CN115894642A

CN115894642A - Fruit type control gene SlGT-2 and homologous gene and application thereof

Info

Publication number: CN115894642A
Application number: CN202110955324.0A
Authority: CN
Inventors: 李传友; 朱强; 邓磊; 杨天霞; 蒋红玲
Original assignee: Institute of Genetics and Developmental Biology of CAS
Current assignee: Institute of Genetics and Developmental Biology of CAS
Priority date: 2021-08-19
Filing date: 2021-08-19
Publication date: 2023-04-04
Anticipated expiration: 2041-08-19
Also published as: CN115894642B

Abstract

The invention discloses a fruit type regulating SlGT-2 protein and a homologous SlGTL1 protein thereof. The invention adopts CRISPR/Cas9 technology, edits the SlGT-2 gene and the SlGTL1 gene at fixed points, knocks out the SlGT-2 gene and the SlGTL1 gene of tomatoes by causing frameshift mutation, and converts round juicy tomato materials into square juicy tomato materials. The invention has great application and popularization values.

Description

Fruit type control gene SlGT-2 and homologous gene and application thereof

Technical Field

The invention relates to a fruit type control gene SlGT-2 in the field of biotechnology and a homologous gene and application thereof.

Background

Tomato is one of the largest vegetables consumed worldwide and is also an important fruit. The fruit type is the most intuitive character of tomato fruits, and not only influences the selection of consumers, but also influences the taste or nutritional quality of the fruits. The fruits of various crops are mostly round, oval, oblate and long, and the fruits are rarely square. The square is an extremely rare shape, has great attraction to consumers and also has great demand on production, but the cultivation of the square fruit is extremely difficult, and a square fruit regulatory gene is not cloned at present.

Disclosure of Invention

The technical problem to be solved by the invention is how to regulate the shape of the plant fruit and how to regulate the water content of pectin.

In order to solve the above technical problems, it is a first object of the present invention to provide a protein which is SlGT-2 and/or its homologous protein SlGTL1;

the SlGT-2 is the protein of the following A1), A2) or A3):

a1 Protein of which the amino acid sequence is a sequence 1 in a sequence table;

a2 Protein which is obtained by substituting and/or deleting and/or adding more than one amino acid residue of an amino acid sequence shown in any one sequence 1 in a sequence table, is derived from the protein shown in A1) and is related to the shape of the plant fruit;

a3 A fusion protein obtained by attaching a protein tag to the N-terminus or/and the C-terminus of A1) or A2);

the SlGTL1 is the protein of A4), A5) or A6) as follows:

a4 Protein of which the amino acid sequence is the sequence 2 in the sequence table;

a5 Protein which is derived from the protein shown in A4) and is related to the shape of the plant fruit, and is obtained by substituting and/or deleting and/or adding more than one amino acid residue in the amino acid sequence shown in any one of the sequences 2 in the sequence table;

a6 A fusion protein obtained by attaching a protein tag to the N-terminus or/and C-terminus of A5) or A6).

Wherein, the sequence 1 in the sequence table is composed of 654 amino acid residues, and the sequence 2 in the sequence table is composed of 651 amino acid residues.

The protein is derived from tomato (Lycopersicon esculentum).

The protein can be artificially synthesized, or can be obtained by synthesizing the coding gene and then carrying out biological expression.

In the above protein, the protein tag (protein-tag) refers to a polypeptide or protein that is expressed by fusion with a target protein using in vitro recombinant DNA technology, so as to facilitate the expression, detection, tracking and/or purification of the target protein. The protein tag may be a Flag tag, a His tag, an MBP tag, an HA tag, a myc tag, a GST tag, and/or a SUMO tag, among others.

In the protein, the protein A2) is SlGT-2+T, and the amino acid sequence of SlGT-2+T is a sequence 9 in a sequence table; a5 The protein is SlGTL1+ AG, and the amino acid sequence of the SlGTL1+ AG is a sequence 10 in a sequence table.

The invention also provides a gene which is an SlGT-2 gene encoding the SlGT-2 or an SlGTL1 gene encoding the SlGTL1, or an SlGT-2+T gene encoding the SlGT-2+T or an SlGTL1+ AG gene encoding the SlGTL1+ AG;

the SlGT-2 gene is a gene shown in the following B1) or B2):

b1 Code sequence (CDS) of the coding strand is a cDNA molecule or a DNA molecule of sequence 3 in the sequence listing;

b2 The nucleotide of the coding strand is a DNA molecule of sequence 5 in the sequence table.

Wherein, the sequence 3 in the sequence table consists of 1965 nucleotides and codes the protein shown in the sequence 1 in the sequence table.

