Mbl protein, encoding gene and use thereof in promoting amylopectin synthesis

Abstract

The application discloses MBL protein, coding genes and application thereof in promoting amylopectin synthesis, and belongs to the technical field of plant genetic engineering. The application first determines the function of metal beta-lactamase MBL in starch synthesis in tomatoes by using reverse genetics, and the MBL promotes the synthesis of amylopectin by interacting with soluble starch synthase III (SS3) and improving the activity of the SS3. Therefore, the activity of the SS3 can be influenced by adjusting the content of the MBL, and then the accumulation level of amylopectin is regulated. The MBL-mediated starch synthesis mechanism discovered by the application has important significance for optimizing the yield and quality of crops such as tomatoes.

Description

Technical Field

[0001] This invention relates to the field of plant genetic engineering technology, specifically to the MBL protein, its encoding gene, and its application in promoting amylopectin synthesis. Background Technology

[0002] Starch, as the main carbohydrate produced by plant photosynthesis, is crucial for maintaining crop environmental adaptability, yield, and quality. Starch is not only vital for human food security but also an important feed component and industrial raw material.

[0003] Starch is composed of amylose and amylopectin. Currently identified key enzymes involved in starch synthesis include ADP-glucose pyrophosphorylase (AGPase), starch synthase (SS), starch branching enzyme (BE), and starch debranching enzyme (DBE). In plants, AGPase catalyzes the reaction of glucose-1-phosphate with ATP to produce ADP-glucose, the substrate for starch synthesis. ADP-glucose then synthesizes amylose and amylopectin under the catalysis of SS, BE, and DBE. The applications of starch in different fields depend on its molecular structure, and the ratio of amylose to amylopectin (amylose-amylose ratio) is the most important parameter affecting starch molecular structure. Therefore, a thorough understanding of the key mechanisms of starch synthesis is crucial for increasing starch accumulation and optimizing its composition.

[0004] The deletion of different members of the starch synthase family (SS1-SS5) has varying degrees of impact on starch content, amylopectin chain length distribution, and starch granule morphology. Except for SS4, the individual deletion of other SS members did not produce obvious phenotypes, while the ss1ss3 double mutant or ss1ss2ss3 triple mutant lines exhibited significant dwarfism and extremely low starch content. This indicates that each SS member plays an important role in starch synthesis both independently and collaboratively. However, current research progress on the regulatory mechanisms of SS is still very limited, and further exploration of the mechanisms of action and regulatory mechanisms at different levels of different SS members is an urgent problem to be solved.

[0005] Metallo-β-lactamases (MBLs) are typical β-lactamases. Bacterial production of β-lactamases can lead to resistance to β-lactam antibiotics (such as penicillins and cephalosporins). The paper "Metallo-β-Lactamases: Structure, Function, Epidemiology, Treatment Options, and the Development Pipeline" (Sara E Boyd, Antimicrob Agents Chemother.) describes the structure and function of MBLs, suggesting that bacterial resistance to β-lactams is primarily due to the disruption of the core β-lactam ring by the produced β-lactamase. This bacterial resistance presents a unique challenge to drug development, making MBLs a hot research topic in the biomedical field. Although MBL proteins also exist in plants, their homology with bacterial MBLs is low, and their specific functions have long remained unknown. Summary of the Invention

[0006] In view of the above-mentioned prior art, the purpose of this invention is to provide the MBL protein, its encoding gene, and its application in promoting amylopectin synthesis. This invention, using reverse genetics, has for the first time clarified the function of the metallo-β-lactamase MBL in starch synthesis in tomato. MBL promotes amylopectin synthesis by interacting with soluble starch synthase III (SS3) and increasing the activity of SS3. Therefore, by regulating the MBL content, the activity of SS3 can be affected, thereby regulating the accumulation level of amylopectin. The MBL-mediated starch synthesis mechanism discovered in this invention is of great significance for optimizing the yield and quality of crops such as tomatoes.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] A first aspect of the invention provides the use of MBL protein in any of the following i)-iii):

[0009] i) Regulate the activity of plant soluble starch synthase III;

[0010] ii) Regulates the synthesis of amylopectin in plants;

[0011] iii) It affects the synthesis of amylopectin in vitro via soluble starch synthase III;

[0012] The amino acid sequence of the MBL protein is shown in SEQ ID NO.1.

