plant (J), fruit setting rate (K), number of grains per ear (L) and yield per plant (M) of ZH8015 and RE10498. Data: mean ± standard deviation (n = 20). P value was obtained by two-tailed t test.

[0054] Figure 3 The husk cytological observation of ZH8015 and RE10498 in the preferred embodiment of the present invention. A: phenotype, scale 5mm; B: husk cross-section, scale 50μm; C: enlarged view of the red frame area in Figure B, scale 100μm; DE: total length of thin-walled cells in the inner layer of the cross section (D), cell width (E); F, scanning electron microscopy of the outer surface of the husk, scale 400μm; GH, number of cells per unit area (G), cell length (H), cell width (I). Data: mean ± standard deviation. P value was obtained by two-tailed t test.

[0055] Figure 4 This is the frequency distribution diagram of the grain test data of 314 individual plants in the F2 generation in the preferred embodiment of the present invention. A: 1000-grain weight; B: length-to-width ratio; C: grain length; D: grain width.

[0056] Figure 5 This is the distribution diagram of Δ_All-index on chromosome in MutMap in a preferred embodiment of the present invention.

[0057] Figure 6 This is a functional verification of OsGLW3 in a preferred embodiment of the present invention. COM is the genome sequence of wild-type OsGLW3 transferred into the mutant, and OE is the OsGLW3 in the mutant overexpressed in the wild type. RE KO-1 is a knockout of OsGLW3 in the wild type. ZH KO-2 is the knockout mutant of OsGLW3 RE A, kernel phenotype; B, kernel length-width ratio; C, kernel length; D, kernel width. Data: mean ± SD (n greater than 6), Duncan multiple comparison.

[0058] Figure 7The expression pattern analysis of OsGLW3 in the preferred embodiment of the present invention. A: The expression level of OsGLW3 gene in various parts of ZH8015 and RE10498, Root: root; Node: node; Leaf, leaf; P0-1cm, P1-3cm, P3-5cm, P5-7cm, P7-10cm, P10-13cm, P13-17cm represent different ear lengths, respectively. GUS staining of transgenic plants, root (B), stem (C), leaf (D), leaf sheath (E), ear (F) and husk (G, H), the scale of BF is 2cm, and the scale of G and H is 2mm. Data: mean ± SD. P value was obtained by two-tailed t test. DETAILED DESCRIPTION

[0059] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention. Unless otherwise specified, the examples are all based on conventional experimental conditions, such as Sambrook et al. Molecular Cloning: a Laboratory Manual (Sambrook J & Russell DW, Molecular Cloning: a Laboratory Manual, 2001), or the conditions recommended by the manufacturer's instructions.

[0060] The rice material used in the present invention, Zhonghui 8015 (ZH8015), is a strong dominant three-line restorer line bred by hybridization with restorer line Zhonghui 218 as the female parent and Zhonghui 8006 as the male parent. The present invention uses ethyl methanesulfonate to induce mutation of ZH8015, and screens the slender grain mutant RE10498 in its mutant library. RE10498 and ZH8015 are hybridized, and the grains of their BC1F2 are tested. The grain width and thousand-grain weight show a continuous normal distribution, and the grain length and length-width ratio show different degrees of separation. The chi-square test is performed on the grain length, and the result shows that the grain length meets the 1:3 separation ratio, indicating that the grain length is controlled by a pair of dominant genes. The candidate gene LOC_Os03g21270 is obtained by using the MutMap method, and the 3119th position of its coding region is changed from thymine (T) to adenine (A), resulting in the mutation of leucine to a stop codon, causing premature termination of translation. We speculated that LOC_Os03g21270 was the target gene and named it OsGLW3. The function of the gene was subsequently verified by transgenic technology.

[0061] Temporal and spatial expression of gene OsGLW3 and GUS staining: RNA was extracted from various rice tissue samples, and the temporal and spatial expression of OsGLW3 was analyzed; the pCAMBLA1305-GUS vector was constructed to verify the temporal and spatial expression results.

