CN114807197A - Method for creating tomato dwarfing material through gene editing and application thereof - Google Patents

Abstract

The invention discloses a method for creating a tomato dwarfing material through gene editing and application thereof, wherein the method for creating the tomato dwarfing material through gene editing can be used for cultivating tomato plant types; the method for creating the tomato dwarfing material by gene editing comprises the following steps of: comprises the selection of a gRNA target site and the construction of a recombinant plasmid SlABIG1-CP 098; and then tomato transformation: taking tomato cotyledons as explants, mediating tomato transformation by positive agrobacterium GV3101, and obtaining T0 generation transgenic tomatoes through kanamycin resistance screening, wherein homozygous editing plants are tomato dwarfing materials; carrying out the identification of the DNA level of the tomato with the SlABIG1 transgenic generation T0 and the phenotypic identification of the tomato with the transgenic SlABIG1 gene; the tomato plants obtained by the invention have the advantages of high-density planting, full utilization of space and land, enhanced photosynthetic efficiency, low water consumption rate and lodging resistance; has important application value.

Description

Method for creating tomato dwarfing material through gene editing and application thereof

Technical Field

The invention belongs to the technical field of tomato plant type cultivation, and particularly relates to a method for creating a tomato dwarfing material through gene editing and application thereof.

Background

Transgenic tomatoes are commercially grown in many regions and countries of the world. The characters of the cultivated transgenic tomato comprise storage resistance, virus resistance, fungus resistance, insect resistance, herbicide resistance, freezing resistance, salt resistance, quality improvement, high yield and the like. The tomato is one of the vegetables with higher global economic benefit. The tomato with virus resistance, fungus resistance and insect resistance can improve the disease resistance of the wild tomato; the tomato with long storage life can reduce the postpartum economic loss of the tomato and balance the supply period; the salt-tolerant tomatoes can adapt to the soil environment with severe conditions, and the land utilization rate is improved. The plant type of the tomato plays an important role in close planting, space illumination utilization, irrigation, environmental resistance and yield.

Disclosure of Invention

The invention aims to provide a method for creating a tomato dwarfing material through gene editing and application thereof, solves the problems in the prior art, and aims to improve the tomato plant type and improve the economic benefit of tomato planting.

In order to achieve the purpose, the invention provides the following technical scheme: a method of creating tomato dwarf material by gene editing comprising the steps of:

firstly, constructing a recombinant vector;

(1) selection of gRNA target sites:

the gRNA target site is selected in the exon region of SlABIG1 gene, 20bp base of the adjacent motif of the prototype spacer sequence is selected as the target site according to the CRISPR/Cas9 target site anchoring principle, namely 5'-N20NGG-3', NGG is PAM sequence, and N20 represents the recognition sequence of 20bp base; the target sequence is: target 1: 5'-GGTTCTTCAAAGCCAACACA-3', respectively; target 2: 5'-GCTGGTAACTGCATGTACAA-3', respectively;

designing a primer according to the target sequence, wherein the primer sequence is as follows: SlABIG 1-CE-F:

5’-AATCTAACAGTGTAGTTTGGGTTCTTCAAAGCCAACACAGTTTTAGAGCTAGAAATAGC-3’；SlABIG1-CE-R：

5’-CTATTTCTAGCTCTAAAACTTGTACATGCAGTTACCAGCCAAACTACACTGTTAGATTC-3’；

(2) and constructing a recombinant plasmid SlABIG1-CP 098:

carrying out PCR amplification by taking a plasmid CP043 as a template and a nucleotide sequence shown by a primer sequence SlABIG1-CE-F and a primer sequence SlABIG1-CE-R as primers to obtain a 599bp PCR amplification product;

the PCR reaction system reagent is as follows: 2X Phanta Flash Master Mix, 25 ul; 2ul of each primer and 2ul of DNA plasmid template; sterile water 19 ul; after the addition, the PCR solution was placed on a PCR instrument for amplification under the following conditions: pre-denaturation at 98 ℃ for 30 s; denaturation at 98 deg.C for 10 s; annealing at 55 deg.C for 5 s; extending at 72 ℃ for 30 s; final extension at 72 deg.C for 1 min; obtaining PCR amplification products after 36 cycles;

carrying out enzyme digestion on the CP098 vector by using Bsa1 to obtain an enzyme digestion product;