[0013] A second aspect of the invention provides the use of a gene encoding the MBL protein in any of the following i)-iv):

[0014] i) Regulate the activity of plant soluble starch synthase III;

[0015] ii) Regulates the synthesis of amylopectin in plants;

[0016] iii) It affects the synthesis of amylopectin in vitro via soluble starch synthase III;

[0017] iv) Plant breeding.

[0018] Furthermore, the gene is a nucleic acid molecule as shown in (1) or (2) below:

[0019] (1) The nucleotide sequence is the nucleic acid molecule shown in SEQ ID NO.2;

[0020] (2) Nucleic acid molecules other than (1) that encode the amino acid sequence shown in SEQ ID NO.1.

[0021] A third aspect of the invention provides the use of a gene-containing expression cassette, recombinant expression vector, or recombinant bacteria in any of the following i)-iv):

[0022] i) Regulate the activity of plant soluble starch synthase III;

[0023] ii) Regulates the synthesis of amylopectin in plants;

[0024] iii) It affects the synthesis of amylopectin in vitro via soluble starch synthase III;

[0025] iv) Plant breeding.

[0026] A fourth aspect of the present invention provides a method for promoting the synthesis of amylopectin in plants, the method being as follows:

[0027] Increasing the content of plant MBL protein or increasing the activity of soluble starch synthase III can promote the synthesis of plant amylopectin.

[0028] The amino acid sequence of the MBL protein is shown in SEQ ID NO.1.

[0029] In a fifth aspect, the present invention provides a method for inhibiting the synthesis of amylopectin in plants, the method being as follows:

[0030] It reduces the content of plant MBL protein or reduces the activity of soluble starch synthase III, thereby inhibiting the synthesis of plant amylopectin.

[0031] The amino acid sequence of the MBL protein is shown in SEQ ID NO.1.

[0032] In a sixth aspect, the present invention provides a method for breeding plants with high amylopectin content, the method being as follows:

[0033] Plants with higher amylopectin content than the target plant were obtained by increasing the content of MBL protein or increasing the activity of soluble starch synthase III.

[0034] The amino acid sequence of the MBL protein is shown in SEQ ID NO.1.

[0035] In a seventh aspect, the present invention provides a method for breeding plants with low amylopectin content, the method being as follows:

[0036] Plants with amylopectin content lower than that of the target plant were obtained by reducing the content of MBL protein or reducing the activity of soluble starch synthase III.

[0037] The amino acid sequence of the MBL protein is shown in SEQ ID NO.1.

[0038] Furthermore, the target plant is tomato.

[0039] The beneficial effects of this invention are:

[0040] This invention utilizes reverse genetics to elucidate for the first time the function of the metallo-β-lactamase MBL in starch synthesis in tomato. MBL promotes amylopectin synthesis by interacting with soluble starch synthase III (SS3) and increasing SS3 activity. Therefore, regulating the MBL content can affect SS3 activity, thereby controlling the accumulation level of amylopectin. The MBL-mediated starch synthesis mechanism discovered in this invention is of great significance for optimizing the yield and quality of crops such as tomatoes. Attached Figure Description

[0041] Figure 1 This represents the mutation site of the MBL mutant.

[0042] Figure 2 The results of qRT-PCR detection of MBL expression levels were obtained from the overexpressing MBL lines MBL-OE1 and MBL-OE2, the RNAi-inhibited MBL expression line MBL-RNAi, and the wild-type line WT.

[0043] Figure 3 Phenotypic representations of tomato MBL overexpression lines, RNAi-repressed expression lines, and Crisper-Cas9-mediated mutant lines. The top and bottom rows show tomato MBL-OE1, MBL-OE2, WT, mbl-1, mbl-2, and MBL-RNAi plants at the seedling and flowering stages, respectively. The top and bottom rows also show photographs of the same subject and experimental group at different stages.

[0044] Figure 4 The dry and fresh weights are for tomato MBL overexpression lines, RNAi-repressed expression lines, and Crisper-Cas9-mediated mutant lines. The upper and lower rows of samples were collected from... Figure 1 The corresponding tomato MBL-OE1, MBL-OE2, WT, mbl-1, mbl-2, and MBL-RNAi seedlings and flowering plants.

[0045] Figure 5 Starch staining analysis of leaves from tomato MBL overexpression lines, RNAi suppressed expression lines, and Crisper-Cas9 mediated mutant lines.

