Application of SbGA20ox3 gene in regulating sorghum variety improvement and breeding

Abstract

This invention discloses the application of the SbGA20ox3 gene in regulating sorghum variety improvement and breeding, belonging to the field of biotechnology. This invention involves targeted editing of the SbGA20ox3 gene in sorghum, resulting in a significant dwarfing phenotype with a dwarfing efficiency of 37.01%–50.67%. Simultaneously, it reduces the number of internodes, shortens internode length, reduces the length of the main spikelet stalk, and increases stem diameter, while increasing cellulose and lignin content, significantly enhancing lodging resistance. Furthermore, it simultaneously optimizes the tannin and amylopectin content of sorghum grains, improving quality traits. The gene-edited sorghum obtained by this invention does not contain transgenic elements, and other agronomic traits are unaffected. It can target and improve sorghum plant architecture, stress resistance, and quality, providing key gene targets and practical technical systems for molecular breeding of dwarfed, lodging-resistant, high-quality, and high-yield sorghum, and has significant agricultural application value.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, specifically to the application of the SbGA20ox3 gene in regulating sorghum variety improvement and breeding. Background Technology

[0002] Sorghum is an important global food, feed, and industrial raw material crop. Dense planting is a key way to achieve high sorghum yields, but it also easily leads to a significant increase in lodging rates in the field, seriously affecting sorghum yields and the efficiency of mechanized harvesting. Dwarf sorghum plants are short and compact, with strong lodging resistance and high light energy utilization, making them ideal varieties for dense, high-yield cultivation. Therefore, screening and breeding dwarf, lodging-resistant sorghum varieties is one of the core directions of sorghum breeding.

[0003] Meanwhile, the quality traits of sorghum grains, such as tannins and amylopectin, directly determine their processing and utilization value. Improving quality while maintaining high yield is a crucial goal of sorghum breeding. Currently, multinational seed companies have established a market advantage through transgenic sorghum breeding technology. However, sorghum varieties still have shortcomings in terms of lodging resistance, yield, and synergistic improvement of quality. There is an urgent need to create new sorghum germplasm with dwarfing, lodging resistance, and high quality through molecular breeding technology.

[0004] Gibberellins (GAs) are key hormones regulating plant height, and GA20 oxidase (GA20ox) is a key rate-limiting enzyme in the gibberellin biosynthesis pathway. The gibberellin synthesis process regulated by its encoding gene directly affects morphogenesis processes such as internode elongation and stem development. SbGA20ox3, as one of the encoding genes for GA20 oxidase, has been preliminarily studied in its role in sorghum plant height regulation. However, the function of this gene in regulating sorghum lodging resistance and grain quality, as well as its molecular breeding applications in achieving synergistic improvement of sorghum plant height, lodging resistance, and quality through gene knockout, remain unclear.

[0005] Gene editing technology is a core tool in crop molecular breeding, and the CRISPR / Cas series systems are widely used due to their simplicity and high editing efficiency. KingCas12, a novel gene-editing enzyme obtained through MAD7 nuclease mutation, can efficiently recognize T-rich PAM sequences in plants, and its smaller molecular weight and higher editing specificity provide a high-quality tool for precise editing of the sorghum genome. Currently, there are no reports on using the CRISPR-KingCas12 system to edit the SbGA20ox3 gene to achieve synergistic improvement in sorghum dwarfing, lodging resistance, and quality.

[0006] Therefore, exploring the regulatory function of the SbGA20ox3 gene and using gene editing technology to directionally edit this gene to create new dwarf, lodging-resistant, and high-quality sorghum germplasm has important theoretical value and application prospects for promoting the development of molecular breeding of sorghum and ensuring the security of my country's sorghum industry. Summary of the Invention

[0007] To address the problems existing in the prior art, this invention provides the application of the SbGA20ox3 gene in regulating sorghum variety improvement and breeding, clarifying that editing this gene can achieve a synergistic effect of reducing sorghum plant height, enhancing lodging resistance, and improving grain quality, and also provides a method for breeding such sorghum varieties.

