MaSPL1 gene and application thereof in regulation and control of banana fruit after-ripening

Abstract

The invention discloses a MaSPL1 gene and application thereof in regulation and control of banana fruit after-ripening. The nucleotide sequence of the MaSPL1 gene is as shown in SEQ ID NO. 1. The invention relates to a key gene MaSPL1 for regulating and controlling banana fruit after-ripening in bananas. The invention provides a theoretical basis for genetic improvement of banana fruit after-ripening, has important breeding application value, and can be applied to cultivation of new storage and transportation-resistant banana varieties.

Description

Technical Field

[0001] The present invention relates to an SPL (squamosa promoter-binding protein-like) transcription factor MaSPL1 gene and applications thereof, in particular to a MaSPL1 gene isolated from banana and applications thereof in regulating the ripening of banana fruits and cultivating new banana varieties that are resistant to storage and transportation, belonging to the field of SPL transcription factor MaSPL1 genes and applications thereof. Background Art

[0002] Bananas, a typical climatic fruit, are susceptible to softening and rotting during transportation and storage at room temperature, resulting in significant postharvest losses, similar to other climatic fruits. Elucidating the regulatory factors and understanding their mechanisms for banana ripening is crucial for maintaining fruit quality, extending shelf life, and enhancing marketability.

[0003] SPL (squamosa promoter-binding protein-like) transcription factors are reportedly involved in plant development, including but not limited to root development, stomatal complex and epidermal hairs, floral organ development, asexual transition and flowering time, plant architecture, grain size and yield, and fruit development and ripening. Therefore, there is a pressing need to identify key SPL genes in bananas that regulate fruit ripening. By manipulating the expression of these genes, we can control postharvest ripening in bananas, promote the development of new ripening and preservation technologies, and ultimately achieve genetic improvement of bananas. Summary of the Invention

[0004] One of the objectives of the present invention is to isolate the SPL (squamosa promoter-binding protein-like) transcription factor MaSPL1 gene from banana;

[0005] The second purpose of the present invention is to apply the identified key gene (SPL transcription factor MaS PL1) that regulates banana fruit ripening and other aspects to the genetic improvement of banana postharvest ripening or banana breeding.

[0006] To achieve the above objectives, the technical solutions adopted by the present invention include:

[0007] The present invention first isolates the MaSPL1 gene from banana, and the polynucleotide sequence of its CDS is shown in (a), (b), (c), (d) or (e):

[0008] (a) the nucleotide sequence shown in SEQ ID No. 1;

[0009] (b) a nucleotide sequence encoding the amino acid shown in SEQ ID No. 2;

[0010] (c) a nucleotide sequence that can hybridize with the complementary sequence of the polynucleotide of SEQ ID NO. 1 under stringent hybridization conditions;

[0011] (d) a nucleotide sequence having at least 90% homology with the nucleotide sequence shown in SEQ ID No. 1;

[0012] (e) Nucleotide sequence variants in which one or more bases are deleted, substituted or inserted based on the polynucleotide shown in SEQ ID NO.1.

[0013] In addition, those skilled in the art can optimize the polynucleotide shown in SEQ ID No. 1 to enhance expression efficiency in plants. For example, the polynucleotide can be synthesized using the preferred codons of the target plant to enhance expression efficiency in the target plant.

[0014] The present invention further provides a protein encoded by the MaSPL1 gene, the amino acid sequence of which is shown in (a) or (b);

[0015] (a) the amino acid sequence shown in SEQ ID No. 2;

[0016] (b) Protein variants derived from the amino acid sequence shown in SEQ ID No. 2 by substitution, deletion and / or insertion of one or more amino acid residues.

[0017] The protein variants described herein can be generated by genetic polymorphisms or human manipulation, as is generally known in the art. For example, amino acid sequence variants or fragments can be prepared by mutation of DNA, as is known in the art for mutagenesis or polynucleotide modification. Conservative substitutions involve replacing one amino acid residue with another having similar properties.