[0062] Example 1 Gene localization

[0063] We identified a new rice grain shape mutant, RE10498. Compared with the wild type, the grain length of RE10498 mutant increased, and the grain width and grain weight decreased. Scanning electron microscopy of the husk revealed that the width of the epidermal cells of the husk of the mutant decreased, and the number of lateral cells decreased, which was consistent with the change in grain shape of the mutant. Genetic analysis showed that the grain length of the mutant was controlled by a pair of single dominant genes. The candidate gene LOC_Os03g21270 was obtained by MutMap method. The single base change T>A at position 3119 in its coding region caused leucine to mutate to a stop codon, causing premature translation termination. Subsequently, transgenic complementation, knockout and overexpression experiments confirmed that LOC_Os03g21270 was the cause of the mutant phenotypic changes, and it was named OsGLW3. Among them, by constructing its CRISPR / Cas9 gene knockout vector and then transforming ZH8015, there was no significant change in the grains of the transgenic plants, but after transforming the mutant RE10498, the grain length of the transgenic plants became shorter, and the grain width and grain weight increased, which was similar to the wild type; the gene function was verified by complementation experiments, and the mutant RE10498 was transformed with the pOsGLW3::OsGLW3 complementation vector, and the grain length, width and aspect ratio were between the wild type and the mutant; the gene function was verified by overexpression, and the pUbi::OsGLW3 RE The wild-type ZH8015 was transformed with the overexpression vector, and the grain length of the transgenic plants became longer, the grain width became narrower, and the grain weight decreased, which was similar to the mutant. OsGLW3 encodes an expression protein of unknown function and is a new gene that has not been reported in rice.

[0064] 1. Grain type phenotype

[0065] We used EMS to induce mutagenesis of ZH8015 and screened the mutant RE10498 with elongated grains in the mutant library. We found that compared with the wild-type ZH8015, the mutant RE10498 produced elongated and lighter grains ( Figure 1 ): The grain length of mutant RE10498 increased by 5.82%, the grain width decreased by 7.53%, the length-to-width ratio increased by 14.73%, and the 1000-grain weight decreased by 5.93%, all of which were extremely significant.

[0066] 2. Agronomic traits and yield traits

[0067] We investigated the plant type and yield-related traits of ZH8015 and RE10498. The results showed that the plant height of mutant RE10498 increased significantly compared with the wild type, the flag leaf became slender, and the plant type was slightly loose ( Figure 2 , A and FH). In terms of ear shape, the ear length, primary branch number and secondary branch number of RE10498 were significantly increased ( Figure 2, B and CE). Compared with the wild type, the total number of grains per plant of the mutant increased significantly, the fruit setting rate decreased significantly, and the number of effective tillers had no significant difference. Since the number of grains per ear of the mutant increased by 7.44%, it compensated for the decrease in grain weight, and ultimately led to no significant difference in yield per plant ( Figure 2 , IM).

[0068] 3. Cytological observation

[0069] Rice grain size is usually regulated by the number and size of glume cells. In order to explore how the mutant changes rice grain size, we observed the cross-section cells of the glume of ZH8015 and RE10498 before flowering. The total length and cell width of the inner thin-walled cells of mutant RE10498 were significantly reduced ( Figure 3 , AE). Scanning electron microscopy of the outer surface cells of the husk showed that the mutant had more cells than the wild type, the cell width decreased significantly, and the cell length did not change significantly ( Figure 3 The above results indicate that OsGLW3 makes the grains slender by regulating the longitudinal proliferation of husk cells and reducing the transverse cell width.

[0070] 4. Genetic analysis and gene localization

[0071] RE10498 and ZH8015 were crossed and their BC1F2 seeds were tested. The grain width and thousand-grain weight showed a continuous normal distribution, and the grain length and length-width ratio showed different degrees of separation ( Figure 4 The chi-square test for grain length showed that the grain length met the 1:3 segregation ratio, indicating that grain length was controlled by a pair of dominant genes (Table 1).