the enzyme digestion reaction system comprises the following reagents: bsa1, 2ul, plasmid template, 5ug, 10x Buffer, 5ul, sterile water is added to 50ul, and PCR reaction conditions are 37 ℃ and 2 h;

recovering PCR amplification products and enzyme digestion products by using a PCR product purification kit to obtain recovered fragments, integrating the PCR amplification products to a CP098 vector by using a homologous recombination method, transferring the recombinant products to escherichia coli competent cells, plating the cells overnight, selecting a single clone to perform sequencing analysis on the plasmid, wherein a sequencing primer is CP 098-seq-F: 5'-aacgacggccagtgccaagc-3', extracting plasmids from the bacterial liquid with correct sequencing result, and transferring the plasmids into agrobacterium GV3101 by a freeze-thaw method;

secondly, tomato transformation: tomato cotyledons are taken as explants, positive agrobacterium GV3101 is used for mediating tomato transformation, and T0 generation transgenic tomatoes are obtained through kanamycin resistance screening; the method comprises the following steps:

and (3) culturing aseptic tomato seedlings: carefully selecting full tomato seeds with consistent size, soaking the tomato seeds in distilled water for 30min, then disinfecting the tomato seeds with 75% ethanol for 1min, then sterilizing the surfaces of the tomato seeds with 50% sodium hypochlorite solution for 15min, finally cleaning the tomato seeds with sterilized ddH2O for 3-4 times, sowing the tomato seeds in 1/2MS culture medium, and placing the tomato seeds in a tissue culture room with the light cycle of 16 h/8 h in the dark for culture for 7-8 d;

activating agrobacterium: the agrobacterium GV3101 of the target vector is drawn on LB culture medium containing the corresponding antibiotic; selecting a single colony, inoculating the single colony into a 20mL LB liquid culture medium containing 50mg/L kanamycin and 35mg/L rifampicin, performing dark culture at the temperature of 200r/min and 28 ℃ for 14-20h, centrifuging the bacterial liquid OD600=1 at 10000r/min for 30s, and suspending and diluting the bacterial body to OD600=0.1-0.3 by using a 0.2MS liquid culture medium for later use;

pre-culturing: culturing aseptic seedlings for 7-8 days, selecting two seedlings with completely unfolded cotyledons, cutting off the cotyledons from petioles, cutting off the petioles and tips of the cotyledons, cutting the leaves into 2-3 sections, flatly placing the cut cotyledons on a KCMS culture medium paved with sterilized filter paper, sealing a culture dish by using a sealing film to prevent bacteria and fungi from invading, and placing the culture dish in a tissue culture room for dark culture for 1 d;

infection: pouring the 0.2MS suspension after high-temperature sterilization into a culture dish, and then suspending the cotyledons cultured in the dark for one day in a 0.2MS liquid culture medium; finally, 100-300uL of agrobacterium tumefaciens suspension with OD600=0.1-0.3 is added, the culture dish is covered and is rotated, and the infection time is 3-4 min; after infection, pouring out bacterial liquid, sucking the bacterial liquid on cotyledons by using sterile filter paper, putting the infected cotyledons back on the original KCMS culture medium, and performing dark culture for 2d in a tissue culture room;

regeneration: transferring the infected cotyledons to a corresponding 2Z culture medium for regeneration, wherein the right sides of the cotyledons face upwards, and the cotyledons are uniformly spaced and cultured in a tissue culture room; when the infected tomato cotyledons grow on a 2Z culture medium for 10-14 days, callus is generated and bud points are formed; more bud points are generated on the callus at 14-20 days, and in order to reduce the bud point aberration rate, the callus generating the bud points is transferred to a culture medium containing corresponding 0.2Z for continuous culture;

obtaining and transplanting transgenic plants: growing the bud point on a 0.2Z culture medium to form a1 cm-long bud, selecting the bud with a complete growing point, cutting off the bud from the stem base part, and performing rooting culture on an R culture medium; culturing for one week, and rooting the transgenic plant; transplanting the seedling into a square pot filled with nutrient soil and vermiculite when the root grows to 5-6cm, covering and moisturizing for 3-4d, gradually uncovering, and irrigating with nutrient water; thus obtaining T0 generation SlABIG1 gene tomato.

