Application of mutant maize ZmERECTA gene in regulating intercalary meristem development and dwarf breeding

Abstract

The application provides application of a mutant maize ZmERECTA gene in regulation of intercalary meristem development and dwarf breeding. The full length of the maize ZmERECTA gene CDS is 2985 bp, and strong mutants er-1, er-2 and er-3 of ZmERECTA are obtained by EMS mutagenesis, and the plant height and ear height of the three are all reduced, which indicates that the gene is a gene that can be used for maize dwarf breeding. Taking er-3 as an example, the intercalary meristem development of the mutant is disordered, which indicates that the gene regulates the intercalary meristem development. Meanwhile, the 1306th base of the ZmERECTA CDS is mutated from (guanine) G to (adenine) A by EMS mutagenesis, and a weak mutant er-750 of EMS mutagenesis is obtained. The plant height of er-750 is obviously reduced relative to B73, and the ten-grain length and ten-grain width of er-750 are obviously increased and widened relative to er-1. It is indicated that the 1306th base of the ZmERECTA CDS is mutated from (guanine) G to (adenine) A, which can reduce the plant height, but the seed size is improved relative to the strong mutant, and it is a mutant site that can be applied to maize dwarf breeding.

Description

TECHNICAL FIELD

[0001] The application belongs to the field of plant biotechnology, and particularly relates to the application of a maize ZmERECTA gene in regulating the development of intercalary meristem and a weak mutant er-750 of the gene in dwarf breeding. BACKGROUND

[0002] Maize is an important food and forage crop in China, and is related to the food security and economic development of China. Therefore, it is urgent to improve the yield of maize in China through modern breeding methods. With the frequent occurrence of extreme climate conditions, lodging has become one of the important factors limiting maize production. The use of heterosis to breed high-stalk maize hybrids to increase the yield increment is becoming smaller and smaller. Dwarf varieties have high yield due to their strong resistance to lodging, suitable height for high-density planting, and high photosynthetic efficiency. Therefore, breeding dwarf maize hybrids has become an important direction of maize breeding in recent years. The excavation and functional identification of dwarf genes are the basis for breeding dwarf varieties through molecular biology methods. Therefore, in-depth study of the genetic mechanism of maize plant height formation, cloning of a batch of new genes with independent intellectual property rights, and elucidation of various allelic variation types and their genetic effects in germplasm resources have important theoretical significance and application value for creating new maize varieties through biological breeding and serving the national strategy of revitalizing the seed industry.

[0003] The change of maize plant height is reflected in the number of nodes and the change of internode distance. From the perspective of anatomy, the number of nodes and the division and elongation of internode cells are regulated by the formation of intercalary meristem (Tsuda et al., 2017; Zhang et al., 2019; Zhang et al., 2018). From the perspective of gene regulation, the formation of intercalary meristem and the division and elongation of internode cells are also regulated by related specific expression genes and plant hormone related genes.

[0004] The intercalary meristem originates from the apical meristem. When the leaf primordium starts, the upper meristem begins to divide to form the intercalary meristem (Sharman, 1945). At the initial stage of plant development, the entire internode can be called the intercalary meristem. The development of the intercalary meristem is basipetal. With the growth and development of the plant, the intercalary meristem in the upper part of the internode continuously divides and differentiates to form mature cells, losing the characteristics of meristematic tissue. The intercalary meristem eventually accumulates at the base of the internode, still retaining some degree of meristematic characteristics, and eventually forms the well-known intercalary meristem. Disordered development of the intercalary meristem can cause changes in plant height. For example, the maize ZmTE1 (TERMINAL EAR) gene is significantly expressed in the intercalary meristem. The number of internodes increases and the internode distance shortens in the loss-of-function mutant. Microscopic observation shows that the intercalary meristem of the mutant develops disorderly and does not form the regular intercalary meristem structure of the wild type. The intercalary meristem is squeezed to the edge of the stem. At the same time, te1 differentiates into parenchyma earlier than the wild type, and the internode differentiates prematurely, ultimately leading to maize dwarfing (Wang et al., 2022). The ZmBLH12 / 14, a member of the maize BLH (BELL1-like homeobox) family of transcription factors, is highly expressed in the intercalary meristem and involved in the maintenance of meristematic tissue. After ZmBLH12 / 14 mutation, the maintenance of meristematic tissue is affected, and the intercalary meristem differentiates into parenchyma prematurely, leading to maize dwarfing (Tsuda et al., 2017). The Growth-Regulating Factor-Interacting Factor1 (GIF1), a co-transcription factor of the plant growth regulator Growth-Regulating Factor (GRF), regulates the proliferation of the intercalary meristem. The maize gif1 mutant exhibits dwarfing, and the leaf, leaf axil, and ear meristems are affected. This is because GIF1 can interact with GRF expressed in different parts to regulate maize growth and development (Zhang et al., 2018). Although multiple genes have been found to regulate the steady-state maintenance of the intercalary meristem and internode cell elongation in maize, the detailed signal transduction pathway and specific molecular mechanisms are still unclear.