[0072] Table 1. Chi-square test of kernel length in BC1F2 generation

[0073]

[0074] Note: GL-L and GL-H represent grain lengths of 9.620-10.370 mm and 10.371-11.15 mm respectively.

[0075] Thirty plants with long and short grains in BC1F2 were selected to construct a mixed pool, and four candidate loci were obtained using the MutMap method ( Figure 5, Table 2). Among them, the thymine (T) at position 3119 in the coding region of OsR498G0305879600.01 (LOC_Os03g21270) was converted to adenine (A), resulting in the mutation of leucine to a stop codon, causing premature termination of translation. The other three candidate sites were all located upstream or downstream of the gene, so we speculated that LOC_Os03g21270 was the target gene and named it OsGLW3.

[0076] Table 2 List of candidate genes

[0077]

[0078] Example 2 Function of gene OsGLW3

[0079] In this example, pOsGLW3::OsGLW3 was transformed into mutant RE10498 by complementation experiment. The grain length and width of transgenic plants were between wild type and mutant. Furthermore, gene knockout experiment verified gene function. By constructing CRISPR / Cas9 gene knockout vector and transforming wild type ZH8015, there was no significant change in transgenic plant grains; while the grain length of transgenic plants transformed into mutant RE10498 was reduced, and the grain width and grain weight were increased. Furthermore, we used OsGLW3 in mutant to RE The coding sequence was used as a template to construct pUBi::OsGLW3 RE The overexpression vector was transformed into wild-type ZH8015, and the grain length of the transgenic plants increased, while the grain width and weight decreased. The details are as follows:

[0080] 1. Complementary verification

[0081] In order to verify whether the variation of the candidate gene OsGLW3 led to the change of the mutant grain shape and plant type, we first carried out a transgenic complementation test. We integrated the full-length 10.543Kb genome of OsGLW3 into the complementation vector (pCAMBIA1300, purchased from CAMBIA), and introduced the constructed complementation vector pOsGLW3::OsGLW3 into the RE10498 mutant through Agrobacterium-mediated transduction. We used hygromycin-specific primer PCR amplification and mutation site sequencing to conduct positive transgenic verification in the T0 generation. The test found that the grain length and width of the T1 generation complemented plants were between the wild type and the mutant ( Figure 6 ).

[0082] 2. Knockout verification

[0083] The knockout vector we used was the pCas9-sgRNA vector (MIAO et al., 2013), which has prokaryotic resistance to spectinomycin and eukaryotic resistance to hygromycin. AarI was used as the restriction site during the construction of the vector. The target sequence of the target gene was designed using the online website (http: / / cbi.hzau.edu.cn / cgi-bin / CRISPR2 / CRISPR), and the sequence N18 (5'-TGAAGCAGACCTCATGAG-3') with a higher score was selected, and the corresponding linker was added to form the F primer (5'-AGATGATCCGTGGCATGAAGCAGACCTCATGAGGTTTTAGAGCTATGC-3'), and the F primer sequence was reversely complementary to form the R primer. Add 1 μL of F and R primers to the 10 μL system, anneal at 94°C for 10 min, 0.1°C / s to 15°C, and keep at 15°C for 10 min to complete annealing; take 1 μL of annealing product and recombinant with pCas9-sgRNA vector plasmid after enzyme digestion, transfer into DH5α, and spread on spectinomycin-resistant solid culture medium; select single clones and sequence successfully, and send the plasmid to Wuhan Boyuan Biotechnology Co., Ltd. for transformation into ZH8015 background. Compared with ZH8015, there is no significant difference in the grain length and length-to-width ratio of the positive plants, and there is a slight difference in grain width ( Figure 6 ). We then transformed the pCas9-sgRNA recombinant plasmid containing the same sgRNA into the RE10498 background. Compared with RE10498, the grain length of the positive plants was reduced and the grain width was increased, showing grains more similar to those of ZH8015 ( Figure 6 ). The above results indicate that RE10498 is a gain-of-function mutant.