[0008] The technical solution of the present invention: First aspect of the invention An application of the SbGA20ox3 gene in regulating sorghum variety improvement and breeding; the nucleotide sequence of the SbGA20ox3 gene is shown in SEQ ID NO.1; the protein encoded by the SbGA20ox3 gene has the amino acid sequence shown in SEQ ID NO.2.

[0009] The regulation refers to the modification of the SbGA20ox3 gene by gene editing technology to remove or reduce its function, so as to achieve the application in sorghum variety improvement and breeding; the variety improvement refers to reducing sorghum plant height, improving sorghum lodging resistance, and increasing tannin and amylopectin content in sorghum grains; the modification of removing or reducing function includes site-directed mutagenesis or base knockout.

[0010] The application is performed by at least one of the following two gene editing methods: (1) inserting 1 bp at the 994 bp position of the sorghum SbGA20ox3 gene, and (2) causing a 1 bp deletion mutation at the 139 bp position of the SbGA20ox3 gene. Both methods result in the loss of function of the SbGA20ox3 gene.

[0011] Application of knocking out the SbGA20ox3 gene in sorghum, wherein the application is any one of the following: A1) Application in reducing sorghum plant height or in the preparation of products that reduce sorghum plant height; A2) Application in improving lodging resistance of sorghum or in the preparation of products that improve lodging resistance of sorghum; A3) Application in cultivating sorghum varieties with reduced plant height; A4) Application in cultivating sorghum varieties with improved lodging resistance; A5) Application in the cultivation of sorghum varieties with increased tannin and amylopectin content in grains; A6) Application in increasing the tannin and amylopectin content of sorghum grains; The nucleotide sequence of the SbGA20ox3 gene is 2703 bp in length from the start codon to the stop codon, containing 3 exons and 2 introns, as shown in SEQ ID No. 1. The 5' UTR is CACTCGCACATCTC, and the 3' UTR is CGAGCGAGCGAGCCGGGCCAAACAAACAAGGGGCAAAGGCCATCTCTTTCGCCGGGGCCCGCGCGCGGGGTTCGCCCACGTGCGCGCCCAGGTGGGCGCTGGCCGCGGGCAGGTGGCGGACATGTGGCCTGCGGGCCCCGCGCCGCCTTCCCATTTTTGGACGCTGCCGCGCATGCCGCATGCGTGCGTCGACGGCCCTACTACT The coding region of the SbGA20ox3 gene is 1241 bp, and the non-coding region is 1462 bp. The SbGA20ox3 gene encodes 413 amino acids, and the amino acid sequence is shown in SEQ ID No. 2.

[0012] The regulation refers to gene editing at position 139bp of the SbGA20ox3 gene to obtain mutant sorghum plants. This gene editing involves knocking out 1bp at position 139bp of the SbGA20ox3 gene. The nucleotide sequence of the mutant is shown in SEQ ID No. 3. The knockout of the base results in a frameshift mutation. The first 46 amino acids of the mutant are identical to those of the original protein, and a stop codon is generated after the 221st amino acid. The amino acid sequence of the mutant is shown in SEQ ID No. 4.