[0018] "Variant" means a substantially similar sequence. For polynucleotides, variants include deletions, insertions, and / or substitutions of one or more nucleotides at one or more sites in a natural polynucleotide. For polynucleotides, conservative variants include those that do not change the encoded amino acid sequence due to the degeneracy of the genetic code. Naturally occurring variants such as these can be identified by existing molecular biology techniques. Variant polynucleotides also include polynucleotides of synthetic origin, such as polynucleotide variants that still encode the amino acid sequence shown in SEQ ID No. 2 obtained by site-directed mutagenesis or by recombinant methods (e.g., DNA shuffling). Those skilled in the art can screen or evaluate the function or activity of the protein encoded by the variant polynucleotide by the following molecular biotechnology means: DNA binding activity, interaction between proteins, activation of gene expression in transient studies, or the effect of expression in transgenic plants, etc.

[0019] The present invention also provides a recombinant plant expression vector containing the MaSPL1 gene and a host cell containing the recombinant plant expression vector.

[0020] The present invention further constructs a proUbi::MaSPL1 vector driven by the Ubi promoter; transforms bananas through an Agrobacterium-mediated method; and the transgenic strains exhibit an early ripening phenotype of banana fruits, indicating that MaSPL1 plays an important role in promoting the ripening of banana fruits.

[0021] The nucleotide sequence of the MaSPL1 gene of the present invention is shown in SEQ ID No. 1, and the amino acid sequence of the protein encoded by the MaSPL1 gene is shown in SEQ ID No. 2.

[0022] Therefore, the present invention provides the use of the MaSPL1 gene in promoting the ripening of banana fruits.

[0023] Preferably, the overexpression of MaSPL1 gene is used to promote the ripening of banana fruit.

[0024] In summary, the present invention identified a key gene MaSPL1 in banana that regulates the ripening of banana fruit. The present invention uses the Ma SPL1 gene to regulate the ripening of banana fruit and further cultivate new banana varieties.

[0025] Therefore, the present invention provides a method for promoting banana fruit ripening: constructing a plant recombinant expression vector containing the MaSPL1 gene; transforming the plant recombinant expression vector into bananas to overexpress the MaSPL1 gene in the bananas, thereby promoting banana fruit ripening; or regulating banana fruit ripening includes delaying banana fruit ripening.

[0026] Accordingly, the present invention also provides a method for delaying the ripening of banana fruit: constructing a knockout vector or a gene editing vector of the MaSPL1 gene; transforming the constructed gene knockout vector or gene editing vector into a banana plant to knock out or mutate the MaSPL1 gene in the banana.

[0027] Among them, the gene editing vector of the MaSPL1 gene or the gene knockout vector of the MaSPL1 gene can be obtained according to conventional construction methods in the art.

[0028] The gene editing CRISPR / Cas9 transgenic technology was used to design two target sites of the MaSPL1 gene in banana for gene editing.

[0029] SnapGene Viewer software and homologous sequence alignment were used to design and screen the specific target sequence of MaSPL1 gene (design sgRNA). In order to ensure the efficiency of gene editing, two optimal target sequences were selected for each gene: these target sequences were introduced into the sgRNA expression cassette. The CRISPR / Cas9 gene editing vector was constructed using a PCR Cloning Kit and cloned into the pUbi::MaCas9-Nos vector. After verification by PCR sequencing, it was used for subsequent genetic transformation. Subsequently, the constructed MaSPL1 CRISPR / Cas9 gene editing vector was transformed into banana plants via Agrobacterium-mediated transfection.

[0030] For reference, the present invention provides a method for producing a plant recombinant expression vector containing the MaSPL1 gene, comprising operably linking the MaSPL1 gene to expression regulatory elements to obtain a plant recombinant expression vector. The plant recombinant expression vector may be composed of a 5' non-coding region, the MaSPL1 gene, and a 3' non-coding region. The 5' non-coding region may include a promoter sequence, an enhancer sequence, and / or a translation enhancing sequence. The promoter may be a constitutive promoter, an inducible promoter, or a tissue- or organ-specific promoter. The 3' non-coding region may contain a terminator sequence, an mRNA cleavage sequence, or the like. Suitable terminator sequences can be obtained from the Ti-plasmid of Agrobacterium tumefaciens, such as the octopine synthase and nopaline synthase terminator regions.

[0031] The recombinant plant expression vector may also contain a selectable marker gene for selecting transformed cells or tissues. Such marker genes include genes encoding antibiotic resistance and genes conferring resistance to herbicidal compounds. Furthermore, such marker genes also include phenotypic markers, such as β-galactosidase and fluorescent protein.