[0084] 3. Overexpression verification

[0085] To further verify the expression of OsGLW3 in mutant RE We used RE10498 cDNA as a template to amplify the mutated sequence of OsGLW3, SEQ ID NO: 5, and homologously recombined it into the overexpression vector pCAMBIA1301 (purchased from CAMBIA, maize Ubiquitin promoter) to construct the fusion vector pUbi::OsGLW3 RE The vector was introduced into ZH8015 (Wuhan Boyuan Biotechnology Co., Ltd.) using Agrobacterium-mediated method. The family with high expression level was selected from the T2 stable positive transgenic plants. The results showed that OsGLW3 was overexpressed in the ZH8015 background. RE This produces longer, narrower, lighter kernels ( Figure 6 ).

[0086] The above research results show that OsGLW3 REIt manifests itself as functional gain: increased grain length, and reduced grain width and weight.

[0087] Example 3 Analysis of the expression pattern of gene OsGLW3

[0088] In order to study the spatial expression of the gene OsGLW3, we used fluorescent quantitative PCR to analyze the expression of the gene in different parts. The results showed that OsGLW3 was expressed in roots, stems, leaves, leaf sheaths and panicles, but the expression level was higher in the panicle ( Figure 7 , A). In addition, we also verified this result by using transgenic plants expressing the β-glucuronidase (GUS) reporter gene driven by the OsGLW3 promoter: GUS staining was detected in roots, stems, leaves, leaf sheaths, and spikes, and the staining was darkest in the spikes ( Figure 7 , BH). This indicates that OsGLW3 is constitutively expressed, and its expression level is highest in the panicle.

[0089] Although the present invention has been described in detail above with general descriptions and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements can be made based on the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.

[0090] References:

[0091] 1. FAN CC, Genetics, 112(6): 1164-1171. DOI: 10.1007 / s00122-006-0218-1.

[0092] 2、LIU J F,CHEN J,ZHENG X M,WU F Q,LIN Q B,HENG Y Q,TIAN P,CHENG Z J,YU X W,ZHOU KN,ZHANG X,GUO X P,WANG J L,WANG H Y and WAN J M,2017.GW5 acts inthe brassinosteroid signalling pathway to regulate grain width and weight inrice.Nature Plants,3:17043.DOI:10.1038 / nplants.2017.43.

[0093] 3、MAO H L,SUN S Y,YAO J L,WANG C R,YU S B,XU C G,LI X H and ZHANG QF,2010.Linking differential domain functions of the GS3 protein to naturalvariation of grain size in rice.Proc Natl Acad Sci U S A,107(45):19579-19584.DOI:10.1073 / pnas.1014419107.

[0094] 4、MIAO J,Guo D S,ZHANG J Z,HUANG Q P,QIN G J,ZHANG X,WAN J M,GU H Yand QU L J,2013.Targeted mutagenesis in rice using CRISPR-Cas system.CellResearch,3;23(10):1233-1236.DOI:10.1038 / cr.2013.123.

[0095] 5、SONG X J,HUANG W,SHI M,ZHU M Z and LIN H X,2007.A QTL for ricegrain width and weight encodes a previously unknown RING-type E3 ubiquitinligase.Nature Genetics,39(5):623-630.DOI:10.1038 / ng2014.

[0096] 6. WANG SK, WU K, YUAN QB, LIU XY, LIU ZB, LIN XY, ZENG RZ, ZHU HT, DONG GJ, QIAN Q, ZHANG GQ, FU XD, 2012. Control of grain size, shape and quality by OsSPL 16in rice.Nature Genetics, 44(8):950-954.DOI: 10.1038 / ng.2327.

[0097] 7.WANG YX, in rice.Nature Genetics, 47(8):944-948.DOI: 10.1038 / ng.3346.

