Application of grape VvCHI4D gene in promotion of early ripening of berry fruits

Abstract

The invention discloses an application of a grape VvCHI4D gene in promoting berry fruits to mature in advance, which comprises the step of performing overexpression on the grape VvCHI4D gene in a plant to obtain a transgenic plant with advanced flowering period and mature period. The protein coded by the grape VvCHI4D gene is as shown in SEQ ID No. 2; the nucleotide sequence of the grape VvCHI4D gene is as shown in SEQ ID No.1 (sequence identifier number 1). According to the invention, the VvCHI4D gene of the grape is expressed in the tomato, so that the function of shortening the growth period of the fruit is achieved, and a theoretical support is provided for promoting the fruit maturation period of the grape and other crops by improving the expression of the VvCHI4D gene of the grape.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and particularly relates to an application of a grape VvCHI4D gene in promoting early ripening of berry fruits. Background Art

[0002] China is the world's largest producer and consumer of table grapes. In 2021, China's grape output value was about 400 billion US dollars (Song and Xu, 2023). Among them, the average economic benefit per mu of table grapes in the southern region can reach more than 20,000 yuan, which is much higher than other fruit trees. Grape cultivation has gradually become a pillar industry of township agricultural production. The time when table grapes are put on the market is directly related to the level of economic benefits. Taking the short-growing grape variety "Summer Black" as an example, the sales price in June was 15-30 yuan / jin, and the sales price dropped to 5-10 yuan / jin in August, which is consistent with the survey results of Mu Weisong et al. (2019). Therefore, the early maturity of grapes is an important means to avoid grape production problems and market risks in the coastal areas of southern China. Moreover, the early maturity of grapes can effectively reduce the use of chemical agents and the number of times they are used, and the ecological benefits are significant. Therefore, studying the formation mechanism of early-maturing grapes can lay a good foundation for assisting the breeding of early-maturing varieties.

[0003] Chitinase (EC 3.2.1.14) is a glycosyl hydrolase that catalyzes the degradation of chitin, a β-1,4-linked polymer of N-acetylglucosamine (GlcNAc) found in fungal cell walls and other natural sources (Watanabe et al., 1999). Chitinases in plants are closely related to pathogen infection and fruit development (Samac et al., 1990; Collinge et al., 1993). Previous studies have found that in grapes, type IV chitinase activity increases sharply during berry ripening (Robinson et al., 1997). Transcriptome analysis and qRT-PCR results of 'Xiahei' and 'Xiahei' bud mutation material 'Tiangong Moyu' found that the expression level of VvCHI4D in 'Tiangong Moyu' was significantly higher than that in 'Xiahei' (Wei et al., 2020).

[0004] This indicates that the grape VvCHI4D gene may be closely related to fruit development. Therefore, by studying the expression of the grape VvCHI4D gene in plants, an important method is provided for the early maturity of berry plants, avoiding problems such as low prices or even unsalable products caused by too concentrated maturity periods. This is of great significance for fruit farmers to increase their income and reduce investment risks. Summary of the invention

[0005] The purpose of the present invention is to provide an application of grape VvCHI4D gene in promoting early ripening of berry fruits, to study the mechanism of shortening the fruit growth period of grape VvCHI4D gene, to improve the expression of grape VvCHI4D gene in plants, and to cultivate early-maturing berry plants.

[0006] To achieve the above purpose, the technical solution of the present invention is as follows:

[0007] An application of a grape VvCHI4D gene in promoting early ripening of berry fruits. By overexpressing the grape VvCHI4D gene in plants, a transgenic plant with early flowering, color change and ripening can be obtained;

[0008] The protein encoded by the grape VvCHI4D gene is shown in SEQ ID No. 2;

[0009] The nucleotide sequence of the grape VvCHI4D gene is shown in SEQ ID No.1;

[0010] The plant is tomato or grape.

[0011] Preferably, when overexpression is performed, the vector plasmid used is pCAMBIA1301.

[0012] Preferably, when overexpression is performed, the host bacteria used is Agrobacterium GV3101 / EH105.