[0005] Receptor-like kinases (RLKs) are located on the plasma membrane and are the primary receptors for signals from the environment or neighboring cells. They transmit extracellular signals to the intracellular by phosphorylation, thereby regulating plant growth and development and responses to environmental stimuli. Studies on the receptor kinase ERECTA in Arabidopsis have found that the signal transduction mediated by ERECTA plays an important role in regulating plant height. However, it is not yet clear whether ERECTA is involved in the development of the intercalary meristem in maize and thus affects plant height. SUMMARY

[0006] The application provides application of a mutant Zea mays ZmERECTA gene in regulation of intercalary meristem development and dwarf breeding. The application obtains a mutant Zea mays ZmERECTA gene, and verifies that mutation of the gene can disorder intercalary meristem of a plant and reduce plant height. Meanwhile, a weak mutant er-750 of the Zea mays ZmERECTA gene is obtained, the er-750 mutant has reduced plant height, and seed size is improved compared with that of a strong mutant er-1, and can be used for dwarf breeding.

[0007] In order to achieve the above purpose, the application adopts the following technical scheme:

[0008] 1. A Zea mays ZmERECTA gene, wherein a CDS sequence of the ZmERECTA gene is shown in a sequence table SEQ ID NO. 1 and a sequence table SEQ ID NO. 3, a nucleotide length is 2985 bp, and a full-length sequence of the gene is shown in the sequence table SEQ ID NO. 3, and a sequence ID published by Phytozome (https: / / phytozome-next.jgi.doe.gov / ) is Zm00001d037114. Figure 7

[0009] 2. A mutant Zea mays ZmERECTA gene, wherein a CDS sequence of the mutant Zea mays ZmERECTA gene is shown in a sequence table SEQ ID NO. 2 and a sequence table SEQ ID NO. 3, a nucleotide length is 2985 bp, a base at a position of 1306 is A, and a mutant produced by the mutant Zea mays ZmERECTA gene is er-750. Figure 7

[0010] 3. A mutant Zea mays ZmERECTA gene, wherein a base G at a position of 4687 in a sequence shown in a sequence table SEQ ID NO. 3 is mutated to A, that is, a boundary mutation (CAG is mutated to CAA) between a 23rd exon and a 22nd intron occurs, variable splicing occurs, and a mutant produced by the mutant Zea mays ZmERECTA gene is er-1.

[0011] 4. A mutant Zea mays ZmERECTA gene, wherein a base G at a position of 2875 in a sequence shown in a sequence table SEQ ID NO. 3 is mutated to A, that is, a boundary mutation (CAG is mutated to CAA) between a 12th exon and an 11th intron occurs, variable splicing occurs, and a mutant produced by the mutant Zea mays ZmERECTA gene is er-2.

[0012] ​​5. A mutant ZmERECTA gene of Zea mays, wherein the mutant ZmERECTA gene is a sequence shown in SEQ ID NO. 3, and a C at 4263 bp is mutated to a T, i.e., a premature termination mutation (CAA mutated to TAA) occurs in the 20th exon, and mRNA translation is prematurely terminated, and the mutant ZmERECTA gene produces a mutant er-3.

[0013] 6. Application of a mutant ZmERECTA gene of Zea mays in maize dwarf breeding, wherein the mutant ZmERECTA gene is shown in SEQ ID NO. 2, and has a nucleotide length of 2985 bp, and 1306 is a base A, and the mutant ZmERECTA gene produces a mutant er-750.