The invention also provides application of the method for creating the tomato dwarfing material through gene editing, which is used for cultivating the tomato plant type.

Compared with the prior art, the invention has the following beneficial effects:

the invention obtains homozygous editing plants, namely the tomato dwarfing material, by carrying out gene editing on the exon regions of the SlABIG1 gene; the tomato plant obtained by the invention has the advantages of high-density planting, full utilization of space and land, enhanced photosynthetic efficiency, low water consumption rate and lodging resistance.

Drawings

In FIG. 1: (a) identifying a sequencing result graph of the pure plant; (b) a phenotypic graph of the inbred plants and the Micro Tom; (c) and a determination chart of the heights of the pure plants and the Micro Tom plants.

FIG. 2 shows the full-length sequence of the gene.

FIG. 3 shows the gene protein sequence.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

The experimental materials were as follows:

adopting a tomato variety of Micro Tom from a solanaceae disease-resistant germplasm innovation research team of gardening college of Henan university;

coli competence: treliestm 5 a; the agrobacterium is infected with the attitude: GV 3101;

the primer synthesis and sequencing are completed by Beijing Optimalaceae Biotechnology Co.

This example provides a method of creating tomato dwarf material by gene editing comprising the steps of:

firstly, constructing a recombinant vector;

(1) selection of gRNA target sites:

the gRNA target site is selected in the exon region of SlABIG1 gene, 20bp base of the adjacent motif of the prototype spacer sequence is selected as the target site according to the CRISPR/Cas9 target site anchoring principle, namely 5'-N20NGG-3', NGG is PAM sequence, and N20 represents the recognition sequence of 20bp base; the target sequence is: target 1: 5'-GGTTCTTCAAAGCCAACACA-3' (SEQ ID NO: 1); target 2: 5'-GCTGGTAACTGCATGTACAA-3' (SEQ ID NO: 2);

designing a primer according to the target sequence, wherein the primer sequence is as follows: SlABIG 1-CE-F: 5'-AATCTAACAGTGTAGTTTGGGTTCTTCAAAGCCAACACAGTTTTAGAGCTAGAAATAGC-3' (SEQ ID NO: 3); SlABIG 1-CE-R: 5'-CTATTTCTAGCTCTAAAACTTGTACATGCAGTTACCAGCCAAACTACACTGTTAGATTC-3' (SEQ ID NO: 4);

(2) and constructing a recombinant plasmid SlABIG1-CP 098:

carrying out PCR amplification by taking the plasmid CP043 as a template and taking nucleotide sequences shown in the sequence 3 and the sequence 4 as primers to obtain a 599bp PCR amplification product;

the PCR reaction system reagent is as follows: 2X Phanta Flash Master Mix, 25 ul; 2ul of each primer and 2ul of DNA plasmid template; sterile water 19 ul; after the addition, the PCR solution was placed on a PCR instrument for amplification under the following conditions: pre-denaturation at 98 ℃ for 30 s; denaturation at 98 deg.C for 10 s; annealing at 55 deg.C for 5 s; extending at 72 ℃ for 30 s; final extension at 72 deg.C for 1 min; obtaining PCR amplification products after 36 cycles;

carrying out enzyme digestion on the CP098 vector by using Bsa1 to obtain an enzyme digestion product;

the enzyme digestion reaction system comprises the following reagents: bsa1, 2ul, plasmid template, 5ug, 10x Buffer, 5ul, sterile water is added to 50ul, and PCR reaction conditions are 37 ℃ and 2 h;

recovering PCR amplification products and enzyme digestion products by using a PCR product purification kit to obtain recovered fragments, integrating the PCR amplification products to a CP098 vector by using a homologous recombination method, transferring the recombinant products to escherichia coli competent cells, plating the cells overnight, selecting a single clone to perform sequencing analysis on the plasmid, wherein a sequencing primer is CP 098-seq-F: 5'-aacgacggccagtgccaagc-3' (sequence 5), extracting plasmids from the bacterial liquid with correct sequencing result, and transferring the plasmids into agrobacterium GV3101 by a freeze-thaw method;

secondly, tomato transformation;

the tomato cotyledon is taken as an explant, positive agrobacterium GV3101 is used for mediating tomato transformation, and the T0 generation transgenic tomato is obtained through kanamycin resistance screening.