[0013] A method for advancing the ripening period of berries, comprising the following steps:

[0014] 1) constructing a recombinant plant expression vector containing the grape VvCHI4D gene;

[0015] 2) transforming the constructed recombinant plant expression vector into plant tissues or plant cells;

[0016] 3) Cultivating and screening transgenic plants with early maturity;

[0017] in,

[0018] The protein encoded by the grape VvCHI4D gene is shown in SEQ ID No. 2;

[0019] The nucleotide sequence of the grape VvCHI4D gene is shown in SEQ ID No.1;

[0020] The plant is tomato or grape.

[0021] A transgenic plant with advanced fruit flowering period, color change period and maturity period obtained by the method for advancing the maturity period of berry fruits.

[0022] The present invention observes the phenotypic data of different fruit development stages of the grape variety 'Xiahei' and its early-maturing bud mutation grape variety 'Early Summer Seedless' and finds that the color change of 'Early Summer Seedless' is earlier than that of 'Xiahei'. Through real-time fluorescence PCR verification, it is found that: compared with 'Xiahei', the VvCHI4D of the flesh of 'Early Summer Seedless' maintains a higher expression level 27-56 days after flowering, and the expression levels of VvCHI4D of the two varieties tend to be consistent 63-77 days after flowering. Compared with 'Xiahei', the VvCHI4D of the flesh of 'Early Summer Seedless' maintains a higher expression level 35-56 days after flowering, and the expression levels of VvCHI4D of the two varieties tend to be consistent 56-77 days after flowering, indicating that VvCHI4D plays an important role in the early stage of fruit development, especially in the color change period, and then the function of VvCHI4D is studied through heterologous expression.

[0023] According to the present invention, it is confirmed in tomatoes that the expression of the grape VvCHI4D gene has the function of shortening the fruit growth period, which may be achieved by promoting fruit flowering and fruit color change in advance. This indicates that increasing the expression of the VvCHI4D gene can promote the early maturity of grape and other crops, solving the problem in the prior art that the late maturity of grape fruit is easily affected by extreme high temperature or typhoon weather, resulting in abnormal softening of the fruit and poor taste and appearance quality. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 These are pictures comparing the phenotypes and fruit quality differences of different grape varieties, 'Xiahei' and 'Zaoxia Seedless'.

[0025] Figure 2 This is a schematic diagram of the relative expression level of VvCHI4D in different fruit growth stages of 'Xiahei' and 'Zaoxia Wuhei' of the present invention.

[0026] Figure 3 Plasmid map of the pCAMBIA1302 vector used for subcellular localization of the present invention.

[0027] Figure 4 This is the subcellular localization map of VvCHI4D in the present invention.

[0028] Figure 5 The plasmid map of the pCAMBIA1301 vector used for heterologous transformation of tomatoes in the present invention.

[0029] Figure 6 This is the electrophoresis diagram of PCR identification of tomato strains overexpressing VvCHI4D in the present invention.

[0030] Figure 7 This is a schematic diagram of the growth phenotype observation of the wild-type and overexpression tomato strains in the present invention at 30 days, 60 days, and 90 days after sowing.

[0031] Figure 8 These are pictures of the fruit phenotype and fruit quality changes of different tomato strains in the present invention 120 days after sowing. DETAILED DESCRIPTION

[0032] The present invention is further described below in conjunction with the embodiments and drawings. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0033] If no specific techniques or conditions are specified in the examples, the techniques or conditions described in the literature in the field (e.g., Molecular Cloning Experiment Guide, 3rd edition, Science Press, by J. Sambrook et al., translated by Huang Peitang et al.) or the product instructions are used. If the manufacturer of the reagents or instruments is not specified, they are all conventional products that can be purchased commercially.

[0034] Test materials and treatments

[0035] The experimental materials selected for this study were mature grape trees produced in the field. Fruit samples were collected 27, 35, 42, 49, 56, 63, 70, and 77 days after flowering (DAA27, 35, 42, 49, 56, 63, 70, and 77) of 'Xiahei' and 'Zaoxia Seedless' varieties. The fruits were separated from the peel and quickly frozen in liquid nitrogen and stored in a -80°C refrigerator for subsequent experiments.

[0036] The tomato seeds used in this study were Micro-Tom.

[0037] The seeds used in this study were purchased from Shanghai Weidi Biotechnology Co., Ltd.