[0032] The transformation protocols described herein and the methods for introducing the polynucleotides or polypeptides into plants can vary depending on the type of plant (monocot or dicot) or plant cell being transformed. Suitable methods for introducing the polynucleotides into plant cells include microinjection, electroporation, Agrobacterium-mediated transformation, direct gene transfer, and high-velocity ballistic bombardment. Transformed cells can be regenerated into stably transformed plants using conventional methods (McCormick et al. Plant Cell Reports. 1986. 5:81-84).

[0033] The target plant includes but is not limited to: a monocotyledonous plant or a dicotyledonous plant. More preferably, the target plant is banana.

[0034] The invention discloses a key gene MaSPL1 in banana that regulates the ripening of banana fruit. The invention provides a theoretical basis for the genetic improvement of banana fruit ripening and has significant breeding application value, and can be used to cultivate new banana varieties that are resistant to storage and transportation.

[0035] Definitions of terms used in this invention

[0036] Unless defined otherwise, 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 invention belongs.

[0037] The term "polynucleotide" or "nucleotide" means deoxyribonucleotides, deoxyribonucleosides, ribonucleosides or ribonucleotides and polymers thereof in single or double stranded form. Unless specifically limited, the term encompasses nucleic acids containing known analogs of natural nucleotides that have binding properties similar to the reference nucleic acid and are metabolized in a manner similar to naturally occurring nucleotides. Unless otherwise specifically limited, the term also means oligonucleotide analogs, including PNA (peptide nucleic acid), DNA analogs used in antisense technology (phosphorothioate, phosphoramidate, etc.). Unless otherwise specified, a specific nucleic acid sequence also implicitly encompasses conservatively modified variants thereof (including but not limited to degenerate codon substitutions) and complementary sequences as well as explicitly specified sequences. In particular, degenerate codon substitutions can be achieved by generating sequences in which position 3 of one or more selected (or all) codons is substituted with mixed-base and / or deoxyinosine residues (Batzer et al., Nucleic Acid Res. 19:5081 (1991); Ohtsuka et al., J. Biol. Chem. 260:2605-2608 (1985); and Cassol et al., (1992); Rossolini et al., Mol Cell. Probes 8:91-98 (1994)).

[0038] The "stringent hybridization conditions" described in the present invention refer to conditions of low ionic strength and high temperature as known in the art. Generally, under stringent conditions, the detectable degree of hybridization of the probe to its target sequence is higher than the detectable degree of hybridization to other sequences (for example, at least 2 times greater than background). Stringent hybridization conditions are sequence-dependent and will be different under different environmental conditions, with longer sequences specifically hybridizing at higher temperatures. By controlling the stringency of hybridization or washing conditions, target sequences that are 100% complementary to the probe can be identified. For detailed guidance on nucleic acid hybridization, reference can be made to the relevant literature (Tijssen, Techniques in Biochemistry and Molecular Biology-Hybridization with Nucleic Probes," Overview of principles of hybridization and the strategy of nucleic acid assays. 1993). More specifically, the stringent conditions are generally selected to be lower than the thermal melting point (Tf) of the specific sequence at a specified ionic strength pH. m ) about 5-10℃. m The temperature at which 50% of the probes complementary to the target hybridize to the target sequence in equilibrium (under specified ionic strength, pH, and nucleic acid concentration) (because the target sequence is present in excess, the target sequence is present at T m Stringent conditions may be those in which the salt concentration is less than about 1.0 M sodium ion concentration, typically about 0.01 to 1.0 M sodium ion concentration (or other salts), at pH 7.0 to 8.3, and the temperature is at least about 30°C for short probes (including but not limited to, 10 to 50 nucleotides) and at least about 60°C for long probes (including but not limited to, greater than 50 nucleotides). Stringent conditions may also be achieved by the addition of destabilizing agents such as formamide. For selective or specific hybridization, a positive signal may be at least twice the background hybridization, optionally 10 times the background hybridization. Exemplary stringent hybridization conditions may be as follows: 50% formamide, 5× SSC, and 1% SDS, incubation at 42°C; or 5× SSC, 1% SDS, incubation at 65°C, wash in 0.2× SSC, and wash in 0.1% SDS at 65°C. The washing may be performed for 5, 15, 30, 60, 120 minutes or longer.