[0046] Figure 6 This study aimed to determine the starch, amylopectin, amylose, and soluble sugar content in the leaves of tomato MBL overexpression lines, RNAi suppressed expression lines, and Crisper-Cas9 mediated mutant lines.

[0047] Figure 7 To observe the starch accumulation in chloroplasts of tomato MBL overexpression lines, RNAi suppressed expression lines, and Crisper-Cas9 mediated mutant lines using transmission electron microscopy, black arrows indicate starch in chloroplasts.

[0048] Figure 8 This study used enzyme profiling to analyze the starch synthase (SS) activity in leaves of tomato MBL overexpression lines, RNAi-inhibited expression lines, and mutant lines. Darker bands indicate higher activity.

[0049] Figure 9 To purify tomato MBL and SS3 proteins through prokaryotic induction, the effect of MBL on SS3 activity was analyzed using enzyme spectrometry. The five samples were MBL-His+SS3-MBP, MBL-His+SS3, etc. MBD2 -MBP, His and SS3-MBP, MBL-His and MBP, SS3 MBD2 -MBP; the darker the band, the higher the activity. Detailed Implementation

[0050] It should be noted that the following detailed descriptions are illustrative and intended to provide further explanation of this application. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains.

[0051] As mentioned earlier, starch, as the main carbohydrate produced by plant photosynthesis, is crucial for maintaining crop environmental adaptability, yield, and quality. Starch is not only vital for human food security but also an important feed component and industrial raw material. Therefore, discovering new mechanisms regulating starch synthesis and identifying novel starch synthesis regulators are essential for crop starch improvement and industrial starch production.

[0052] Based on this, this invention conducted an in-depth study on proteins that affect the starch content and starch synthase activity of tomatoes. Bacterial metallo-β-lactamases (MBLs) are typical β-lactamases that depend on Zn. 2+ The active site for cleaving the β-lactam ring of almost all β-lactam antibiotics is one of the main factors contributing to bacterial resistance. MBL proteins also exist in plants, but they have low homology with bacterial MBLs and their functions are unknown. Our research found that tomato MBL is a key factor controlling starch content; this protein enhances SS activity and promotes amylopectin synthesis by interacting with SS3. The MBL protein involved in this invention contains 337 amino acids, and its amino acid sequence is shown in SEQ ID NO.1, as follows:

[0053]

[0054] This invention is the first to conduct an in-depth study on the function of plant MBL protein and clone the MBL protein encoding gene. The MBL protein encoding gene is 1014 bp in length, and its nucleotide sequence is shown in SEQ ID NO.2, as follows:

[0055]

[0056] To further elucidate the function of MBL, this invention utilized to obtain tomato MBL overexpression lines, RNAi-repressed expression lines, and Crisper-Cas9-mediated mutant lines. Analysis showed that the MBL overexpression lines exhibited higher starch synthase activity, greater starch accumulation, and higher biomass than the wild type; while the RNAi-repressed expression lines and mutant lines showed lower starch synthase activity, less starch accumulation, and significantly lower biomass than the wild type. Further experimental analysis indicated that this was due to the interaction between MBL and SS3, a member of the soluble starch synthase family, leading to enhanced SS3 activity. Therefore, this invention provides a novel starch synthesis regulatory pathway and identifies a new starch synthesis regulator.

[0057] Based on the above findings, the scope of protection of this invention also includes the function of DNA fragments homologous to the MBL gene, provided that the proteins they encode are functionally equivalent to the protein shown in SEQ ID NO. 1. The term "functionally equivalent to the protein shown in SEQ ID NO. 1" as used herein means that the protein encoded by the target DNA fragment is the same as or similar to the protein shown in SEQ ID NO. 1 in this invention in terms of biological function and physiological and biochemical characteristics. The typical biological function of the protein shown in SEQ ID NO. 1 is to increase amylase activity and increase amylopectin content.

[0058] These DNA fragments homologous to the MBL gene include alleles, homologous genes, mutant genes, and derived genes corresponding to the nucleotide sequence (SEQ ID NO.2) of this invention; the proteins they encode are similar to the protein shown in SEQ ID NO.1 of this invention, or there are substitutions, deletions, or insertions of one, several, or dozens of amino acids, all of which fall within the scope of this invention.

[0059] The present invention also provides a method for regulating the amount of amylopectin in plants by changing the expression level of the MBL gene in plants, which can be done by decreasing or increasing the expression level of MBL.