The SlGTL1 gene is a gene shown in the following B3) or B4):

b3 Code sequence (CDS) of the coding strand is a cDNA molecule or a DNA molecule of sequence 4 in the sequence listing;

b4 Nucleotide of coding strand is DNA molecule of sequence 6 in the sequence table.

Wherein, the sequence 4 in the sequence table consists of 1956 nucleotides and codes the protein shown in the sequence 2 in the sequence table.

The nucleotide of the coding chain of the SlGT-2+T gene is a DNA molecule of a sequence 11 in a sequence table;

the nucleotide of the coding chain of the SlGTL1+ AG gene is a DNA molecule of a sequence 12 in a sequence table.

The invention also provides a method for regulating and controlling the shape of the plant fruit, which comprises the following steps: and inhibiting the expression of the SlGT-2 gene and the SlGTL1 gene in the receptor round fruit plant to obtain the target plant of the square fruit.

In the method, the inhibition of the expression of the SlGT-2 gene of claim 2 and the SlGTL1 gene of claim 2 in the recipient round fruit plant is achieved by gene editing of the SlGT-2 gene and the SlGTL1 gene in the plant. The gene editing is realized by means of a CRISPR/Cas9 system.

In the method, the CRISPR/Cas9 system comprises a plasmid for expressing Cas9 and sgRNA, wherein the sgRNA can be sgRNA1 aiming at a target sequence 1 and sgRNA2 aiming at a target sequence 2, the target sequence 1 can be the 40 th to 59 th positions of a sequence 5 in a sequence table, and the target sequence 2 can be the 116 th to 136 th positions of a sequence 6 in the sequence table.

In the method, the nucleotide sequence of the sgRNA1 is shown as a sequence 7 in a sequence table, and the nucleotide sequence of the sgRNA2 is shown as a sequence 8 in the sequence table.

In the above method, the target plant is a plant satisfying the following conditions: a plant mutated in both the target sequence 1 and target sequence 2 regions.

In the above method, the expression of the SlGT-2 gene and the SlGTL1 gene in the receptor cone plant is X1 and X2 as follows:

the SlGT-2 gene shown in a sequence 5 in a sequence table is mutated into a SlGT-2+T gene by X1, and the coding sequence of the SlGT-2+T gene is shown as a sequence 11;

and X2, the SlGTL1 gene shown in a sequence 6 in the sequence table is mutated into a SlGTL1+ AG gene, and a coding sequence of the SlGTL1+ AG gene is shown in a sequence 12.

In the method, the amino acid sequence of the mutant SlGT-2 protein in the target plant is shown as a sequence 9, and the coding sequence is shown as a sequence 11; the amino acid sequence of the mutant SlGTL1 protein is shown as a sequence 10, and the coding sequence is shown as a sequence 12.

In the method, the target plant can be a plant of which the SlGT-2 gene and the SlGTL1 gene are homozygous mutated.

The plant described above may be any of F1) -F4):

f1 Tubular plants of the order florida;

f2 ) plants of the Solanaceae family;

f3 ) plants of the genus Lycopersicon;

f4 ) tomatoes.

In order to solve the technical problems, the invention also provides a reagent for regulating and controlling the fruit type of the plant, wherein the active component of the reagent is a substance for inhibiting the expression of the genes coding the SlGT-2 and the SlGTL1, reducing the abundance of the SlGT-2 protein and the SlGTL1 protein and/or knocking out the SlGT-2 gene and the SlGTL1 gene.

In the above reagent, the substance contains the following F1), F2) or F3):

f1 sgRNA, siRNA, shRNA, miRNA, or antisense RNA targeting the gene;

f2 A DNA molecule that produces a sgRNA that targets the gene, a DNA molecule that produces an siRNA that targets the gene, a DNA molecule that produces an shRNA that targets the gene, a DNA molecule that produces a miRNA that targets the gene, or a DNA molecule that produces an antisense RNA that targets the gene;

f3 An expression vector that generates sgrnas targeting the gene, an expression vector that generates sirnas targeting the gene, an expression vector that generates shrnas targeting the gene, an expression vector that generates mirnas targeting the gene, or an expression vector that generates antisense RNAs targeting the gene.

The active ingredients of the above agents may also contain other biological or/and non-biological components, and the other active ingredients of the above agents can be determined by those skilled in the art according to the fruit type of the plant.

The invention also provides application of the protein, the gene or the reagent in regulating and controlling the shape of the fruit and/or the water content of pectin of a plant.

The invention also provides application of the method in regulating and controlling the water content of plant pectin.