[0014] 7. Application of a mutant ZmERECTA gene of Zea mays in regulating intercalary meristem development, wherein the mutant ZmERECTA gene is shown in SEQ ID NO. 2, and has a nucleotide length of 2985 bp, and 1306 is a base A, and the mutant ZmERECTA gene produces a mutant er-750.

[0015] 8. Application of the mutant ZmERECTA gene of Zea mays in regulating intercalary meristem development according to any one of 3-5, wherein the mutant ZmERECTA gene produces mutants er-1, er-2, and er-3 in sequence.

[0016] 9. Application of the mutant ZmERECTA gene of Zea mays in maize dwarf breeding according to any one of 3-5, wherein the mutant ZmERECTA gene produces mutants er-1, er-2, and er-3 in sequence.

[0017] 10. A method for verifying the application of a ZmERECTA gene of Zea mays in regulating intercalary meristem development, comprising: (1) EMS mutagenesis of strains er-1, er-2, and er-3, (2) observation of the plant height of er-1, er-2, and er-3, (3) observation of the intercalary meristem of er-3, and (4) analysis of ZmERECTA tissue expression.

[0018] 11. A method for verifying the application of a weak mutant er-750 of a ZmERECTA gene of Zea mays in maize dwarf breeding, comprising: (1) EMS mutagenesis of strain er-750, (2) observation of the plant height of er-750, and (3) observation of the seed morphology of er-750.

[0019] Compared with the prior art, the present application has the beneficial effects that the ZmERECTA gene is isolated and cloned from corn, and three EMS mutagenesis mutants (strong mutants) of the ZmERECTA gene are obtained, the mutant plants are compared with wild type plants grown under the same conditions, the plants with ZmERECTA gene mutation show the phenotypes of disordered intercalary meristem and dwarfed plant height, and the corn ZmERECTA gene is highly expressed in plant nodes and internodes, so it is proved that the ZmERECTA gene is involved in the regulation of plant intercalary meristem development. At the same time, the EMS mutagenesis weak mutant er-750 is obtained. The plant height of er-750 is significantly reduced relative to B73, and the ten-grain length and ten-grain width of er-750 are significantly improved relative to er-1. It is indicated that by mutating the 1306th base of ZmERECTA from (guanine) G to (adenine) A, the plant height can be reduced, but the seed size is improved relative to the strong mutant, and it is a mutant site that can be applied to corn dwarf breeding. Therefore, the ZmERECTA gene has high agricultural application value. BRIEF DESCRIPTION OF DRAWINGS

[0020] Figure 1 .EMS mutagenesis mutant er-1, er-2, er-3, er-750 mutant site schematic diagram

[0021] Figure 2 .EMS mutagenesis mutant er-1, er-2, er-3, er-750 mutant site sequencing detection Figure 2 A, er-1 mutant site sequencing detection Figure 2 B, er-2 mutant site sequencing detection Figure 2 C, er-3 mutant site sequencing detection Figure 2 D, er-750 mutant site sequencing detection

[0022] Figure 3 .zmerecta mutant phenotype observation Figure 3 A, zmerecta mutant plant height phenotype; scale = 20 cm Figure 3 B, zmerecta mutant ear position height phenotype; scale = 20 cm Figure 3 C and Figure 3 D are plant height data statistics and ear position height data statistics, respectively, and Student's t-tests are used to analyze the differences between B73 and EMS mutagenesis mutants (ns, p>0.1; *, p<0.1; **, p<0.01; ***, p<0.001).

[0023] Figure 4 .er-750 mutant phenotype observation Figure 4A, er-750 mutant plant height phenotype; scale = 20 cm; Figure 4 B, er-750 mutant seed size phenotype; scale = 2 cm; Figure 4 C, Figure 4 D, Figure 4 E, respectively, plant height data statistics, ten-grain length data statistics, ten-grain width data statistics, Student's t-tests were used to analyze the differences between B73 and EMS mutagenesis mutants (ns, p > 0.1; *, p < 0.1; **, p < 0.01; ***, p < 0.001).

[0024] Figure 5 Intermediary meristem observation of ZmERECTA mutant. B73 and er-3 intermediary meristem phenotype; scale = 500 μm.