The method mainly comprises the following steps:

and (3) culturing aseptic tomato seedlings: carefully selecting full tomato seeds with consistent size, soaking the tomato seeds in distilled water for 30min, then disinfecting the tomato seeds with 75% ethanol for 1min, then sterilizing the surfaces of the tomato seeds with 50% sodium hypochlorite solution for 15min, finally cleaning the tomato seeds with sterilized ddH2O for 3-4 times, sowing the tomato seeds in 1/2MS culture medium, and placing the tomato seeds in a tissue culture room with the light cycle of 16 h/8 h in the dark for culture for 7-8 d;

activating agrobacterium: the agrobacterium GV3101 of the target vector is drawn on LB culture medium containing the corresponding antibiotic; selecting a single colony, inoculating the single colony into a 20mL LB liquid culture medium containing 50mg/L kanamycin and 35mg/L rifampicin, performing dark culture at the temperature of 28 ℃ at 200r/min for 14-20h, and enabling the common bacterial liquid OD600= 1; centrifuging at 10000r/min for 30s, and suspending and diluting the thallus by using a 0.2MS liquid culture medium until the OD600=0.1-0.3 is reserved;

pre-culturing: culturing aseptic seedlings for 7-8 days, selecting two seedlings with completely unfolded cotyledons, cutting off the cotyledons from petioles, cutting off the petioles and tips of the cotyledons, cutting the leaves into 2-3 sections, flatly placing the cut cotyledons on a KCMS culture medium paved with sterilized filter paper, sealing a culture dish by using a sealing film to prevent bacteria and fungi from invading, and placing the culture dish in a tissue culture room for dark culture for 1 d;

infection: pouring the 0.2MS suspension after high-temperature sterilization into a culture dish, and then suspending the cotyledons cultured in the dark for one day in a 0.2MS liquid culture medium; finally, 100-300uL of agrobacterium tumefaciens suspension with OD600=0.1-0.3 is added, the culture dish is covered and is rotated, and the infection time is 3-4 min; after infection, pouring out bacterial liquid, sucking the bacterial liquid on cotyledons by using sterile filter paper, putting the infected cotyledons back on the original KCMS culture medium, and performing dark culture for 2d in a tissue culture room;

regeneration: transferring the infected cotyledons to a corresponding 2Z culture medium for regeneration, keeping the right side of the cotyledons upward, keeping a proper distance between the cotyledons and a proper density, and culturing for about 2 weeks in a tissue culture room; when the infected tomato cotyledons grow on a 2Z culture medium for 10-14 days, a large amount of callus begins to be generated and bud points are formed; the callus tissue generates a large amount of buds in 14-20 days, and in order to reduce the bud aberration rate, the callus tissue generating the buds is transferred to a culture medium containing corresponding 0.2Z for continuous culture;

obtaining and transplanting transgenic plants: when the bud point grows on the 0.2Z culture medium for about 2 weeks, a bud with the length of about 1cm is formed, at the moment, the bud with a complete growing point is selected to be cut off from the stem base part, and rooting culture is carried out on an R culture medium; culturing for about one week to make transgenic plant take root; transplanting the seedling into a square pot filled with nutrient soil and vermiculite when the root grows to 5-6cm, covering and moisturizing for 3-4d, gradually uncovering, and irrigating with nutrient water; and transforming the tomato to obtain the T0 generation transformed SlABIG1 gene tomato.

Thirdly, identifying the tomato DNA level of the T0 generation-transformed SlABIG1 gene;