The inventor of the invention utilizes a fruit type-related protein SlGT-2 and a homologous protein SlGTL1 thereof which are separated and cloned from tomatoes in a laboratory, edits the SlGT-2 gene and the SlGTL1 gene at fixed points by adopting a CRISPR/Cas9 technology, knocks out the SlGT-2 gene and the SlGTL1 gene in the tomatoes by causing frameshift mutation, and can convert round juicy tomato materials into square juicy tomato materials. The invention has great application and popularization values.

Drawings

FIG. 1 is a photograph of the appearance of the 1-SlGT-2-SlGTL1 gene editing plant and the control wild type tomato variety AC fruit in example 1 of the present invention.

FIG. 2 is a photograph of a cut-through of the 1-SlGT-2-SlGTL1 gene editing plant and a control wild type tomato variety AC fruit in example 1 of the present invention.

FIG. 3 shows the measured positions of the width of the top 5% position of the fruit in the longitudinal direction and the width of the middle position of the fruit in the longitudinal direction according to the calculation formula of the present invention. Wherein, the first horizontal line from top to bottom indicates the position of 5% of the top end of the fruit in the longitudinal direction, and the second horizontal line from top to bottom indicates the position of the middle of the fruit in the longitudinal direction.

FIG. 4 is a statistical result chart of AC fruit orientation of 1-SlGT-2-SlGTL1 gene editing plants and control wild type tomato varieties in example 1 of the present invention. Data shown are mean ± sd, repeat number 15, and significance differences were analyzed in each group as t-test, representing significance analysis results P <0.01.

Detailed Description

The present invention is described in further detail below with reference to specific embodiments, which are given for the purpose of illustration only and are not intended to limit the scope of the invention. The examples provided below serve as a guide for further modifications by a person skilled in the art and do not constitute a limitation of the invention in any way.

In the quantitative tests in the following examples, three replicates were set up and the results averaged.

The experimental procedures in the following examples are conventional unless otherwise specified. Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.

The tomato variety AC (Ailsa Craig) in the following examples is the American tomato genetic resource center product (TGRC, http:// TGRC. Ucdavis. Edu /), numbered LA2838A.

The CRISPR/Cas9 vector pTX041 in the following examples is described in non-patent document "Deng et al," Efficient generation of pin-free protocols using CRISPR/Cas9 system. Journal of Genetics and Genomics 45 (2018) 51-54, "publicly available from the institute of Genetics and developmental biology of the Chinese academy of sciences, to repeat the experiments of the present application, and is not useful for other applications.

The preparation method of the seed growth culture medium in the following examples is (1L is an example): dissolving 2.2g of MS salt and 10g of cane sugar in water to a constant volume of 1L, adjusting the pH value to 5.8-6.0 by using 1mol/L KOH, adding 8g of agar, and sterilizing under high pressure.

The preparation method of the preculture medium in the following examples was as follows (1L as an example): dissolving 4.4g of MS salt, 1.0mg of Zeatin (Zeatin) and 30g of sucrose in water to a constant volume of 1L, adjusting the pH value to 5.8-6.0 by using 1mol/L KOH, adding 8g of agar, and sterilizing under high pressure.

The preparation method of the liquid MS culture medium in the following examples is as follows (1L as an example): 4.4g of MS salt and 30g of cane sugar are dissolved in water to be constant volume to 1L, the pH value is adjusted to be between 5.8 and 6.0 by using 1mol/L KOH, and the mixture is sterilized by high pressure.

The preparation method of the differentiation medium for screening in the following examples was as follows (1L for example): 4.4g of MS salt, 2.0mg of zeatin, 50mg of kanamycin, 100mg of inositol, 0.5mg of folic acid and 30g of sucrose are dissolved in water to be constant volume to 1L, the pH is adjusted to be between 5.8 and 6.0 by using 1mol/L of KOH, 8g of agar is added, and autoclaving is carried out.

The preparation method of the rooting medium in the following examples is as follows (1L is an example): dissolving 4.4g of MS salt, 50mg of kanamycin, 0.5mg of folic acid, 0.5mg of indolebutyric acid and 30g of sucrose in water to constant volume of 1L, adjusting the pH value to between 5.8 and 6.0 by using 1mol/L of KOH, adding 8g of agar, and autoclaving.