[0013] The amino acid sequence of the protein encoded by the SbGA20ox3 gene is SEQ ID No. 2: MVSQERQEPALPLPSNSSSAKRAAASMDASSPAPPLLLRAPTPSPSIDLPAAAGKAAAVFDLRREPKIPAPFLWPHEEARPTSAAELEVPVVDVGVLRNGDRAGLRRAAAQVASACATHGFFQVCGHGVDAALGRAALDGASDFFRLPLADKQRARRVPGTVSGYTSAHADRFASKLPWKETLSFGFHDGAASPVVVDYFTGTLGQ DFEPMGRVYQRYCEKMKELSLTIMELLELSLGVERGYYREFFEDSRSIMRCNYYPPCPEPERTLGTGPHCDPTALTILLQDDVGGLEVLVDGEWRPVRPVPGAMVINIGDTFMALSNGRYKSCLHRAVVNQRQERRSLAFFLCPREDRVVRPPASSATPRQYPDFTWADLMRFTQRHYRADTRTLDAFTRWLSHGPVPAQEAAAPCT The nucleotide sequence of the SbGA20ox3 mutant gene is SEQ ID No. 3: Amino acid sequence of the protein encoded by the SbGA20ox3 mutant gene SEQ ID No.4: MVSQERQEPALPLPSNSSSAKRAAASMDASSPAPPLLLRAPTPSPSIDLPAAAGKAAAVFDLRREPKIPAPFLWPHEEARPTSAAELEVPVVDVGVLRNGDRAGLRRAAAQVASACATHGFFQVCGHGVDAALGRAALDGASDFFRLPLADKQRARRVPGTVSGYTSAHADRFASKLPWKETLSFGFHDGAASPVVVDYFTGTLGQDFEPMGRVYQRYCEKMKELSLTIMELLELSLGVERGYYREFFEDSRSIMRCNYYPPCPEPERTLGTGPHCDPTGADHPPAGRRRRAGGAGGRRVAPRPARPRRHGHQHRRHLHGAVERAVQELPAPRGGEPAAGAAVAGLLPVPARGPGGAAAGQQRHAAAVPGLHLGRPHALHAAPLPRRHPHAGRLHPLALPRPSPSPGGGGSLHLASERAGPNKQGAKAISFAGARARGSPTCAPRWALAAGRWRTCGLRAPRRLPIFGRCRACRMRASTALLLLLLLLRLLVYIRKNTYIRIFYIYIYKQGGPPVTFSLFLSTTVF Nucleotide sequence of the SbGA20ox3 mutant gene SEQ ID No.5: 5’- The amino acid sequence of the protein encoded by the SbGA20ox3 mutant gene is SEQ ID No. 6: MVSQERQEPALPLPSNSSSAKRAAASMDASSPAPPLLLRAPTPSPSLTSPLPLARPRPCSTCGGSPRSRRHSCGRTRRARPRPRSWRFRWWTWACCAMATARGCGAPRRRWPRRARRTGSSRCAGTAWTRPWGAPRWTAPATSSGCRWPTSSAPGASPAPCPGTRARTPTGSRPSSPGRRPCPSASTTAPRRPSSWTTSPAPSAKISSQWGGCTRGTARR The reduction in sorghum plant height is specifically manifested in a dwarfing efficiency of 37.01%~50.67%, with dwarfing characteristics resulting from a decrease in the number of internodes, a shortening of internode length, and a reduction in the length of the main spikelet stalk. The improvement in lodging resistance is specifically manifested in an increase in stem diameter and an increase in cellulose and lignin content, resulting in shorter and stronger sorghum stems and a significantly enhanced lodging resistance potential. The quality improvement is specifically manifested in the simultaneous optimization of tannin and amylopectin content in the grains, thereby improving the processing and utilization value of sorghum.

[0014] In this regard, the present invention also provides an SbGA20ox3 mutant gene that regulates sorghum plant height, lodging resistance, tannin and amylopectin content in grains, the nucleotide sequence of the SbGA20ox3 mutant gene being shown in SEQ ID NO.3 and SEQ ID NO.5, and the amino acid sequence of the protein encoded by the SbGA20ox3 mutant gene being shown in SEQ ID NO.4 and SEQ ID NO.6.

[0015] This paper also provides an application of the aforementioned SbGA20ox3 mutant gene in regulating sorghum variety improvement and breeding. By using gene editing technology to perform site-directed mutations or loss-of-function modifications on the SbGA20ox3 gene in sorghum, the following applications are achieved: A1) Application in reducing the height of sorghum plants; A2) Application in improving lodging resistance of sorghum; A3) Application in the cultivation of sorghum varieties with reduced plant height and / or improved lodging resistance; A4) Application in the cultivation of sorghum varieties with increased tannin and amylopectin content in grains.

[0016] Second aspect of the invention A method for cultivating sorghum with reduced plant height and / or increased stem diameter includes: mutating the sorghum by reducing the content and / or activity of SbGA20ox3 protein in the sorghum to obtain sorghum with reduced plant height and / or increased stem diameter, wherein the amino acid sequence of the SbGA20ox3 protein is shown in SEQ ID No. 2.