primers were designed based on cas9 protein and the sequences were as follows: cas 9-F: 5'-atggattacaaggaccacgac-3' (SEQ ID NO: 6); cas 9-R: 5'-gagcctagcggacaggatag-3' (SEQ ID NO: 7). Respectively extracting genome DNA of the transgenic tomato plants, performing PCR amplification by using the genome DNA as a template, and if the PCR amplification product contains a DNA fragment with the size of 765bp, the corresponding T0 generation SlABIG1 gene-transformed tomato is a T0 generation positive plant; if the PCR amplification product does not contain a DNA fragment with the size of 765bp, the corresponding T0 generation SlABIG1 gene-transferred tomato is a negative plant. And designing a primer for target site amplification according to the upstream and downstream sequences of the target site, wherein the primer sequence is as follows: CRABIG 1-F: 5'-CCATCCCTTTTCTCTTTTCC-3' (SEQ ID NO: 8); CRABIG 1-R: 5'-CGGATTGTTGAATGGACTGT-3' (SEQ ID NO: 9). And (3) carrying out target site amplification on the template with the correct cas9 band, sequencing the amplified product, and judging whether the regenerated plant is subjected to gene editing. The plants edited by the SlABIG1 gene of 3 strains identified by the method are named as SlABIG1-1, SlABIG1-8 and SlABIG 1-14. Three materials were planted in a climatic chamber and transgenic seeds for the T1 generation were obtained by selfing. And (3) sowing seeds of T1 generation into a plug tray, extracting genomic DNA of tomato plants after seedlings grow, and identifying the pure plants which do not contain cas9 protein and have edited target sites by using a PCR method. SlABIG1-8-3 is found to have a homozygous mutation at the target site position, and a base T is inserted, so that a homozygous line of SlABIG1-8-3 is obtained, as shown in figure 1 (a).

Fourthly, phenotype identification of the transgenic SlABIG1 gene pure line tomato;

and the plants to be detected are the SlABIG1 gene editing pure line plants obtained in the third step and Micro Tom (used as control plants).

Culturing the plant to be tested in an artificial climate chamber, and managing normal water and fertilizer to ensure that the water and fertilizer conditions of all the plants are basically consistent. The plants are not forked and are allowed to grow naturally. After 5 weeks of phytotron culture, the height of the gene editing inbred material was significantly less than the control group, see (b) and (c) in fig. 1.

In conclusion, the invention provides a method for creating a tomato dwarfing material through gene editing, which is characterized in that homozygous edited plants are obtained by performing gene editing on an exon region of a SlABIG1 gene, namely the tomato dwarfing material. The tomato plant obtained by the invention has the advantages of high-density planting, full utilization of space and land, enhanced photosynthetic efficiency, low water consumption rate and lodging resistance.

FIG. 2 the full length sequence of the gene, the underlined sequences being intron sequences; the shaded portion is the gene recognition site. FIG. 3 Gene protein sequence.

Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.

Claims (2)

1. A method of creating tomato dwarf material by gene editing comprising the steps of:

firstly, constructing a recombinant vector;

(1) selection of gRNA target sites:

the gRNA target site is selected in the exon region of SlABIG1 gene, 20bp base of the adjacent motif of the prototype spacer sequence is selected as the target site according to the CRISPR/Cas9 target site anchoring principle, namely 5'-N20NGG-3', NGG is PAM sequence, and N20 represents the recognition sequence of 20bp base; the target sequence is: target 1: 5'-GGTTCTTCAAAGCCAACACA-3', respectively; target 2: 5'-GCTGGTAACTGCATGTACAA-3', respectively;

designing a primer according to the target sequence, wherein the primer sequence is as follows: SlABIG 1-CE-F: 5'-AATCTAACAGTGTAGTTTGGGTTCTTCAAAGCCAACACAGTTTTAGAGCTAGAAATAGC-3', respectively; SlABIG 1-CE-R: 5'-CTATTTCTAGCTCTAAAACTTGTACATGCAGTTACCAGCCAAACTACACTGTTAGATTC-3', respectively;

(2) and constructing a recombinant plasmid SlABIG1-CP 098:

carrying out PCR amplification by taking a plasmid CP043 as a template and a nucleotide sequence shown by a primer sequence SlABIG1-CE-F and a primer sequence SlABIG1-CE-R as primers to obtain a 599bp PCR amplification product;

the PCR reaction system reagent is as follows: 2X Phanta Flash Master Mix, 25 ul; 2ul of each primer and 2ul of DNA plasmid template; sterile water 19 ul; after the addition, the PCR solution was placed on a PCR instrument for amplification under the following conditions: pre-denaturation at 98 ℃ for 30 s; denaturation at 98 deg.C for 10 s; annealing at 55 deg.C for 5 s; extending at 72 ℃ for 30 s; final extension at 72 deg.C for 1 min; obtaining PCR amplification products after 36 cycles;

carrying out enzyme digestion on the CP098 vector by using Bsa1 to obtain an enzyme digestion product;