Example 1 editing tomato SlGT-2 gene and its homologous gene SlGTL1 by CRISPR/Cas9 method to obtain tomato with square juicy fruit

The inventor finds the SlGT-2 protein and the homologous protein SlGTL1 thereof in tomato for the first time, and the protein controls the formation of square fruits. The amino acid sequence of the SlGT-2 protein is sequence 1 in a sequence table, the gene coding the SlGT-2 protein is a SlGT-2 gene, the nucleotide sequence of the SlGT-2 gene between an initiation codon and a termination codon in genomic DNA is sequence 5 in the sequence table (wherein, the 1 st to 284 th sites and the 1037 th to 2717 th sites are exons), and the open reading frame in cDNA is sequence 3 in the sequence table. The amino acid sequence of the SlGTL1 protein is sequence 2 in a sequence table, the gene coding the SlGTL1 protein is a SlGTL1 gene, the nucleotide sequence of the SlGTL1 gene between an initiation codon and a termination codon in genomic DNA is sequence 6 in the sequence table (wherein, the 1 st to 320 th site and the 729 th to 2364 th site are exons), and the open reading frame in cDNA is sequence 4 in the sequence table.

1. Construction of recombinant vectors

1. Selecting a target sequence: selecting the 40 th-59 th site of a sequence 5 in a sequence table as a target sequence 1 aiming at the SlGT-2 gene; aiming at the SlGTL1 gene, the 116 th to 136 th sites of a sequence 6 in a sequence table are selected as a target sequence 2.

Two PCR primers 1 and 2 are synthesized, then a target fragment containing two gRNA and U6 promoter sequences is amplified from the template plasmid pTX041 vector, and the target fragment is purified and recovered.

The nucleotide sequences of the two PCR primers F and R are respectively as follows:

F:5’-ATATAT

GTTTGGGTGATAATCCAGAAAGCGGGTTTTAGAGCTAGAAATAGC-3' (the underlined sequence binds specifically to positions 40-59 of sequence 5 in the sequence table, and the wavy line indicates a BsaI enzyme recognition site);

R:5’-ATTATT

GAAACACCACTCTCCGGAGCTTCTTCAAACTACACTGTTAGATTC-3' (the underlined sequence specifically binds to positions 116-136 of sequence 6 in the sequence listing, and the wavy line indicates the BsaI enzyme recognition site).

2. And (3) carrying out enzyme digestion on the recovered target fragment by using restriction enzyme BsaI, purifying and recovering to obtain the enzyme digestion target fragment. And carrying out enzyme digestion on the CRISPR/Cas9 empty vector pTX041 by using restriction enzyme BsaI, purifying and recovering to obtain a linearized pTX041 vector skeleton.

3. And (3) connecting the enzyme digestion target fragment obtained in the step (2) with a pTX041 vector framework to obtain a recombinant plasmid, wherein the enzyme digestion target fragment is inserted between BsaI enzyme digestion sites of the pTX041 vector, and other sequences of the pTX041 vector are kept unchanged. Through sequencing verification, the recombinant plasmid expresses sgRNA1 (aiming at a target sequence 1) shown in a sequence 7 in a sequence table and sgRNA2 (aiming at a target sequence 2) shown in a sequence 8 in the sequence table, and the recombinant plasmid is named as pTX041-sgRNA1-sgRNA2.

The sgRNA1 aims at a target sequence 1 (40 th to 59 th positions of a sequence 5 in a sequence table), and a target sequence binding region in the sgRNA1 is shown as 2 nd to 21 nd nucleotides of a sequence 7 in the sequence table.

The sgRNA2 aims at a target sequence 2 (116 th to 136 th in a sequence 6 in a sequence table), and a target sequence binding region in the sgRNA1 is shown as nucleotides 2 nd to 21 th in a sequence 8 in the sequence table.

2. Preparation of Gene-edited plants

1. And (3) introducing the recombinant plasmid pTX041-sgRNA1-sgRNA2 prepared in the step one into agrobacterium GV3101 to obtain a recombinant agrobacterium, which is named as GV3101-sgRNA1-sgRNA2.

2. Taking recombinant agrobacterium GV3101-sgRNA1-sgRNA2, and carrying out genetic transformation on a wild tomato variety AC by an agrobacterium-mediated method to obtain a T0 generation plant, wherein the specific process comprises the following steps:

(1) Seeds of wild type tomato variety AC were selected, filled and large seeds were soaked with 75% aqueous ethanol for 2min, then soaked with 10% aqueous NaClO solution for 10min, then rinsed 7 times with sterile water, and then the seeds were sown in seed growth medium and cultured for 8 days. Taking cotyledons, cutting the cotyledons into small blocks under the aseptic condition, inoculating the cotyledons blocks into a pre-culture medium and culturing for 2 days to obtain cotyledons blocks serving as explants.

(2) Taking recombinant agrobacterium GV3101-sgRNA1-sgRNA2, and re-suspending with 50mL liquid MS culture medium to obtain OD _600nm And (3) adding 50 mu L of 0.074mol/L acetosyringone aqueous solution into the bacterial suspension, namely the infection solution.