[0038] Example 1: VvCHI4D gene cloning and sequence analysis

[0039] Primers VvCHI4D-F1: 5'-ATGGCAGCCAAGCTACTAACA GT-3' (SEQ ID No. 3); VvCHI4D-R1: 5'-TCAGCAAGTGAGGTTGTC ACCA-3' (SEQ ID No. 4) were designed based on the genomic sequence of Pinot Noir on NCBI (https: / / blast.ncbi.nlm.nih.gov / Blast.cgi). The ORF region of VvCHI4D in 'Xiahei' grape was obtained by PCR amplification, as shown in SEQ ID No. 1:

[0040] atggcagccaagctactaacagtccttcttgttggggcccttttcggagctgcagtggctcagaactgtgggtgtgcctcaggcctatgttgcagtaagtacgggtactgtggcaccggcagtgactactgcggtgacggttgccaatcgggtccttgtgattcaagtagcggcagtggtagctcggtttctgatattgtgacacagtcatttttcgatgggataattaatcaagctgcttcgagttgtgctgggaagaatttctacacccgtgcagcgtttctcagtgccttgaattcgtattctgggttcggcaacgatggttctaccgatgctaataagcgcgagattgcagctttcttcgctcatgtcacacacgagactggacacttttgttatattgaagaaatcaatggtgcctctcataactactgtgattcaagcaatacccaatatccatgtgtctccggtcaaaattactacggccgtggaccgcttcaactaacgtggaactacaactatggcgctgctggaaacagcattggattcaatggcttgagcaaccctggaattgttgcaactgacgtggttacttcattcaagaccgcattatggttttggatgaataatgttcactctgtcataggccaaggttttggtgccacaattcgagccatcaatggtgccgtcgaatgtaatggtggaaacacagctgccgttaacgcccgcgttcagtattacaaggactactgcagtcagctcggggtttcacctggtgacaacctcacttgctga

[0041] The protein sequence encoded by this gene is shown in SEQ ID No.2:

[0042] MAAKLLTVLLVGALFGAAVAQNCGCASGLCCSKYGYCGTGSDYCGDGCQSGPCDSSSGSGSSVSDIVTQSFFDGIINQAASSCAGKNFYTRAAFLSALNSYSGFGNDGSTDANKREIAAFFAHVTHETGHFC YIEEINGASHNYCDSSNTQYPCVSGQNYYGRGPLQLTWNYNYGAAGNSIGFNGLSNPGIVATDVVTSFKTALWFWMNNVHSVIGQGFGATIRAINGAVECNGGNTAAVNARVQYYKDYCSQLGVSPGDNLTC

[0043] Example 2: Real-time fluorescence quantitative PCR analysis of the expression pattern of VvCHI4D

[0044] The fruit and peel materials of 'Xiahei' and 'Zaoxia Wushi' stored at ultra-low temperature were taken, and total RNA was extracted using an RNA extraction kit (Oemga) according to the instructions. The purity was detected by 1% agarose electrophoresis, and the concentration was determined by an ultraviolet spectrophotometer (ThermoScientific, NANO DROP2000) and then stored in a -70℃ refrigerator for later use. 1μg RNA was taken and reverse transcribed to synthesize the first strand cDNA using PrimeScriptII 1st strand cDNA Synthesis Kit for subsequent experiments.

[0045] Real-time fluorescence quantitative PCR reaction was performed by Roche The reaction was performed on a 480II real-time fluorescence quantitative PCR instrument using SYBR Premix Ex TaqTM (Perfect Real Time) reagent (TaKaRa). The reaction system was 20 μL, which contained 10 μL of SYBR Premix Ex TaqTM (2x), 1 μL of cDNA, 0.3 μL of each upstream and downstream primer (10 μmol / L), and 8.4 μL of ddH2O. Each sample was repeated 3 times. The reaction conditions were pre-denaturation at 95°C for 30 s; 40 cycles of 95°C for 5 s and 60°C for 34 s. The fluorescence intensity was set to detect and collect signals at the end of 60°C (refer to the TaKaRa company's operating manual).

[0046] The internal reference gene was GAPDH, whose GenBank accession number was VIT_17s0000g10430, and the primer sequences were: 5'-TTCCGTGTTCCTACTGTTG-3' (SEQ ID No. 5) and 5'-CCTCTGACTCCTCCTTGAT-3' (SEQ ID No. 6); the primer sequences of the target gene VvCHI4D were: 5'-AGTGGCTCAGAACTGTGGGTGTG-3' (SEQ ID No. 7), 5'-GGCAACCGTCACCGCAGT-3' (SEQ ID No. 8). The relative expression of the target gene was calculated using 2 -ΔΔc Law.