the enzyme digestion reaction system comprises the following reagents: bsa1, 2ul, plasmid template, 5ug, 10x Buffer, 5ul, sterile water is added to 50ul, and PCR reaction conditions are 37 ℃ and 2 h;

recovering PCR amplification products and enzyme digestion products by using a PCR product purification kit to obtain recovered fragments, integrating the PCR amplification products to a CP098 vector by using a homologous recombination method, transferring the recombinant products to escherichia coli competent cells, plating the cells overnight, selecting a single clone to perform sequencing analysis on the plasmid, wherein a sequencing primer is CP 098-seq-F: 5'-aacgacggccagtgccaagc-3', extracting plasmids from the bacterial liquid with correct sequencing result, and transferring the plasmids into agrobacterium GV3101 by a freeze-thaw method;

secondly, tomato transformation: tomato cotyledons are taken as explants, positive agrobacterium GV3101 is used for mediating tomato transformation, and T0 generation transgenic tomatoes are obtained through kanamycin resistance screening; the method comprises the following steps:

and (3) culturing aseptic tomato seedlings: carefully selecting full tomato seeds with consistent size, soaking the tomato seeds in distilled water for 30min, then disinfecting the tomato seeds with 75% ethanol for 1min, then sterilizing the surfaces of the tomato seeds with 50% sodium hypochlorite solution for 15min, finally cleaning the tomato seeds with sterilized ddH2O for 3-4 times, sowing the tomato seeds in 1/2MS culture medium, and placing the tomato seeds in a tissue culture room with the light cycle of 16 h/8 h in the dark for culture for 7-8 d;

activating agrobacterium: agrobacterium GV3101 of the target vector is streaked on LB medium containing the corresponding antibiotic; selecting a single colony, inoculating the single colony into a 20mL LB liquid culture medium containing 50mg/L kanamycin and 35mg/L rifampicin, performing dark culture at the temperature of 200r/min and 28 ℃ for 14-20h, centrifuging the bacterial liquid OD600=1 at 10000r/min for 30s, and suspending and diluting the bacterial body to OD600=0.1-0.3 by using a 0.2MS liquid culture medium for later use;

pre-culturing: culturing aseptic seedlings for 7-8 days, selecting two seedlings with completely unfolded cotyledons, cutting off the cotyledons from petioles, cutting off the petioles and tips of the cotyledons, cutting the leaves into 2-3 sections, flatly placing the cut cotyledons on a KCMS culture medium paved with sterilized filter paper, sealing a culture dish by using a sealing film to prevent bacteria and fungi from invading, and placing the culture dish in a tissue culture room for dark culture for 1 d;

infection: pouring the 0.2MS suspension after high-temperature sterilization into a culture dish, and then suspending the cotyledons cultured in the dark for one day in a 0.2MS liquid culture medium; finally, 100-300uL of agrobacterium tumefaciens suspension with OD600=0.1-0.3 is added, the culture dish is covered and is rotated, and the infection time is 3-4 min; after infection, pouring out bacterial liquid, sucking the bacterial liquid on cotyledons by using sterile filter paper, putting the infected cotyledons back on the original KCMS culture medium, and performing dark culture for 2d in a tissue culture room;

regeneration: transferring the infected cotyledons to a corresponding 2Z culture medium for regeneration, enabling the right sides of the cotyledons to face upwards, uniformly spacing the cotyledons, and culturing in a tissue culture room; when the infected tomato cotyledons grow on a 2Z culture medium for 10-14 days, callus is generated and bud points are formed; more bud points are generated on the callus at 14-20 days, and in order to reduce the bud point aberration rate, the callus generating the bud points is transferred to a culture medium containing corresponding 0.2Z for continuous culture;

obtaining and transplanting transgenic plants: growing the bud point on a 0.2Z culture medium to form a1 cm-long bud, selecting the bud with a complete growing point, cutting off the bud from the stem base part, and performing rooting culture on an R culture medium; culturing for one week, and rooting the transgenic plant; transplanting the seedling into a square pot filled with nutrient soil and vermiculite when the root grows to 5-6cm, covering and moisturizing for 3-4d, gradually uncovering, and irrigating with nutrient water; thus obtaining T0 generation SlABIG1 gene tomato.

2. Use of a method of creating tomato dwarf material by gene editing according to claim 1, characterized in that: is used for cultivating tomato plant types.

Images