(3) And (3) taking the explant obtained in the step (1), immersing the explant in the staining solution obtained in the step (2) for 10min, and then inoculating the explant on a pre-culture medium for culturing for 2 days.

(4) After step (3) is completed, the explants are taken and inoculated into a screening differentiation medium for 8 weeks (subculture every 2 weeks), and then the explants grow resistant buds.

(5) And (4) when the resistant bud length is 3cm, cutting the resistant bud, transferring the resistant bud to a rooting culture medium for culture, wherein the rooted plant is the T0 generation plant.

Culture conditions of the whole process: at 25 ℃ for 16h light/8 h dark.

3. And screening out plants in which the SlGT-2 gene and the homologous gene SlGTL1 thereof are homozygously mutated from the T0 generation plants.

The screening method comprises the following steps: taking the genome DNA of the plant leaf, respectively carrying out PCR amplification by using two pairs of primers, namely a primer pair 1 (consisting of F1 and R1) and a primer pair 2 (consisting of F2 and R2), recovering PCR amplification products and sequencing.

F1：5′-GGTGTAATTGCTAACTTGCTTGG-3′；

R1：5′-GCATGATGAAACTGGTGCAGC-3′；

F2：5′-ATGCTTGGTGTTTCTTCAAG-3′；

R2：5′-AACTTCTTCCCATAACGGTCC-3′。

Screening 1 plant of which the SlGT-2 gene and the homologous gene SlGTL1 thereof are homozygously mutated, and naming the plant as a plant 1.

Compared with a wild tomato variety AC, the SlGT-2 gene in the plant 1 has a mutation of inserting one nucleotide, specifically, a nucleotide T is inserted between the 56 th site and the 57 th site of the sequence 5 in the sequence table, and the mutation is homozygosis mutation (namely, the two chromosomes have the same mutation). The insertion causes the CDS sequence of the gene to generate frame shift mutation from the 57 th position, a stop codon is generated in advance, and a truncated protein losing a DNA binding structural domain is translated, so that the function of the SlGT-2 protein is lost (the plant does not contain the SlGT-2 protein). The mutated gene is named as SlGT-2+T gene, the protein coded by SlGT-2+T gene is SlGT-2+ T, and the amino acid sequence of SlGT-2+T is sequence 9. Meanwhile, compared with a wild tomato variety AC, the SlGTL1 gene in the plant 1 has mutation of inserting two nucleotides, specifically, two nucleotides of AG are inserted between the 131 th site and the 132 th site of the sequence 6 in the sequence table, and the mutation is homozygous (namely, the two chromosomes have the same mutation). The insertion causes the CDS sequence of the gene to generate frame shift mutation from the 132 th position, a stop codon is generated in advance, and a truncated protein losing a DNA binding structural domain is translated, so that the function of the SlGTL1 protein is lost (the SlGTL1 protein is not contained in a plant). The mutated gene is named as SlGTL1+ T gene, the protein coded by the SlGTL1+ T gene is SlGT-2+ T, and the amino acid sequence of SlGT-2+T is sequence 10.

That is, compared with the wild tomato variety AC, for the SlGT-2 gene, the SlGT-2 genes in two homologous chromosomes are mutated into the SlGT-2+T gene, and the coding sequence of the SlGT-2+T gene is a sequence 11; for the SlGTL1 gene, both SlGTL1 genes in two homologous chromosomes are mutated into SlGTL1+ AG genes, and the coding sequence of the SlGTL1+ AG gene is a sequence 12.

4. Normally culturing the plant 1, selfing to obtain seeds, namely T1 generation seeds, and culturing the T1 generation seeds into a plant group, namely a T1 generation plant 1 group.

5. Plants without exogenous DNA were selected from the T1 generation plant 1 population.

The screening method comprises the following steps: separately extracting genome DNA from leaves of each individual plant of the T1 generation plant 1 group, performing PCR amplification by using a primer pair consisting of F3 and R3 (the target sequence of the primer pair consisting of F3 and R3 is located in the Cas9 gene in the CRISPR/Cas9 vector pTX041, and the amplification product is expected to be about 402 bp), wherein if the amplification product is obtained, the plant contains exogenous DNA, and if the amplification product is not obtained, the plant does not contain the exogenous DNA.

F3：5′-TTGACAAGCTGTTCATCCAG-3′；

R3：5′-CCTTCGTAATCTCGGTGTTC-3′。

About 1/4 of the individuals in the T1 generation plant 1 population did not contain exogenous DNA.

6. And (4) taking the plants which are obtained by screening in the step (5) and do not contain the exogenous DNA, extracting the genome DNA of the leaves, respectively adopting primers consisting of F1 and R1 and F2 and R2 to carry out PCR amplification, recovering PCR amplification products and sequencing.