[0047] Figure 1 The following are pictures comparing the phenotypes and fruit quality differences of different grape varieties, 'Summer Black' and 'Early Summer Seedless'. A is a phenotype diagram of different grape varieties during the fruit development period; B is the difference in soluble solids content, titratable acid and single fruit weight of different grape varieties during the fruit development period. DAA27 means 27 days after flowering, and the others are similar. As can be seen from the figure, the skin color of 'Early Summer Seedless' grapes began to change on the 42nd day, with a higher soluble solids content and a lower titratable acid content, proving that its maturity period is earlier than that of 'Summer Black' grapes.

[0048] Figure 2 The figure is a schematic diagram of the relative expression of VvCHI4D in the fruit growth period of different grape varieties of the present invention, 'Summer Black' and 'Early Summer Seedless'. A is the relative expression of VvCHI4D in the pulp; B is the relative expression of VvCHI4D in the peel. As can be seen from the figure, the results of real-time fluorescence quantitative PCR show that the expression of VvCHI4D gene in the peel and pulp of 'Early Summer Seedless' is higher than that of 'Summer Black', and the difference is greater 35 to 56 days after flowering, which is the fruit color change period, indicating that VvCHI4D plays a major role in the fruit color change period, indirectly affecting the promotion of grape fruit maturity in advance. The variance analysis is based on the relative expression of VvCHI4D in different grape varieties at the same time point. *: p<0.1; ns: no significant difference.

[0049] Example 3: Localization of VvCHI4D gene in subcellular

[0050] like Figure 3As shown, pCAMBIA1302 vector was used, KpnCHI4D-F1: 5'-CTCGAGGGGGGGCCCGGTACCATGGCAGCCAAGCTACTAACAGTC C-3' (SEQ ID No. 13), BamCHI4D-R1: 5'-GATCTGCAGCCCGGGGGATCCTCAGCAAGTGAGGTTGTCACCAGG-3' (SEQ ID No. 14); VvCHI4D and pCAMBIA1302 were double-digested with KpnI / BamHI, the target gene fragment and the vector backbone were recovered, and Escherichia coli were connected and transformed. One PCR-positive transformant was selected for plasmid extraction and sequencing. The sequencing results showed that the target gene sequence was completely correct and was connected to the N-terminus of the GFP gene. That is, the subcellular localization expression vector pCAMBIA1302::VvCHI4D::GFP was successfully constructed.

[0051] The constructed vector plasmid was then transferred into Agrobacterium GV3101. Finally, the Agrobacterium suspension was injected into tobacco seedlings (Nicotiana rustica var Pavonii) and cultured under weak light conditions for 2 days. The tobacco leaves were then observed and photographed using a confocal microscope (C2-ER; Nikon). The excitation wavelength of GFP green fluorescence is 488nm and the emission wavelength is 507nm.

[0052] In this example, a vector for VvCHI4D fusion expression driven by CaMV35S promoter was constructed. The vector was transformed into tobacco leaves for transient expression using GFP empty vector as control. The results are shown in Figure 2. Figure 4 As shown, VvCHI4D is localized on the cell membrane.

[0053] Example 4: Preparation of VvCHI4D overexpressing tomatoes and study of its involvement in fruit growth period

[0054] like Figure 5As shown, pCAMBIA1301-35SN vector was used, primer sequence KpnCHI4D-F1: 5'-CTCGAGGGGGGGCCCGGTACCATGGCAGCCAAGCTACTAACAGTC C-3' (SEQ ID No. 13), BamCHI4D-R1: 5'-GATCTGCAGCCCGGGGGATCC TCAGCAAGTGAGGTTGTCACCAGG-3' (SEQ ID No. 14); VvCHI4D and pCAMBIA1301 were double-digested by KpnI / BamHI, the target gene fragment and vector backbone were recovered, and Escherichia coli were connected and transformed. One PCR-positive transformant was selected for plasmid extraction. The sequencing results showed that the target gene sequence was completely correct, that is, the pCAMBIA1301::VvCHI4D::GFP vector was successfully constructed.