[0025] Figure 6 ZmERECTA tissue expression analysis. qRT-PCR was used to detect the expression of ZmERECTA in various tissues of wild type, including primary root, seminal root, crown root, crown root node, mesocotyl, and from bottom to top, three nodes and internodes, tassel, ear, and leaf of six-week-old plants. Primary root: main root; Seminal root: seed root; Crown root: crown root; Crown root node: crown root node; Mesocotyl: mesocotyl; 1st node: first node; 1st internode: first internode; 2nd node: second node; 2nd internode: second internode; 3rd node: third node; 3rd internode: third internode; Tassel: tassel; Ear: ear; Leaf: leaf.

[0026] Figure 7 Maize ZmERECTA gene, in which the underlined is the start and stop codon, and the box is the 1306 site of er-750 mutant, which is mutated from G in wild type to A in er-750 mutant. DETAILED DESCRIPTION

[0027] The methods and devices used in the following examples of the present application are all conventional methods and devices unless otherwise specified; the equipment and reagents used are all conventional equipment and reagents purchased from reagent companies. In order to make the purpose, technical scheme and advantages of the present application clearer, the specific embodiments of the present application will be described in detail below with reference to the drawings. The examples of these preferred embodiments are illustrated in the drawings. The embodiments of the present application shown in the drawings and described according to the drawings are merely exemplary, and the present application is not limited to these embodiments. Here, it also needs to be explained that, in order to avoid obscuring the technical scheme of the present application due to unnecessary details, only the processing steps closely related to the scheme according to the present application are shown in the drawings, and other details not closely related are omitted.

[0028] Example 1

[0029] This embodiment provides the source and phenotype detection method of zmerecta mutant, specifically comprising:

[0030] 1. Obtaining of zmerecta mutant

[0031] The applicant ordered three EMS mutagenesis mutants (strong mutants) of ZmERECTA gene from the near-saturation B73 background corn EMS mutagenesis mutant library (http: / / maizeems.qlnu.edu.cn / search / lists.html), which are named er-1, er-2 and er-3 respectively. And through sequencing, it is found that er-1 is a mutation (CAG is mutated to CAA) at the boundary between 4687bp base, i.e. 23rd exon and 22nd intron, which causes alternative splicing. Er-2 is a mutation (CAG is mutated to CAA) at the boundary between 2875bp base, i.e. 12th exon and 11th intron, which causes alternative splicing. Er-3 is a premature termination mutation (CAA is mutated to TAA) at 4263bp, i.e. in the 20th exon, which causes premature termination of mRNA translation, and the results are shown in Figure 1 .

[0032] The weak mutant er-750 is also ordered from the near-saturation B73 background corn EMS mutagenesis mutant library (http: / / maizeems.qlnu.edu.cn / search / lists.html), which is a non-synonymous mutation caused by mutation of 1306th base of ZmERECTA CDS from (guanine) G to (adenine) A, and the results are shown in Figure 1 .

[0033] 2. Detection of zmerecta mutant

[0034] In order to identify the zmerecta mutant, the applicant designed four pairs of primers for PCR amplification and sequencing identification, and the identification results are as follows:Figure 2 Figure 1 shows the phenotype of the er-1 mutant (A), the er-2 mutant (B), the er-3 mutant (C) and the er-750 mutant (D) compared to the wild type (WT) maize plant. Figure 2 A: sequencing detection of the er-1 mutant site; Figure 2 B: sequencing detection of the er-2 mutant site; Figure 2 C: sequencing detection of the er-3 mutant site; Figure 2 D: sequencing detection of the er-750 mutant site), the mutation of the relevant site can be clearly observed.

[0035] er-1-F: 5'-ATCATCTTGGTGGTCTGATTCC-3';

[0036] er-1-R: 5'-ATCAGGAGGATAACAAGTCCACC-3';

[0037] er-2-F: 5'-GGGTTTGCGGGGAAATCATT-3';

[0038] er-2-R: 5'-TGCTCTAGGTTGCCAATGGA-3';

[0039] er-3-F: 5'-TTGGACACGTTGTAAGCCTTC-3';

[0040] er-3-R: 5'-CTTACGTTTGTCTCGGTGGC-3';

[0041] er-750-F: 5'-ATGGAACCATTCCTCGTTCG-3';

[0042] er-750-R: 5'-AAACTCCGCAGGGATGAATC-3'.