The plants which are obtained by screening from the T1 generation plant 1 population and do not contain exogenous DNA have homozygotic insertion which is the same as that of the plant 1 in the SlGT-2 gene and the homologous gene SlGTL1 thereof. The result shows that the mutation generated by introducing the recombinant plasmid pTX041-sgRNA1-sgRNA2 prepared in the step one into the wild type tomato variety AC can be stably inherited from the T0 generation to the T1 generation.

Plants which are obtained by screening from the T1 generation plant 1 group and do not contain exogenous DNA are named as 1-SlGT-2-SlGTL1 gene editing plants.

3. Fruit inspection

The test plants were: wild type tomato variety AC, 1-SlGT-2-SlGTL1 gene editing plant.

1. The seedlings of the tested plants were cultured at 25 ℃ under 16h light/8 h dark conditions until the plants grew to 4-5 leaves.

2. After step 1 was completed, plants (10 plants per test plant and grown consistently) were transplanted into the greenhouse. The plant spacing and the row spacing are more than 60 cm; randomly distributing tested plants; normal water and fertilizer management ensures that the water and fertilizer conditions of all plants are basically consistent.

After 3 months of transplanting, the plants begin to enter the fruiting period, which lasts about 6 months. And collecting mature fruits in the whole fruiting period.

1-SlGT-2-SlGTL1 gene editing plant, wherein the number of mature fruits harvested in the whole fruiting period of each plant is 55 on average, and the mature fruits are square fruits.

Wild type tomato variety AC, on average 58 mature fruits per plant were harvested over the whole fruiting period, all round fruits.

An exemplary fruit profile of a part of the fruit is shown in figure 1.

An exemplary fruit slit of a part of the fruit is shown in figure 2.

The formula for calculating the squareness is as follows:

squareness = width at 5% position of top end of fruit in longitudinal direction/width at middle position of fruit in longitudinal direction (see fig. 3).

The fruit squareness statistics for each test plant are shown in figure 4.

Compared with the fruit of the wild tomato variety AC, the fruit of the 1-SlGT-2-SlGTL1 gene editing plant is changed from a round shape to a square shape, and the water content of pectin is changed from more to less.

The results show that the SlGT-2 gene and the homologous gene SlGTL1 thereof can regulate the fruit type of the tomato, and round juicy tomato materials can be converted into square juicy tomato materials by carrying out gene editing on the SlGT-2 gene and the homologous gene SlGTL1 thereof.

In conclusion, the inventor discovers the SlGT-2 protein and the homologous protein SlGTL1 thereof in tomato for the first time, the protein controls the formation of square fruits, round fruits can be changed into squares after the genes are knocked out by utilizing a non-transgenic gene editing technology, and meanwhile, the pectin state, particularly the water content, is obviously changed. The target gene provided by the invention can be rapidly cultured into square fruits by using a gene editing technology, and only a half year to a year is needed in tomatoes.

The present invention has been described in detail above. It will be apparent to those skilled in the art that the invention can be practiced in a wide range of equivalent parameters, concentrations, and conditions without departing from the spirit and scope of the invention and without undue experimentation. While the invention has been described with reference to specific embodiments, it will be appreciated that the invention can be further modified. In general, this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains. The use of some of the essential features is possible within the scope of the claims attached below.

Sequence listing

<110> institute of genetics and developmental biology of Chinese academy of sciences

<120> fruit type control gene SlGT-2 and homologous gene and application thereof

<130> GNCSY212244

<160> 12

<170> SIPOSequenceListing 1.0

<210> 1

<211> 654

<212> PRT

<213> tomato (Lycopersicon esculentum)