[0060] The MBL gene in this invention can be used in the field of plant genetic engineering, as its increase or decrease can lead to upregulation or downregulation of amylopectin content in tomatoes. Therefore, by increasing the expression level of the MBL gene in plants, the amylopectin level in crops such as tomatoes can be controlled, thereby improving crop yield and quality.

[0061] To enable those skilled in the art to better understand the technical solution of this application, the technical solution of this application will be described in detail below with reference to specific embodiments.

[0062] The test materials used in the embodiments of this invention, unless otherwise specified, are all conventional test materials in the art and can be purchased through commercial channels.

[0063] Example 1: Obtaining MBL mutant lines and overexpression lines

[0064] (1) Obtaining MBL mutant lines

[0065] MBL coding sequences were obtained through BLAST analysis. https: / / phytozome.jgi.doe.gov / pz / portal.html The database, MBL number is Solyc03g111700). The target sequence for Crisper Cas9, designed using https: / / crispr.cos.uni-heidelberg.de / , is as follows:

[0066] 5'-ATTCATCTTCACTTCCGTTG-3'; (SEQ ID NO.3)

[0067] 5'-TAGAGGGGAATAGCTGGTTG-3'. (SEQ ID NO.4)

[0068] The expression vector, pHSE401 (described in the journal article "A CRISPR / Cas9toolkit for multiplex genome editing in plants"), was constructed into the vector. This vector was then transformed into Agrobacterium strain LBA4404 via freeze-thaw conversion, and subsequently into tomato (Micro-TOM) using the Agrobacterium-mediated leaf disc method to obtain the MBL mutant line. The mutations in the MBL mutant lines can be categorized into two types: one involves adding AT after 147bp from ATG in the CDS, forming TAA at 223bp; the other involves adding T after 149bp from ATG in the CDS, forming TAA at 175bp. The "mbl-1 mutant line" and "mbl-2 mutant line" used in subsequent experiments in this application represent... Figure 1 The two mutation types shown are illustrated. The obtained strains were identified using DNA sequencing.

[0069] (2) Obtaining RNAi-repressed expression lines

[0070] A BLAST-specific sequence for the MBL coding region was obtained (database: https: / / phytozome.jgi.doe.gov / pz / portal.html, MBL ID: Solyc03g111700), and this sequence was amplified and ligated into the PC336 plant RNAi interference vector (provided by Shandong Agricultural University). This vector was transformed into Agrobacterium strain LBA4404 using a freeze-thaw method, and then into tomato (Micro-TOM) using the Agrobacterium-mediated leaf disc method to obtain an RNAi-inhibited expression line for MBL, named MBL-RNAi. The expression level of the obtained line was detected by qRT-PCR.

[0071] (3) Obtaining MBL overexpression lines

[0072] The full-length MBL coding region sequence (excluding the terminator) was constructed into the pSuper1300-GFP plant expression vector, transformed into Agrobacterium strain LBA4404 via freeze-thaw conversion, and then transformed into tomato (Micro-TOM) using the Agrobacterium-mediated leaf disc method. Overexpression lines of MBL were obtained and named MBL-OE1 and MBL-OE2. The obtained lines were identified by real-time quantitative PCR and PCR amplification.

[0073] The expression levels of MBL were detected by qRT-PCR in the overexpressing lines MBL-OE1 and MBL-OE2, the RNAi-inhibited expression line MBL-RNAi, and the wild-type line WT. (See attached table for details.) Figure 2 .

[0074] Example 2: Greenhouse Cultivation Experiment

[0075] The greenhouse was located at the State Key Laboratory of Wheat Breeding, Shandong Agricultural University. All plants were grown in the greenhouse. The tomato varieties (Micro-TOM) were named WT; the MBL overexpression lines MBL-OE1 and MBL-OE2 obtained in Example 1 were named MBL-1 and MBL-2; and the MBL RNAi line MBL-RNAi obtained in Example 1 was also included. Tomato seeds were used for cultivation. The seeds were homozygous, obtained from the aforementioned line samples through plant tissue culture and self-pollination. The seeds were sown in pots containing nutrient soil and vermiculite, and watered with the same nutrient solution. Each pot contained 8-10 tomato plants, and each sample from each line was divided into three replicates, for a total of 18 pots. The incubation period was 16 hours of daylight followed by 8 hours of darkness, at a room temperature of 25°C and an air humidity of 60-70%. One week after germination, robust seedlings were thinned to 3 plants per pot. For seedling experiments, two seedlings per pot were selected that had grown to 4-5 leaves in 2-3 weeks, and then continued to be cultivated. For flowering experiments, seedlings from the aforementioned seedling stage were selected and continued to grow for 4-5 weeks. All starch-related test samples were taken at the end of the day (when the lights were turned off).