[0043] 3. Phenotype detection of zmerecta mutant

[0044] The wild type maize B73 was used as a control (WT). The phenotypes of the er-1, er-2 and er-3 mutant maize plants were observed and statistically analyzed. The results are shown in Figure 2. Figure 3 Figure 2 shows the phenotype of the er-1 mutant (A), the er-2 mutant (B) and the er-3 mutant (C) compared to the wild type (WT) maize plant. Figure 3 A, the phenotype of the zmerecta mutant plant height; Figure 3 B, the phenotype of the zmerecta mutant ear height; Figure 3 C and Figure 3 D are the statistical data of the plant height and the statistical data of the ear height, respectively), the three alleles of er-1, er-2 and er-3 all show a significant decrease in plant height, which also indicates that the decrease in plant height of the mutant is caused by the mutation of ZmERECTA.

[0045] Wild type corn B73 was used as a control (WT). The phenotypes of er-1 and er-750 mutant corn plants and seeds were observed and counted. The results are shown in Table 1. Figure 4 Figure 4 A, er-750 mutant plant height phenotype; Figure 4 B, er-750 mutant seed size phenotype; Figure 4 C, Figure 4 D, Figure 4 E, respectively, plant height data statistics, ten grain length data statistics, and ten grain width data statistics), the plant height of er-1 and er-750 mutants was significantly reduced, but the seed size of er-750 mutant was improved compared with er-1.

[0046] Example 2

[0047] This example provides a method for observing the intercalary meristem of zmerecta mutants, which specifically comprises:

[0048] 1. Fixation:

[0049] FAA fixing solution (70% alcohol 90ml, glacial acetic acid 5ml, formaldehyde 5ml) was used to fix the wild type and zmerecta mutant tender stem tips grown for 27 days for more than 48 hours but not more than 72 hours.

[0050] 2. Dehydration:

[0051] 70% alcohol→80% alcohol→95% alcohol→100% alcohol→100% alcohol. 2h per level. 1.5h in 100% alcohol. The 70% alcohol can be stored for a long time.

[0052] 3. Transparency:

[0053] Xylene: anhydrous ethanol (1:1) (2h)→xylene (1.5h)→xylene (1.5h)

[0054] 4. Wax immersion:

[0055] The treated material was placed in paraffin (solid powder): xylene (1:1) in a 37℃ oven overnight (must be done in a fume hood).

[0056] 5. Embedding:

[0057] First, raise the temperature of the incubator to 60℃, replace the pure wax 3 times, 1-2h each time, use hard electric light paper, cowhide paper, and paper box, place it on the ironing board at 45-60℃, pour the material, place the material, put the label (front outward) on the bottom, add paraffin, pour it gently into the cold water basin, make sure the bottom surface is in contact with the cold water in the basin, and take it out to dry after the paraffin is completely solidified. It can also be placed in a cold water basin overnight. ​

[0058] 6. Slicing

[0059] After trimming, sticking, and finishing the embedded material, make sure that the material is surrounded by paraffin on all sides. However, do not use too much. The upper and lower edges of the section should be parallel. Dip a heated scalpel in a small amount of paraffin debris, and quickly flatten the debris around the paraffin block to make the paraffin block firmly stick to the table. Check the microtome, install the sectioning knife, adjust the angle of the knife, and adjust the angle and position between the paraffin block and the knife edge, and then start sectioning.

[0060] 7. Mounting

[0061] Place the adhesive on the glass slide, then take the section and place it on the adhesive, and then place it on the sectioning table to expand and flatten the section. The material should not have wrinkles. Finally, arrange the sections in order, use filter paper to absorb excess water, and use a marker to number the glass slides. Place the glass slides in a drying oven at a temperature of 30-40°C overnight.

[0062] 8. De-paraffinization

[0063] Use xylene for de-paraffinization. Place the baked glass slide in a dye vat containing xylene: xylene (10-20 min) → xylene (5-10 min)

[0064] 9. Staining

[0065] Stain the completely de-paraffinized material with 0.25% toluidine blue staining solution for 2 min (the staining time can be appropriately extended or shortened), wash off the staining solution with ultrapure water, and dry the glass slide at 42°C.