<400> 1

Met Leu Gly Val Ser Gly Leu Val Ser Ser Glu Gly Gly Gly Asp Asn

1 5 10 15

Pro Glu Ser Gly Gly Gly Ala Gly Ser Gly Gly Ser Ser Glu Ile Gly

20 25 30

Leu Gly Gly Gly Ser Gly Gly Gly Gly Gly Ser Ser Gly Gly Phe Met

35 40 45

Thr Glu Asp Gly Glu Arg Asn Ser Gly Gly Asn Arg Trp Pro Arg Gln

50 55 60

Glu Thr Ile Ala Leu Leu Lys Ile Arg Ser Glu Met Asp Val Ile Phe

65 70 75 80

Arg Asp Ser Ser Leu Lys Gly Pro Leu Trp Glu Glu Val Ser Arg Lys

85 90 95

Met Ala Asp Leu Gly Phe His Arg Ser Ser Lys Lys Cys Lys Glu Lys

100 105 110

Phe Glu Asn Val Tyr Lys Tyr His Lys Arg Thr Lys Asp Gly Arg Ala

115 120 125

Ser Lys Ala Asp Gly Lys Asn Tyr Arg Phe Phe Glu Gln Leu Glu Ala

130 135 140

Leu Glu Asn Ile Thr Ser His His Ser Leu Met Pro Val Pro Ser Ser

145 150 155 160

Asn Thr Arg Pro Pro Pro Pro Pro Leu Glu Ala Thr Pro Ile Asn Met

165 170 175

Ala Met Pro Met Ala Ser Ser Asn Val Gln Val Thr Ala Ser Gln Gly

180 185 190

Thr Ile Pro His His Val Thr Ile Ser Ser Ala Pro Pro Pro Pro Asn

195 200 205

Ser Leu Phe Ala Pro Ser His Gln Asn Ala Pro Ser Ser Ser Pro Val

210 215 220

Pro Leu Pro Pro Pro Pro Ser Gln Gln Pro Ser Pro Gln Pro Ala Val

225 230 235 240

Asn Pro Ile Asn Asn Ile Pro Gln Gln Val Asn Ala Ser Ala Met Ser

245 250 255

Tyr Ser Thr Ser Ser Ser Thr Ser Ser Asp Glu Asp Ile Gln Arg Arg

260 265 270

His Lys Lys Lys Arg Lys Trp Lys Asp Tyr Phe Glu Lys Phe Thr Lys

275 280 285

Asp Val Ile Asn Lys Gln Glu Glu Ser His Arg Arg Phe Leu Glu Lys

290 295 300

Leu Glu Lys Arg Glu His Asp Arg Met Val Arg Glu Glu Ala Trp Lys

305 310 315 320

Val Glu Glu Met Ala Arg Met Asn Arg Glu His Asp Leu Leu Val Gln

325 330 335

Glu Arg Ala Met Ala Ala Ala Lys Asp Ala Ala Val Ile Ser Phe Leu

340 345 350

Gln Lys Ile Thr Glu Gln Gln Asn Ile Gln Ile Pro Asn Ser Ile Asn

355 360 365

Val Gly Pro Pro Ser Ala Gln Val Gln Ile Gln Leu Pro Glu Asn Pro

370 375 380

Leu Ser Ala Pro Val Pro Thr Gln Ile Gln Pro Thr Thr Val Thr Ala

385 390 395 400

Ala Ala Pro Pro Gln Pro Ala Pro Val Pro Val Ser Leu Pro Val Thr

405 410 415

Ile Pro Ala Pro Val Pro Ala Leu Ile Pro Ser Leu Ser Leu Pro Leu

420 425 430

Thr Pro Pro Val Pro Ser Lys Asn Met Glu Leu Val Pro Lys Ser Asp

435 440 445

Asn Gly Gly Asp Ser Tyr Ser Pro Ala Ser Ser Ser Arg Trp Pro Lys

450 455 460

Ala Glu Val Glu Ala Leu Ile Lys Leu Arg Thr Asn Leu Asp Val Lys

465 470 475 480

Tyr Gln Glu Asn Gly Pro Lys Gly Pro Leu Trp Glu Glu Ile Ser Ser

485 490 495

Gly Met Lys Lys Ile Gly Tyr Asn Arg Asn Ala Lys Arg Cys Lys Glu

500 505 510

Lys Trp Glu Asn Ile Asn Lys Tyr Phe Lys Lys Val Lys Glu Ser Asn

515 520 525

Lys Lys Arg Pro Glu Asp Ser Lys Thr Cys Pro Tyr Phe His Gln Leu

530 535 540

Asp Ala Leu Tyr Lys Glu Lys Ala Lys Asn Pro Glu Thr Ala Ser Ser

545 550 555 560

Thr Ser Ser Phe Asn Pro Ser Phe Ala Leu Asn Pro Asp Asn Asn Gln

565 570 575

Met Ala Pro Ile Met Ala Arg Pro Glu Gln Gln Trp Pro Leu Pro Gln

580 585 590

His His Glu Ser Thr Thr Arg Ile Asp His Glu Asn Glu Ser Asp Asn

595 600 605

Met Asp Glu Asp Asp His Asp Asp Glu Glu Asp Glu Asp Asp Glu Asp

610 615 620

Glu Asn Asn Ala Tyr Glu Ile Val Ala Asn Lys Gln Gln Ser Ser Met

625 630 635 640

Ala Ala Ala Asn Thr Thr Thr Ser Thr Ala Thr Thr Thr Val

645 650

<210> 2

<211> 651

<212> PRT

<213> tomato (Lycopersicon esculentum)