[0017] Specifically, the method includes one or two of the following: 1) Deleting 1 bp from the 139 bp position of the sorghum SbGA20ox3 gene using knockout technology to obtain a dwarfing mutant AGHYZ02; the sequence of the mutant AGHYZ02 is shown in SEQ ID No. 5. 2) Inserting 1 bp from the 994 bp position of the sorghum SbGA20ox3 gene using mutagenesis technology to obtain a dwarfing mutant AGHYZ01; the nucleotide sequence of the mutant AGHYZ01 is shown in SEQ ID No. 3.

[0018] More specifically, the mutated amino acid contains 221 amino acids, with the first 46 amino acids being the same as those in SbGA20ox3.

[0019] The cultivated sorghum plants are gene-edited sorghum without transgenic elements, and their agronomic traits are unaffected except for plant height, lodging resistance, and grain quality.

[0020] Furthermore, a method for cultivating sorghum with reduced plant height and / or increased lodging resistance and / or improved quality includes the following steps: 1. Gene editing vector construction: Using the CRISPR-KingCas12 gene editing system, a single-target vector was designed and constructed. SbGA20ox3 Gene editing vector AGHYZ0101; the KingCas12 enzyme is obtained by mutating the MAD7 nuclease at the K169R and N589H amino acid sites, and can efficiently and specifically recognize the T-rich PAM sequence (YTTV) in plants. 2. Agrobacterium-mediated transformation: The constructed gene-editing vector was introduced into sorghum recipient material via Agrobacterium-mediated transformation to achieve gene editing in the sorghum genome. SbGA20ox3 Targeted gene editing; 3. Screening and identification of positive plants: Positive plants were selected through PCR and sequencing analysis. SbGA20ox3 Gene-knockout positive transgenic plants were used to breed gene-edited sorghum lines that do not contain transgenic elements and have stable traits through offspring selection; 4. Agronomic trait identification: The selected gene-edited sorghum lines were identified for plant height, stem morphology, lodging resistance and grain quality, and sorghum plants with reduced plant height and improved lodging resistance were obtained.

[0021] Third aspect of the invention SbGA20ox3 The application of genes or their encoded proteins in the screening / preparation of sorghum breeding materials, to SbGA20ox3 By targeting specific genes and using gene editing to knock out or insert gene mutants, the resulting mutants can serve as core materials for breeding dwarfing, lodging-resistant, high-quality, and high-yield sorghum varieties. The gene editing refers to gene mutation or gene knockout.

[0022] This invention utilizes the CRISPR-KingCas12 gene editing system to... SbGA20ox3 The gene was edited without introducing any foreign genes. The resulting gene-edited sorghum does not contain transgenic elements, and other agronomic traits are unaffected except for the target improved traits, which meets the requirements of agricultural production for varieties.

[0023] Beneficial effects 1. This invention is the first to clearly define SbGA20ox3 This gene can simultaneously regulate sorghum plant height, lodging resistance, and grain quality. Knocking out this gene can achieve synergistic improvement of these three factors, filling the gap in the functional research of this gene in sorghum lodging resistance and quality regulation, and providing a new theory for elucidating the molecular mechanisms of sorghum plant architecture, stress resistance formation, and quality regulation.

[0024] 2. This invention utilizes the CRISPR-KingCas12 gene editing system. SbGA20ox3 Precise gene knockout resulted in sorghum with a dwarfing efficiency of 37.01% to 50.67%, short and sturdy stems, significantly enhanced lodging resistance, and increased tannin and amylopectin content in the grains. This achieved the breeding goal of "dwarfing + lodging resistance + high quality" and provided excellent germplasm for high-density and high-yield sorghum cultivation.

[0025] 3. The breeding method used in this invention is precise and efficient, and the resulting gene-edited sorghum does not contain transgenic elements, thus avoiding the application restrictions of transgenic varieties. Furthermore, the KingCas12 enzyme provides a practical tool and technical system for molecular breeding of sorghum in my country.