[0076] See results Figure 3 and Figure 4 ,pass Figure 3 Phenotypic observation and Figure 4 The weighing results showed that the aboveground biomass of the MBL overexpression lines was higher than that of the wild type, while the MBL RNAi suppressed expression lines and their mutant lines were significantly lower than that of the wild type.

[0077] Starch staining analysis of leaves from tomato MBL overexpression lines, RNAi suppressed expression lines, and mutant lines was performed using Lugol's iodine solution:

[0078] The results are as follows Figure 5 As shown, the starch content in the leaves of MBL overexpression lines was significantly higher than that of wild type, while the starch content in the leaves of MBL mutant lines was lower than that of wild type.

[0079] The starch, amylopectin, amylose, and soluble sugar contents of leaves from tomato MBL overexpression lines and Crisper-Cas9-mediated mutant lines were determined. Starch content was measured using the DF-2-Y starch content test kit, amylose content test kit (ZDF-2-Y), and amylopectin test kit (ZHDF-2-Y) from Suzhou Keming Biotechnology Co., Ltd. Soluble sugar content was measured using the soluble sugar (SS) kit (G0501F) from Suzhou Gres Biotechnology Co., Ltd.

[0080] The results are as follows Figure 6 As shown, the amylopectin content in the leaves of MBL overexpression lines was higher than that of wild type, while the amylopectin content in MBL mutant lines was lower than that of wild type; however, the amylose content did not differ significantly among the above lines.

[0081] Transmission electron microscopy was used to observe the accumulation of starch grains in the chloroplasts of MBL-overexpressing, mutant, and wild-type leaves.

[0082] The results are as follows Figure 7 As shown, the accumulation of starch granules in the chloroplasts of leaves from MBL-overexpressing lines, mutant lines, and wild-type lines was observed (the white lumps in the chloroplasts are starch granules, indicated by the black arrows).

[0083] Observational analysis revealed that the starch accumulation (number of starch granules) in the chloroplasts of MBL overexpression lines was higher than that in the wild type, while the starch accumulation (number of starch granules) in the chloroplasts of MBL mutant lines was lower than that in the wild type.

[0084] The above experimental results indicate that MBL affects the accumulation level of starch in tomatoes. This suggests that tomato MBL is a key factor regulating starch synthesis in tomato chloroplasts.

[0085] Determination of total SS activity in each strain

[0086] The activities of soluble starch synthase in the MBL overexpression, mutant, and wild-type lines in Example 1 were determined by enzyme spectrometry. Non-denatured total protein was extracted from the tomato leaves using standard methods (1M Tris-HCl, pH 7.5, 0.5M EDTA, 5M NaCl, 1M MgCl2, 50% Glycerol, CPIC). The obtained protein samples were quantified using the Bradford method and then added in equal volumes to a Native-PAGE gel containing 0.3% glycogen for electrophoresis. After electrophoresis, the gel was removed and placed in a reaction solution (50M glycine, pH 9, 100mM [NH4]2SO4, 5mM β-mercaptoethanol, 5mM MgCl2, 0.25g / L BSA, 1mM ADP-Glc) overnight. The band differences were then observed after staining with Lugol's stain.

[0087] Total protein was extracted from leaves of MBL overexpression lines, RNAi-repressed expression lines, and mutant lines, and its enzyme profile was analyzed.

[0088] See results Figure 8 The total SS activity in MBL overexpression lines was higher than that in wild type, while the total SS activity in RNAi suppressed expression lines and mutant lines was lower than that in wild type.