[0066] 10. Sealing

[0067] Add neutral gum dropwise, and cover with a cover glass to seal.

[0068] 11. Microscopy and photography

[0069] The results of the intercalary meristem are shown in Figure 5 The wild-type B73 can clearly observe the layer-by-layer intercalary meristem structure, while the zmerecta mutant has almost no such layer-by-layer structure, indicating that the intercalary meristem of the mutant is disordered.

[0070] Example 3

[0071] The present example provides a method for analyzing the tissue-specific expression of ZmERECTA, which specifically comprises:

[0072] 1. Extraction of RNA: Use the Tiangen DP432 RNA extraction kit to extract total RNA from corn;

[0073] (1) Material collection, the materials are taproot, seed root, crown root, crown root node, mesocotyl, and the three nodes and internodes from bottom to top, tassel, ear, leaf and other parts of the growth of one week, each material takes 3 parts, and is immediately frozen with liquid nitrogen after collection;

[0074] (2) Homogenate treatment, the material is rapidly ground into powder in liquid nitrogen, 450 μL RL (check whether β-mercaptoethanol has been added before use, and make the final concentration 1%) is added, vortexed vigorously, and incubated at 56°C for 1-3 min;

[0075] (3) Transfer all solutions to filter column CS (filter column CS is placed in a collection tube), centrifuge at 12000 rpm for 2-5 min, carefully suck the supernatant in the collection tube into an RNase-Free centrifuge tube, and try to avoid touching the cell debris precipitate in the collection tube with the suction head;

[0076] (4) Slowly add 0.5 times the supernatant volume of anhydrous ethanol, mix well, and transfer to the adsorption column CR3, centrifuge at 12000 rpm for 30-60 sec, discard the waste liquid in the collection tube, and place the adsorption column CR3 back into the collection tube;

[0077] (5) Add 350 μL deproteinization solution RW1 to the adsorption column CR3, centrifuge at 12000 rpm for 30-60 sec, discard the waste liquid in the collection tube, and place the adsorption column CR3 back into the collection tube;

[0078] (6) Preparation of DNase I working solution: take 10 μL DNase I storage solution into a new RNase-Free centrifuge tube, add 70 μL RDD buffer, and mix gently;

[0079] (7) Add 80 μL of DNase I working solution to the center of the adsorption column CR3, and place it at room temperature for 15 min;

[0080] (8) Add 350 μL deproteinization solution RW1 to the adsorption column CR3, centrifuge at 12000 rpm for 30-60 sec, discard the waste liquid in the collection tube, and place the adsorption column CR3 back into the collection tube;

[0081] (9) Add 500 μL of rinse solution RW (check whether ethanol has been added before use) to the adsorption column CR3, stand at room temperature for 2 min, centrifuge at 12000 rpm for 30-60 sec, discard the waste liquid in the collection tube, and place the adsorption column CR3 back into the collection tube;

[0082] (10) Repeat step 9;

[0083] (11) 12000rpm centrifugation for 2min, and pour out the waste liquid. Put the adsorption column CR3 in room temperature for several minutes to dry the residual rinse liquid in the adsorption material completely;

[0084] (12) Put the adsorption column CR3 in a new RNase-Free centrifuge tube, and add 30-100μL RNase-Free ddH2O to the middle of the adsorption membrane, and then put it in room temperature for 2min, and centrifuge at 12000rpm for 2min to obtain the RNA solution.

[0085] 2. RNA reverse transcription:

[0086] After dissolving the extracted RNA, the RNA concentration was determined, and then the 1st Strand cDNA Synthesis SuperMix reverse transcription kit was used for reverse transcription. Take 5μg total RNA, add SuperMix 10μL, and add water to 20μL, and incubate at 25℃ and 42℃ for 5min and 30min respectively, and then inactivate the enzyme at 85℃ for 5min.