[0026] 4. The new sorghum variety developed in this invention has strong lodging resistance and excellent quality. It is suitable for mechanized harvesting and high-density planting and high-yield cultivation, which can effectively improve the benefits of sorghum planting. It is of great significance to promoting the high-quality development of my country's sorghum industry and ensuring food security. Attached Figure Description

[0027] Figure 1 The image shows the vector pattern of AGHYZ0101.

[0028] Figure 2 The results are from PCR detection of exogenous gene insertion.

[0029] Figure 3 This is a comparison diagram of the nucleotide sequence of mutant AGHYZ01 and the recipient sorghum SbGA20ox3.

[0030] Figure 4 This is a comparison diagram of the amino acid sequences of the protein encoded by the mutant AGHYZ01 and the recipient sorghum SbGA20ox3 gene.

[0031] Figure 5 This is a comparison diagram of the nucleotide sequences of the mutant AGHYZ02 and the recipient sorghum SbGA20ox3.

[0032] Figure 6 This is a comparison diagram of the amino acid sequences of the protein encoded by the mutant AGHYZ02 and the recipient sorghum SbGA20ox3 gene.

[0033] Figure 7 This is a comparison of plant height phenotypes between conventional sorghum and gene-edited sorghum AGHYZ01.

[0034] Figure 8 The figure shows a comparison of plant height between conventional sorghum and gene-edited sorghum AGHYZ01 at different growth stages. In the figure, **P<0.01, ***P<0.005, and ****P<0.001 are considered.

[0035] Figure 9 This is a bar graph comparing the stem diameters of conventional sorghum and gene-edited sorghum AGHYZ01. P < 0.001 is shown in the figure.

[0036] Figure 10 This is a comparison of lodging resistance in the field between conventional sorghum and gene-edited sorghum AGHYZ01.

[0037] Figure 11 The bar chart shows the comparison of the starch and other nutrient content of conventional sorghum and gene-edited sorghum AGHYZ01. *P<0.05, **P<0.01 in the figure. Detailed Implementation

[0038] Example 1: Construction of gene editing vector 1. Preparation of KingCas12 enzyme: Using MAD7 nuclease as a template, the K169R and N589H amino acid sites were mutated by site-directed mutagenesis to obtain KingCas12 enzyme, which can efficiently and specifically recognize T-rich PAM sequences (YTTV) in plants.

[0039] 2. Target design and vector construction: Based on sorghum SbGA20ox3 The gene coding sequence was used to design a specific single-target gRNA, which was then linked to the KingCas12 coding sequence to construct the single-target gene editing vector AGHYZ0101. See details below. Figure 1 The vector was transferred into Agrobacterium competent cells to obtain Agrobacterium engineered bacteria containing the recombinant vector.

[0040] Example 2 Agrobacterium-mediated genetic transformation of sorghum Using sorghum as the recipient material, genetic transformation was carried out using Agrobacterium-mediated transformation. The specific steps included: 1. Select plump sorghum seeds, disinfect their surfaces, and germinate them under aseptic conditions to obtain explants; 2. Immerse the explants in an infection solution containing recombinant Agrobacterium engineered bacteria, infect at room temperature, then transfer to a co-culture medium and incubate in the dark at 25°C for 3-5 days; 3. The co-cultured explants were transferred to a selection medium, and after induction differentiation, redifferentiation, elongation, and rooting culture, resistant regenerated plants were obtained; 4. The Bar test strip method was used to preliminarily identify the T0 generation regenerated plants and screen for positive transgenic plants.

[0041] PCR test results as follows Figure 2 As shown in the figure, M represents the Marker; 1 represents the AGHYZ0101 plasmid. KingCas12 Gene PCR results; 2 represents AGHYZ01 E0 generation. KingCas12 Gene PCR results; 3 represents AGHYZ01 E1 generation. KingCas12 Gene PCR results; 4 represents AGHYZ01 E2 generation. KingCas12 Gene PCR results; 5 represents sorghum KingCas12 Gene PCR results; 6 represents the AGHYZ0101 plasmid. bar Gene PCR results; 7 represents AGHYZ01 E0 generation. bar Gene PCR results; 8 represents AGHYZ01E1 generation. bar Gene PCR results; 9 represents AGHYZ01 E2 generation. bar Gene PCR results; 10 represents sorghum bar Gene PCR results.