[0055] Select plump, uniform, fresh tomato (Micro-Tom) seeds and rinse them repeatedly with sterile water for several times. Disinfect the tomato seeds with 70% alcohol for 30 seconds and 10% sodium hypochlorite for 10 minutes, rinse with sterile water 4-5 times, dry with sterile filter paper, and inoculate in seed germination medium. After dark culture until most seeds germinate and turn white, place them in a light environment of 16 hours per day, with a light intensity of 1600-1800 lux and a temperature of (24±2)℃. Select true leaves, cotyledons, and hypocotyls as explants for transformation. Cut off the leaf tips and petioles of true leaves and cotyledons, and cut the rest into leaf pieces of 0.5cm×0.5cm; cut the hypocotyls into segments of about 0.5-0.6mm in length and place them horizontally on the pre-culture medium, with 15-20 pieces per dish. The culture conditions are the same as above, and the pre-culture is 1 day.

[0056] Take the competent Agrobacterium stored at -80℃ and place it at room temperature or in the palm of your hand for a while until it partially melts. When it is in an ice-water mixture, insert it into ice. Add 0.1μg (volume not more than 10μl) plasmid DNA to every 100μL competent state, stir the bottom of the tube by hand to mix, and let it stand on ice for 5min, liquid nitrogen for 5min, 37℃ water bath for 5min, and ice bath for 5min. Add 700μL LB liquid culture medium without antibiotics and culture at 28℃ and 200rpm for 2-3h. Centrifuge at 6000rpm for one minute to collect the bacteria, keep about 100μL of supernatant, gently blow and resuspend the bacteria block, apply it on the LB plate containing the corresponding antibiotics, and invert it in a 28℃ incubator for 2-3d. Randomly select 2-3 single colonies, do colony PCR, identify the correct Agrobacterium monoclonal clone and mark it for use. Use a sterile pipette to pick up the marked Agrobacterium monoclone and inoculate it into 5 mL of LB liquid medium containing the corresponding antibiotics and culture it at 28°C, 200 rpm for 24 hours. Centrifuge at 20°C, 4000 rpm for 15 minutes to collect the bacteria; blow the bacteria evenly with the transformation solution and resuspend them to OD 600 =about 0.5.

[0057] Remove the explants from the pre-culture medium and dilute them to OD 600 = 0.5 of Agrobacterium, transform for 30 minutes, remove the explants, blot dry on sterile paper, put them back into the pre-culture medium, and co-culture for 1-2 days. The co-cultured explants are transferred to the screening medium for selective culture. After a few days of selective culture, the cotyledons begin to thicken and the hypocotyls begin to thicken. The transformed explants will form callus and adventitious buds on the selective culture medium. Subculture once every 14 days. Complete plants are obtained by elongating and rooting the adventitious buds.

[0058] DNA was extracted from the transformed tomato seedlings of the T1 generation, and positive plants were identified by PCR using primers HYG1-F: 5'-GCTTCTGCGGGCGATTTGTGT-3' (SEQ ID No. 15) and HYG1-R: 5'-GGTCGCGGAGGCTATGGATGC-3' (SEQ ID No. 16). Figure 6 It can be seen that the transgenic lines VvCHI4D-OE1, VvCHI4D-OE3, and VvCHI4D-OE5 are positive plants and can be used for further analysis. Figure 6 Middle M: ​​DL2000 DNA Marker; 2-7: PCR results of tomato overexpression lines OE1, OE2, OE3, OE4, OE5, OE6; 8: CHI4D plasmid PCR results; 9: Micro-Tom gene DNA PCR results; 10: H 2 O is the PCR result of the template.

[0059] The wild-type tomato (Micro-Tom) and the three homozygous transgenic lines VvCHI4D-OE1, VvCHI4D-OE3, and VvCHI4D-OE5 were planted at the same time for plant growth and development observation and fruit quality test, and photos were taken. Each experiment was repeated three times.

[0060] like Figure 7 and Figure 8 As shown in the figure, the phenological period statistics showed that the flowering and maturity periods of the transgenic lines VvCHI4D-OE1, VvCHI4D-OE3, and VvCHI4D-OE5 were significantly higher than those of wild-type tomatoes. The fruits of the control lines were still green 120 days after sowing, but the fruits of the transgenic lines VvCHI4D-OE1, VvCHI4D-OE3, and VvCHI4D-OE5 were all orange-red. This result shows that the grape VvCHI4D gene plays an important role in the growth and development of plant fruits.