[0089] Example 3: In vitro detection of the effect of tomato MBL protein on SS3 activity

[0090] The above MBL sequence was homologously recombinated using BamHI and SalI and constructed into the prokaryotic expression vector pET-30A (provided by Shandong Agricultural University). The above SS3 sequence was homologously recombinated using BamHI and SalI and constructed into the prokaryotic expression vector pMal-c2x-mbp (provided by Shandong Agricultural University). SS3 was obtained from SS3-MBP using overlap technology. ΔMBD2 -MBP was transferred into *Escherichia coli* strain BL21 using a heat shock method. The strain was induced to grow with 0.2 mM isopropyl thiogalactoside (IPTG) in a shaker at 37°C. Tomato MBL and SS3 proteins were induced. The bacterial culture was sonicated, and the supernatant proteins were separated and purified to obtain purified His, MBP, MBL-His, SS3-MBP, and SS3 proteins. ΔMBD2Five MBP proteins were analyzed and quantified using the Bradford method. Samples with the same protein content were subjected to zymography analysis using the same method. MBD2 is the binding site of MBL to SS3 protein and belongs to a domain of SS3 protein; the SS3 protein sequence number is NCBI Sequence ID: XP_004232219.1, and the SS3 CDS sequence number is NCBI Sequence ID: NM_001247694.1.

[0091] See results Figure 9 The five samples are MBL-His+SS3-MBP, MBL-His+SS3, etc. MBD2 -MBP, His and SS3-MBP, MBL-His and MBP, SS3 MBD2 -MBP; the darker the band, the higher the activity. It is evident that the activity of SS3 is significantly enhanced in the presence of MBL.

[0092] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. Application of MBL protein in any of the following i)-V): i) Increase the content of MBL protein to enhance the activity of tomato soluble starch synthase III; ii) Reduce the content of MBL protein to decrease the activity of tomato soluble starch synthase III; iii) Increase the content of MBL protein to promote the synthesis of amylopectin in tomatoes; iV) reduces the content of MBL protein to inhibit the synthesis of amylopectin in tomatoes; V) Enhance the activity of tomato soluble starch synthase III in vitro to promote amylopectin synthesis; The amino acid sequence of the MBL protein is shown in SEQ ID NO.

1.

2. Use of the gene encoding the MBL protein as described in claim 1 in any one of the following i)-Vi): i) Increase the expression level of the MBL gene to enhance the activity of tomato soluble starch synthase III; ii) Reduce the expression level of the MBL gene to decrease the activity of tomato soluble starch synthase III; iii) Increase the expression level of the MBL gene to promote the synthesis of amylopectin in tomatoes; iV) reduces the expression level of the MBL gene to inhibit the synthesis of amylopectin in tomatoes; V) Enhance the activity of tomato soluble starch synthase III in vitro to promote amylopectin synthesis; Vi) Increasing the expression level of the MBL gene in tomato breeding to increase amylopectin content.

3. The application according to claim 2, characterized in that, The gene is a nucleic acid molecule as shown in (1) or (2) below: (1) The nucleotide sequence is the nucleic acid molecule shown in SEQ ID NO.2; (2) Nucleic acid molecules other than (1) that encode the amino acid sequence shown in SEQ ID NO.

1.

4. The use of the expression cassette, recombinant expression vector, or recombinant bacteria containing the gene as described in claim 3 in any one of the following i)-iV): i) Increase the activity of soluble starch synthase III in tomatoes; ii) Promotes the synthesis of amylopectin in tomatoes; iii) To promote amylopectin synthesis by increasing the activity of tomato soluble starch synthase III in vitro; iV) Tomato plant breeding to increase amylopectin content.

5. A method for promoting the synthesis of amylopectin in plants, characterized in that, The method is as follows: To increase the content of plant MBL protein and promote the synthesis of plant amylopectin; the plant is tomato; The amino acid sequence of the MBL protein is shown in SEQ ID NO.

1.

6. A method for inhibiting the synthesis of amylopectin in plants, characterized in that, The method is as follows: The study aimed to reduce the content of plant MBL protein and inhibit the synthesis of plant amylopectin; the plant in question was tomato. The amino acid sequence of the MBL protein is shown in SEQ ID NO.

1.

7. A breeding method for plants with high amylopectin content, characterized in that, The breeding method is as follows: By increasing the content of MBL protein in the target plant, plants with a higher amylopectin content than the target plant were obtained; the plant was tomato. The amino acid sequence of the MBL protein is shown in SEQ ID NO.

1.

8. A method for breeding plants with low amylopectin content, characterized in that, The breeding method is as follows: By reducing the content of MBL protein in the target plant, plants with amylopectin content lower than that of the target plant were obtained; the target plant was tomato. The amino acid sequence of the MBL protein is shown in SEQ ID NO.1.

Images