[0039] The "multiple" mentioned in the present invention generally means 2-8, preferably 2-4, depending on the position of the amino acid residues or the type of amino acids in the three-dimensional structure of the transcription factor; the "replacement" refers to the replacement of one or more amino acid residues with different amino acid residues; the "deletion" refers to a reduction in the number of amino acid residues, that is, the lack of one or more amino acid residues; the "insertion" refers to a change in the amino acid residue sequence, and the change results in the addition of one or more amino acid residues relative to the natural molecule.

[0040] The terms "polypeptide," "peptide," and "protein" are used interchangeably herein to refer to a polymer of amino acid residues. That is, a description directed to a polypeptide applies equally to describing a peptide and describing a protein, and vice versa. The terms apply to naturally occurring amino acid polymers as well as amino acid polymers in which one or more amino acid residues is a non-naturally encoded amino acid. As used herein, the terms encompass amino acid chains of any length, including full-length proteins (i.e., antigens), in which the amino acid residues are linked via covalent peptide bonds.

[0041] The term "recombinant host cell strain" or "host cell" refers to a cell comprising a polynucleotide of the present invention, regardless of the method used for insertion to produce the recombinant host cell, such as direct uptake, transduction, f-mating, or other methods known in the art. The exogenous polynucleotide may be maintained as a non-integrating vector, such as a plasmid, or may be integrated into the host genome. The host cell may be a prokaryotic cell or a eukaryotic cell, and the host cell may also be a monocotyledonous or dicotyledonous plant cell.

[0042] The term "operably linked" refers to a functional connection between two or more elements. Operably linked elements may be contiguous or non-contiguous.

[0043] The term "plant recombinant expression vector" refers to one or more DNA vectors used to achieve plant transformation; these vectors are often referred to in the art as binary vectors. Binary vectors, along with vectors with helper plasmids, are most commonly used for Agrobacterium-mediated transformation. Binary vectors typically include the cis-acting sequences required for T-DNA transfer, a selectable marker engineered for expression in plant cells, and the heterologous DNA sequence to be transcribed.

[0044] The term "transformation" refers to a process by which a heterologous DNA sequence is introduced into a host cell or organism.

[0045] The term "expression" refers to the transcription and / or translation of an endogenous gene or a transgene in a plant cell.

[0046] Detailed description of the overall technical solution of the present invention

[0047] This invention utilizes the Ubi promoter-driven proUbi::MaSPL1-GFP vector for Agrobacterium-mediated transformation of bananas. The resulting overexpression transgenic strain exhibits a phenotype that promotes fruit ripening. Therefore, this invention provides a theoretical basis for genetically improving banana fruit ripening and has significant breeding application value, potentially enabling the development of new banana varieties that are durable in storage and transportation. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 Effects of MaSPL1 overexpression on banana fruit ripening: (A) RT-qPCR analysis of MaSPL1 expression in MaSPL1-overexpressing lines. (B) MaSPL1 overexpression leads to earlier ripening of banana fruit compared to the control. Scale bar, 5 cm. (C) MaSPL1 overexpression leads to faster yellowing of banana fruit compared to the control. Hue angles of MaSPL1-overexpressing and control banana fruits and (D) MaSPL1 overexpression leads to a reduced firmness phenotype in banana fruit compared to the control. Data are presented as mean ± SE of three or four biological replicates, using t-tests (*P < 0.05, ***P < 0.001). DETAILED DESCRIPTION

[0049] The present invention will be further described below with reference to specific embodiments, and the advantages and features of the present invention will become clearer as the description proceeds. However, these embodiments are merely exemplary and do not limit the scope of the present invention in any way. It should be understood by those skilled in the art that the details and forms of the present invention may be modified or replaced without departing from the spirit and scope of the present invention, and such modifications and replacements fall within the scope of protection of the present invention.

[0050] Example 1: Construction and phenotypic observation of banana MaSPL1 overexpression

[0051] (1) In this experiment, a Ubi promoter-driven proUbi::MaSPL1-GFP vector was constructed and transformed into banana via Agrobacterium-mediated method.

[0052] The specific construction method is as follows:

[0053] The pCAMBIA1300-Ubi-MCS-GFP binary vector (nucleotide sequence shown in SEQ ID NO. 3) containing a green fluorescent protein (GFP) reporter system controlled by the ubiquitin (Ubi) promoter was used. The MaSPL1 gene (shown in SEQ ID NO. 1, encoding the amino acid sequence of the protein shown in SEQ ID NO. 2) was inserted into the linearized pCAMBIA1300-Ubi-MCS-GFP vector via a directional cloning strategy using SacI / KpnI double digestion and the In-Fusion HD Cloning Kit (Clontech) to achieve seamless ligation of the target fragment based on homologous recombination.