[0061] Figure 8 In the figure, A: fruit phenotype of different tomato strains; B: single fruit weight, soluble solid content and organic acid content of different tomato strains. Different lowercase letters indicate that the indicators of different tomato strains are significantly different (p<0.1).

[0062] like Figure 8 As shown in the figure, the results of fruit quality measurement data showed that there were no significant differences in single fruit weight and soluble solid content between the control strain and the transgenic strains VvCHI4D-OE1, VvCHI4D-OE3, and VvCHI4D-OE5, but the organic acid content of VvCHI4D-OE1, VvCHI4D-OE3, and VvCHI4D-OE5 was significantly lower than that of the control strain, indicating that overexpression of the grape VvCHI4D gene has an effect on the accumulation of organic acid content in the fruit.

[0063] It should be noted that: Figure 1-2 Among them, 'Xiahei' is the main grape variety, and 'Zaoxia Seedless' is an early-maturing bud mutation of 'Xiahei';

[0064] Figure 6-8 In the figure, Micro-Tom is the wild type, and OE1, OE3 and OE5 are three tomato lines overexpressing VvCHI4D.

[0065] The present invention observes the phenotypic data of different fruit development stages of the grape variety 'Summer Black' and its early-maturing bud mutation grape variety 'Early Summer Seedless' and finds that 'Early Summer Seedless' changes color earlier than 'Summer Black', and finds that compared with 'Summer Black', the gene VvCHI4D is highly expressed in both the skin and the flesh of 'Early Summer Seedless'. Through real-time fluorescence PCR verification, it is found that: compared with 'Summer Black', the VvCHI4D in the flesh of 'Early Summer Seedless' maintains a higher expression level at 35 to 56 days after flowering, and the expression levels of VvCHI4D in the two varieties tend to be consistent at 63 to 77 days after flowering. This shows that VvCHI4D plays an important role in the early stage of fruit development, especially in the color change period, and then the function of VvCHI4D is studied by heterologous expression.

[0066] According to the present invention, the grape VvCHI4D gene is expressed in tomatoes and has the function of shortening the growth period of tomato fruits, which may be achieved by promoting fruit flowering and fruit color change in advance.

[0067] The invention provides a theoretical basis for promoting the early maturity of grapes and other crops by increasing the expression amount of VvCHI4D.

[0068] The above is only a preferred embodiment of the present invention, and is not intended to limit the scope of the present invention. The above embodiments of the present invention can also be modified in various ways. All simple, equivalent changes and modifications made according to the claims and the description of the present invention fall within the scope of protection of the claims of the present invention. The contents not described in detail in the present invention are all conventional technical contents.

Claims

1. An application of grape VvCHI4D gene in promoting early ripening of berry fruits, characterized in that: By overexpressing the grape VvCHI4D gene in plants, a transgenic plant with early flowering, color change and maturity can be obtained; The protein encoded by the grape VvCHI4D gene is shown in SEQ ID No. 2; The nucleotide sequence of the grape VvCHI4D gene is shown in SEQ ID No.1; The plant is tomato or grape.

2. The use of the grape VvCHI4D gene in promoting early ripening of berry fruits as claimed in claim 1, characterized in that: When overexpression was performed, the vector plasmid used was pCAMBIA1301.

3. The use of the grape VvCHI4D gene in promoting early ripening of berry fruits as claimed in claim 1, characterized in that: When overexpression was performed, the host bacteria used was Agrobacterium GV3101 / EHA105.

4. A method for advancing the ripening period of berry fruits, characterized in that: The steps include: 1) constructing a recombinant plant expression vector containing the grape VvCHI4D gene; 2) transforming the constructed recombinant plant expression vector into plant tissues or plant cells; 3) Cultivating and screening transgenic plants with early maturity; in, The protein encoded by the grape VvCHI4D gene is shown in SEQ ID No. 2; The nucleotide sequence of the grape VvCHI4D gene is shown in SEQ ID No.1; The plant is tomato or grape.

5. A transgenic plant with advanced fruit flowering, color change and maturity obtained by the method for advancing the maturity of berry fruits according to claim 4.

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