[0042] Example 3: Identification and Screening of Gene-Edited Sorghum Plants 1. Molecular identification: Genomic DNA was extracted from T0 generation positive plants, and the editing region of the SbGA20ox3 gene was amplified by PCR. The PCR products were sequenced, and homozygous mutant plants with SbGA20ox3 gene knockout were screened to obtain the results. 2. Offspring selection: The homozygous mutant plants were self-pollinated to obtain T1 and T2 generation lines. Lines without transgenic elements and with stable SbGA20ox3 gene knockout traits were screened by molecular detection and named AGHYZ01 and AGHYZ02.

[0043] See details Figures 3-6 Nucleotide sequence comparison diagram of mutants AGHYZ01 and AGHYZ02 with the SbGA20ox3 gene of the recipient sorghum, and amino acid sequence comparison diagram of the protein encoded by the SbGA20ox3 gene.

[0044] Example 4: Identification of agronomic traits of gene-edited sorghum AGHYZ01 Using wild-type sorghum as a control, the plant height, stem morphology, lodging resistance, and grain quality of the AGHYZ01 line were evaluated. Each trait was replicated three times. The results are as follows: 1. Plant height assessment: The dwarfing efficiency of the AGHYZ01 line reached 37.01%~50.67%. The reduction in plant height was mainly mediated by a decrease in the number of internodes, a shortening of internode length, and a reduction in the length of the main spikelet stalk; for details, see [link to relevant data]. Figure 7 and Figure 8 .

[0045] 2. Lodging resistance assessment: The stem diameter of the AGHYZ01 strain was significantly increased compared to the wild type, with an increase rate of 11.76%~18.87%; the stem cellulose and lignin content were also increased, with cellulose (dry weight) increasing by 11.62%~30.64% and lignin (dry weight) increasing by 4.08%~11.88%; the stems were short and stout, significantly enhancing lodging resistance in the field; for details, see [link to relevant documentation]. Figure 9 and Figure 10 .

[0046] 3. Grain quality assessment: The AGHYZ01 strain showed a simultaneous increase in tannin and amylopectin content compared to the wild type, with tannin content increasing by 27.52%–29.53% and amylopectin content increasing by 1.68%–2.87%, demonstrating significant improvement in quality traits; for details, see [link to product description]. Figure 11 .

[0047] 4. Other agronomic traits: The heading date, flowering date, and seed setting rate of the AGHYZ01 line were not significantly different from those of the wild type, indicating that knockout... SbGA20ox3 Genes only regulate the target trait and do not affect the normal growth and development of sorghum.

[0048] Example 5: Intermediate field trial of gene-edited sorghum AGHYZ01 and AGHYZ02 Gene-edited sorghum varieties AGHYZ01 and AGHYZ02 were subjected to field trials in Shandong, Guizhou, and Yunnan provinces to evaluate their lodging resistance, agronomical trait stability, and quality performance under natural conditions. The results showed that AGHYZ01 and AGHYZ02 exhibited stable dwarfing and lodging resistance phenotypes in different ecological zones, maintained high levels of tannin and amylopectin content in their grains, and demonstrated strong adaptability. They can be further demonstrated and promoted in the aforementioned regions.

[0049] Table 1. Statistical analysis of plant height traits in T3 generation SbGA20ox3 gene-edited sorghum AGHYZ01 and AGHYZ02 (n=30, - x ± s )

[0050] Note: Compared with conventional sorghum, ****: P<0.0001.

[0051] This invention is not limited to the specific textual description above. Various changes can be made to this invention within the scope outlined in the claims, and all such changes are within the scope of this invention.

Claims

1. An application of the SbGA20ox3 gene in regulating sorghum variety improvement and breeding; the nucleotide sequence of the SbGA20ox3 gene is shown in SEQ ID NO.1; the protein encoded by the SbGA20ox3 gene has the amino acid sequence shown in SEQ ID NO.

2.