[0054] The amplification primer sequences for the MaSPL1 gene are as follows:

[0055] OE-MaSPL1-proUbi-F: GTTTGGTGTTACTTCTGCAGGAGCTC ATGGAGCACCGGAGG CCGTCG;

[0056] OE-MaSPL1-proUbi-R: TCGACTCTAGAGGATCCCCGGGTACC TCTGATCTGGAAGTG CTTGTAC.

[0057] Transform pUbi::MaSPL1 into Agrobacterium tumefaciens EHA105 to obtain Agrobacterium tumefaciens carrying pUbi::MaSPL1. Inoculate this Agrobacterium EHA105 into LB liquid medium and incubate at 200 rpm for 48 hours to obtain a bacterial suspension. Load 0.3 ml of the bacterial suspension into a needle-free syringe (e.g., the Injex-30™ needle-free injection system). Inject 2-3 injections per banana onto a relatively flat surface. Seal the injection site with scotch tape. One day after injection, immerse the transformed fruit in an ethephon solution (200 mg / L) for 1 minute and incubate at 22°C for 6 days. Agrobacterium EHA105 transformed with the empty vector pCA MBIA1300-Ubi-MCS-GFP served as a control (Emply Vector). Thus, a MaSPL1 overexpressing strain (pUbi::MaSPL1) was obtained by injecting Agrobacterium EHA105 carrying pUbi::MaSPL1 and a blank control (Empty Vector) was obtained by injecting Agrobacterium EHA105 carrying the empty vector pCAMBIA1300-Ubi-MCS-GFP.

[0058] (2) RT-qPCR analysis of the expression level of MaSPL1 in MaSPL1 overexpressing strains. The specific method is as follows:

[0059] Banana samples were collected, frozen with liquid nitrogen, and stored at -80°C. Total RNA was extracted using the SteadyPure Plant RNA Extraction Kit (Hunan Aikerui Bioengineering Co., Ltd.). ⅢqPCR first-strand cDNA synthesis ready-to-use premix ( 1-2 μg of total RNA was reverse transcribed using Ⅲ1st Strand cDNA Synthes is SuperMix, Yisheng Biotechnology (Shanghai) Co., Ltd. A premixed solution for real-time quantitative PCR amplification (Hieff Real-time quantitative polymerase chain reaction (RT-qPCR) was performed using SYBR Green Master Mix (Shanghai Yisheng Biotechnology Co., Ltd.) and a LightCycler 480 system (Roche), with MaCAC as the reference gene. The primer sequences used are as follows:

[0060] MaCAC-qRT-F:GCTTTGATGTCGACAGAACTTT;

[0061] MaCAC-qRT-R:CGAGAAATCAGCACGTACTTTT;

[0062] MaSPL1-qRT-F: AGAGCACGAAGGAGAAGACGA;

[0063] MaSPL1-qRT-R:TGAGGTCAGCGGTGCAATTC;

[0064] The results are as follows Figure 1 As shown in A, the expression level of MaSPL1 in MaSPL1-overexpressing bananas was significantly higher than that in bananas transformed with an empty vector.

[0065] (3) The effect of MaSPL1 gene overexpression on banana fruit ripening.

[0066] The firmness and color of banana fruits were measured using a texture analyzer (BosinTech) and a color analyzer (Konica Minolta). Figure 1 ).

[0067] The banana pulp was punctured using a texture analyzer (BosinTech). The maximum stress during the puncture process was the firmness of the fruit. The results showed that the firmness of banana fruits in the overexpressing transgenic line (pUbi::MaSPL1) was significantly reduced after one day compared to the control with the empty vector ( Figure 1 D).

[0068] The banana skin was measured using a color analyzer (Konica Minolta). This experiment used color (h value) to analyze the color changes of the banana's appearance, ranging from 80° to 180°. The closer to 180°, the greener the banana's appearance, while the closer to 80°, the yellower the banana's appearance. The results showed that compared to the control, the degree of yellowing of banana fruit in the overexpressing transgenic line (pUbi::MaSPL1) increased significantly after one day ( Figure 1 B and C). Based on analysis of banana fruit firmness and color, the results indicate that overexpression of MaSPL1 can promote banana fruit ripening.