[0087] 3. Real-time fluorescent quantitative PCR

[0088] Use the primer and ZmERECTA gene primer pair to analyze the tissue expression pattern of ZmERECTA by qRT-PCR:

[0089] ACTIN-F: 5'-GAGCGGGAGATTGTCAGGG-3';

[0090] ACTIN-R: 5'-AAGGGATGGTTGGAACAGCA-3';

[0091] ZmERECTA-F: 5'-GACAAACCGCCAATCTCAAAGG-3';

[0092] ZmERECTA-R: 5'-AAAGCAGGTGGATGGTGTGG-3';

[0093] Use the qRT-PCR special 96-well plate and high light transmittance sealing film, and use the fluorescent quantitative PCR instrument Applied Biosystems StepOnePlus TM Real-Time PCR system to perform qRT-PCR analysis, 3 times repeated for each sample, and the reaction system refers to qPCR SYBR Green Master Mix(High Rox Plus) instruction, the reaction condition is as follows: 95℃ pre-denaturation 5min; amplification procedure is 95℃, 10sec, 60℃, 30sec, 40 cycles in total.

[0094] Results as shown in Fig. 1, the maize ZmERECTA gene is highly expressed in the corn node and internode. Figure 6

[0095] The above evidence shows that the phenotype of zmerecta mutant is specifically the development disorder of intercalary meristem, causing the mutant plant height to be dwarf, and the maize ZmERECTA gene is highly expressed in the corn node and internode. At the same time, the plant height of the weak mutant er-750 is significantly lower than that of the wild type, but the seed size of the er-750 mutant is improved compared with the strong mutant er-1, which indicates that the mutation site of er-750 is a better mutation site for dwarf breeding.

[0096] The above is only a specific embodiment of the present application, it should be noted that for those skilled in the art, without departing from the principles of the present application, can make a number of improvements and refinements, these improvements and refinements should also be considered as the protection scope of the present application.

[0097] Main references:

[0098] Sharman B.C.(1945).Leaf and bud ination in the Gramineae.Bot.Gaz.106:269-289.

[0099] Tsuda, K., Abraham-Juarez, M.J., Maeno, A., Dong, Z., Aromdee, D., Meeley, R., Shiroishi, T., Nonomura, K.I., and Hake, S. (2017). KNOTTED1 cofactors, BLH12 and BLH14, regulate internode patterning and vein anastomosis in maize. Plant Cell 29: 1105-1118.

[0100] ​Wang, F., Yu, Z., Zhang, M., Wang, M., Lu, X., Liu, X., Li, Y., Zhang, X., Tan, B., Li, C., and Ding, Z. (2022). ZmTE1 promotes plant height by regulating intercalary meristem formation and internode cell elongation in maize. Plant Biotechnology Journal 20: 526-537.

[0101] Zhang, D., Sun, W., Singh, R., Zheng, Y., Cao, Z., Li, M., Lunde, C., Hake, S., and Zhang, Z.

[0102] (2018). GRF-interacting factor 1 Regulates Shoot Architecture and Meristem Determinacy in Maize. Plant Cell 30: 360-374.

[0103] Zhang, X., Hou, X., Liu, Y., Zheng, L., Yi, Q., Zhang, H., Huang, X., Zhang, J., Hu, Y., Yu, G., Liu, H., Li, Y., Huang, H., Zhan, F., Chen, L., Tang, J., and Huang, Y. (2019). Maize brachytic2

[0104] (br2) suppresses the elongation of lower internodes for excessive auxin accumulation in the intercalary meristem region. BMC Plant Biology 19: 589.

Claims

1. Mutant maize ZmERECTA gene characterized by, The mutant corn ZmERECTA The gene CDS is shown in the sequence listing SEQ ID NO. 2, with a nucleotide length of 2985 bp, wherein the base at position 1306 is A.

2. Mutant maize ZmERECTA Use of the gene in breeding for maize dwarfing, characterized in that, The mutant corn ZmERECTA The gene is shown in SEQ ID NO. 2 of the sequence listing, with a nucleotide length of 2985 bp, of which 1306 are A bases.

3. Mutant maize ZmERECTA The use of the gene in regulating the development of the intercalary meristem is characterized in that, The mutant corn ZmERECTA ZmERECTA The gene is shown in SEQ ID NO. 2 of the sequence listing, with a nucleotide length of 2985 bp, of which 1306 are A bases.

Images