2. The application as described in claim 1, characterized in that, The regulation refers to the use of gene editing technology to mutate or knock out bases in the SbGA20ox3 gene to achieve the application in sorghum variety improvement and breeding; the variety improvement refers to reducing sorghum plant height, improving sorghum lodging resistance, and increasing the tannin and amylopectin content of sorghum grains.

3. The application as described in claim 1 or 2, characterized in that, The gene editing involves inserting a 1 bp base at position 994 bp in the sorghum SbGA20ox3 gene, or performing a 1 bp deletion mutation at position 139 bp in the SbGA20ox3 gene.

4. Application of knocking out the SbGA20ox3 gene in sorghum, wherein the application is any one of the following: A1) Application in reducing sorghum plant height or in the preparation of products that reduce sorghum plant height; A2) Application in improving lodging resistance of sorghum or in the preparation of products that improve lodging resistance of sorghum; A3) Application in cultivating sorghum varieties with reduced plant height; A4) Application in cultivating sorghum varieties with improved lodging resistance; A5) Application in the cultivation of sorghum varieties with increased tannin and amylopectin content in grains; A6) Application in increasing the tannin and amylopectin content of sorghum grains; The nucleotide sequence of the SbGA20ox3 gene is shown in SEQ ID NO.1; The protein encoded by the SbGA20ox3 gene has the amino acid sequence as described in SEQ ID NO.

2.

5. A method for cultivating sorghum with reduced plant height and / or increased stem diameter, characterized in that, include: By using gene editing technology to perform site-directed mutations or knock out bases in the SbGA20ox3 gene in sorghum, sorghum with reduced plant height and / or increased stem diameter can be obtained. The amino acid sequence of the protein encoded by the SbGA20ox3 gene is shown in SEQ ID No.

2.

6. The method for cultivating sorghum with reduced plant height and / or increased stem diameter as described in claim 5, characterized in that, The method includes any one or two of the following: (1) inserting 1 bp at position 994 bp of the sorghum SbGA20ox3 gene through gene editing, wherein the nucleotide sequence of the SbGA20ox3 mutant gene is shown in SEQ ID No. 3, and the amino acid sequence of the protein encoded by the SbGA20ox3 mutant gene is shown in SEQ ID No. 4; (2) deleting 1 bp at position 139 bp of the sorghum SbGA20ox3 gene through gene editing, wherein the nucleotide sequence of the SbGA20ox3 mutant gene is shown in SEQ ID No. 5, and the amino acid sequence of the protein encoded by the SbGA20ox3 mutant gene is shown in SEQ ID No. 6; both editing methods result in dwarfing mutants; the gene editing is gene mutation or gene knockout.

7. A method for cultivating sorghum with reduced plant height and / or increased stem diameter as described in claim 5 or 6, characterized in that, The cultivated sorghum plants are gene-edited sorghum without transgenic elements, and their agronomic traits are unaffected except for plant height, lodging resistance, and grain quality.

8. The application of the SbGA20ox3 gene or its encoded protein in screening and / or preparing sorghum breeding materials, characterized in that, The application involves knocking out the SbGA20ox3 gene through gene editing to obtain a sorghum mutant, which is then used as a breeding variety for dwarfing, lodging-resistant, high-quality, and high-yield sorghum.

9. An SbGA20ox3 mutant gene, characterized in that, The mutant gene contains mutations that result in reduced or absent function of the encoded protein. The nucleotide sequence of the SbGA20ox3 mutant gene is shown in SEQ ID NO.3 and SEQ ID NO.5, and the amino acid sequence of the encoded protein of the SbGA20ox3 mutant gene is shown in SEQ ID NO.4 and SEQ ID NO.

6.

10. The application of the SbGA20ox3 mutant gene as described in claim 9 in regulating sorghum variety improvement and breeding, characterized in that, By reducing or eliminating the function of the SbGA20ox3 gene, the following applications can be achieved: A1) Application in reducing the height of sorghum plants; A2) Application in improving lodging resistance of sorghum; A3) Application in the cultivation of sorghum varieties with reduced plant height and / or improved lodging resistance; A4) Application in the cultivation of sorghum varieties with increased tannin and amylopectin content in grains.

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