[0098] 8. Li Yang, Xu Xiaoyan, Yan Ming, Feng Fangjun, Ma Xiaosong and Mei Hanwei, 2016. Research on improving rice grain shape using GS3 gene functional molecular markers. Shanghai Journal of Agricultural Sciences (1): 1-5. DOI: 10.15955 / j.issn1000-3924.2016.01.01.

Claims

1. Rice gene OsGLW3, characterized in that It is a gene encoding the following protein (A) or (B): (A) a protein consisting of the amino acid sequence shown in SEQ ID NO: 3; or (B) A protein derived from (A) with equivalent functions, wherein one or more amino acids are substituted, deleted or added to the sequence shown in SEQ ID NO:

3.

2. The gene OsGLW3 according to claim 1, characterized in that Its nucleotide sequence is: I) the nucleotide sequence shown in SEQ ID NO: 1; II) a nucleotide sequence in which one or more nucleotides of the nucleotide sequence shown in SEQ ID NO: 1 are replaced, deleted and / or added and the nucleotide sequence expresses a protein with the same function; III) a nucleotide sequence that hybridizes with the sequence shown in SEQ ID NO: 1 under stringent conditions and expresses a protein with the same function, wherein the stringent conditions are hybridization at 65°C in a 0.1×SSPE solution containing 0.1% SDS or a 0.1×SSC solution containing 0.1% SDS, and the membrane is washed with the solution; or IV) is a nucleotide sequence that has more than 90% homology with the nucleotide sequence of I), II) or III) and expresses a protein with the same function.

3. Any of the following uses of the gene OsGLW3 according to claim 1 or 2 or a biological material containing the gene: (1) Regulate plant grain shape, plant shape and ear shape; (2) Used for plant variety improvement; (3) Used for preparing transgenic plants; The grain shape includes grain length, grain width and grain length-to-width ratio; The plant type includes plant height; The ear type includes ear length; Preferably, the plant is a grass plant, preferably rice.

4. Rice OsGLW3 gene mutant OsGLW3 RE , characterized in that, It is a gene encoding the following protein (a) or (b): (a) a protein consisting of the amino acid sequence shown in SEQ ID NO: 4; or (b) A protein derived from (a) with equivalent functions, wherein one or more amino acids are substituted, deleted or added to the sequence shown in SEQ ID NO:

4.

5. A biological material containing the gene mutant according to claim 4, wherein the biological material is recombinant DNA, an expression cassette, a transposon, a plasmid vector, a viral vector or an engineered bacterium.

6. Any of the following uses of the gene mutant according to claim 4 or the biomaterial according to claim 5: (1) Regulate plant grain shape, plant shape and ear shape; (2) Used for plant variety improvement; (3) Used for preparing transgenic plants; The grain shape includes grain length, grain width and grain length-to-width ratio; The plant type includes plant height; The ear type includes ear length; Preferably, the plant is a grass plant, more preferably rice.

7. A method for making rice grains longer, narrower, with a higher aspect ratio, lower grain weight, higher plant height, thinner flag leaves, and longer panicle length, characterized in that: The method comprises: using genetic engineering means to overexpress the gene mutant OsGLW3 in rice RE Among them, the gene mutant OsGLW3 RE As described in claim 4. Preferably, the plant is a grass plant, more preferably rice.

8. The method according to claim 7, characterized in that The overexpression method is selected from the following 1) to 5), or an optional combination: 1) by introducing a plasmid carrying the gene; 2) by increasing the copy number of the gene on the plant chromosome; 3) by changing the promoter sequence of the gene on the plant chromosome; 4) by operably linking a strong promoter to the gene; 5) By introducing enhancers.

9. Use of the transgenic rice obtained according to the method of claim 7 or 8 in plant breeding.

10. The use according to claim 9, characterized in that: Breeding methods include transgenic, hybridization, backcrossing, selfing or asexual reproduction.

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