[0069] The nucleotide sequence of MaSPL1 gene is SEQ ID NO.1

[0070] ATGGAGCACCGGAGGCCGTCGACCGAGCCCTCAAGGAAGAGCACGAAGGAGAAGACGAGGAAGGACACGGCGGTCGCCACGGGCAACGACGAGGACGAGGAGGAGCAGGAGGAAGCCGCGGCGGAGGCGGACAAGAAGCGAAGGCCGCCCTCCTCCTCCTCCTCAGCTGCTCGCAGAGGAGTGAGCGGCGGCGGCGGCGGAGGAGGAGCGCCGCAACCCTGCTGTCAGGCCGAGAATTGCACCGCTGACCTCACCGAGGCGAAGCGCTACCACCGCCGGCACAAGGTCTGCGAGGCCCACTCCAAGGCCGCCGTCGTCATGGTCGCCGGCTTCCGCCAGAGGTTCTGCCAGCAATGCAGCAGGTTCCACGAGCTAGCCGAGTTCGACGACTCCAAGCGGAGCTGCCGCCGCCGCCTGGCCGGCCACAACGAGCGGCGCCGGAAGAGCTCCTCCGACGCGCAGGCCGGCGAAGGCTCGAACCGGTGCAGGCAGGCAGATCAAGACGGGAGGATGCAGATAAGCCTCCCCGGGAAGCCCACGTACAAGCACTTCCAGATCAGATGA

[0071] Amino acid sequence of MaSPL1 protein SEQ ID NO.2

[0072] MEHRRPSTEPSRKSTKEKTRKDTAVATGNDEDEEEQEEAAAEADKKRRPPSSSSSAARRGVSGGGGGGGAPQPCCQ AENCTADLTEAKRYHRRHKVCEAHSKAAVVMVAGFRQRFCQQCSRFHELAEFDDSKRSCRRRLAGHNERRRKSSSDAQAG EGSNRCRQADQDGRMQISLPGKPTYKHFQIR。

[0073] pCAMBIA1300 + Ubi + MCS + GFP SEQ ID NO.3

[0074]

Claims

1. The MaSPL1 gene, whose nucleotide sequence is shown in (a), (b), (c), (d), or (e): (a) the nucleotide sequence shown in SEQ ID No. 1; (b) a nucleotide sequence encoding the amino acid shown in SEQ ID No. 2; (c) a nucleotide sequence that can hybridize with the complementary sequence of the polynucleotide of SEQ ID NO. 1 under stringent hybridization conditions; (d) a nucleotide sequence having at least 90% homology with the nucleotide sequence shown in SEQ ID No. 1; (e) Nucleotide sequence variants in which one or more bases are deleted, substituted or inserted based on the polynucleotide shown in SEQ ID NO.

1.

2. MaSPL1 protein, the amino acid sequence of which is shown in (a) or (b); (a) the amino acid sequence shown in SEQ ID No. 2; (b) Protein variants derived from the amino acid sequence shown in SEQ ID No. 2 by substitution, deletion and / or insertion of one or more amino acid residues.

3. A recombinant plant expression vector containing the MaSPL1 gene according to claim 1.

4. A host cell containing the recombinant plant expression vector according to claim 3.

5. Use of the MaSPL1 gene according to claim 1 in regulating the ripening of plant fruits.

6. The use according to claim 5, characterized in that The plant is banana.

7. The use according to claim 6, characterized in that The invention relates to the application of overexpressing MaSPL1 gene in promoting the ripening of banana fruit.

8. The use according to claim 6, characterized in that It is the application of knocking out the MaSPL1 gene in delaying the ripening of banana fruit.

9. A method for promoting the ripening of banana fruits, characterized in that: A plant recombinant expression vector containing the MaSPL1 gene according to claim 1 is constructed; the plant recombinant expression vector is transformed into bananas, the MaSPL1 gene is overexpressed in the bananas, and the ripening of banana fruits is promoted.

10. A method for delaying the ripening of banana fruits, characterized in that: Constructing the knockout vector or gene editing vector of the MaSPL1 gene according to claim 1; transforming the constructed gene knockout vector or gene editing vector into a banana plant to knock out or mutate the MaSPL1 gene in the banana.

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