<400> 2

Met Leu Gly Val Ser Ser Ser Leu Ile Ala Ser Ser Asn Thr Ser Ile

1 5 10 15

Thr Ala Gly Ala Ala Gly Asp Gly Ala Ala Ile Ser Ala Ala Pro Ser

20 25 30

Gln Leu Ala Pro Pro Pro Gln Glu Ala Pro Glu Ser Gly Gly Ser Ser

35 40 45

Glu Gly Gly Gly Gly Gly Gly Asp Leu Ser Ile Gly Gly Glu Asp Gly

50 55 60

Glu Arg Asn Ser Gly Gly Asn Arg Trp Pro Arg Gln Glu Thr Leu Ala

65 70 75 80

Leu Leu Lys Ile Arg Ser Glu Met Asp Val Val Phe Lys Asp Ser Ser

85 90 95

Leu Lys Gly Pro Leu Trp Glu Glu Val Ser Arg Lys Leu Ala Glu Leu

100 105 110

Gly Tyr His Arg Ser Ala Lys Lys Cys Lys Glu Lys Phe Glu Asn Val

115 120 125

Tyr Lys Tyr His Arg Arg Thr Lys Asp Gly Arg Ala Ser Lys Ala Asp

130 135 140

Gly Lys Thr Tyr Arg Phe Phe Asp Gln Leu Gln Ala Leu Glu Asn Asn

145 150 155 160

Pro Ser Ser His Ser Asn Ile Pro Pro Pro Pro Leu Ala Ala Thr Pro

165 170 175

Ile Thr Met Ala Met Pro Met Arg Ser Gly Asn Asn Ser Ala Asn Pro

180 185 190

Pro Met Pro Thr Pro Thr Pro Thr Pro Gln Asn His Asn His Phe Phe

195 200 205

Ser Val Ser Gln Lys Ser Val Val Thr Gly Ala Ala Gln Pro Ala Val

210 215 220

Met Thr Ala Pro Ala Leu Pro Leu Ser Gln Val Pro Ile Gly Asn Asn

225 230 235 240

Asn Leu Asn Gln Met His Arg Pro Gln Gly Asn Thr Thr Thr Thr Lys

245 250 255

Thr Ser Phe Leu Ser Asn Ser Thr Ser Ser Ser Ser Ser Thr Ser Ser

260 265 270

Asp Glu Asp Ile Gln Arg Arg Gln Met Lys Lys Arg Lys Trp Lys Glu

275 280 285

Phe Phe Glu Ser Leu Met Lys Asp Val Ile Glu Lys Gln Glu Glu Leu

290 295 300

Gln Lys Lys Phe Leu Glu Thr Leu Glu Lys Arg Glu Arg Asp Arg Leu

305 310 315 320

Met Arg Glu Glu Ala Trp Arg Val Gln Glu Met Ala Arg Leu Asn Arg

325 330 335

Glu His Asp Leu Leu Val Gln Glu Arg Ser Met Ala Ala Ala Lys Asp

340 345 350

Ala Thr Ile Ile Ala Phe Leu Gln Lys Ile Thr Glu Gln Gln Asn Thr

355 360 365

Gln Thr Pro Asn Ser Thr Asn Asn Thr Ser Pro Ser Pro Phe Pro Ile

370 375 380

Ala Gln Ile Gln Leu Lys Leu Ser Glu Lys Pro Phe Ser Thr Pro Pro

385 390 395 400

Gln Pro Gln Pro Gln Pro Ser Ala Thr Ala Val Ser Leu Pro Met Thr

405 410 415

Ile His Thr Pro Thr Pro Ala Pro Pro Gln Thr Leu Thr Leu Pro Val

420 425 430

Val Ser Ser Lys Ser Leu Glu Pro Pro Lys Ser Asp Asn Gly Gly Glu

435 440 445

Asn Phe Ser Pro Ala Ser Ser Ser Arg Trp Pro Lys Glu Glu Ile Glu

450 455 460

Ala Leu Ile Ser Leu Arg Thr Cys Leu Asp Leu Lys Tyr Gln Glu Asn

465 470 475 480

Gly Pro Lys Gly Pro Leu Trp Glu Glu Ile Ser Ser Gly Met Arg Lys

485 490 495

Ile Gly Tyr Asn Arg Asn Ala Lys Arg Cys Lys Glu Lys Trp Glu Asn

500 505 510

Ile Asn Lys Tyr Phe Lys Lys Val Lys Glu Ser Asn Lys Lys Arg Pro

515 520 525

Glu Asp Ser Lys Thr Cys Pro Tyr Phe His Gln Leu Glu Ala Leu Tyr

530 535 540

Lys Glu Lys Ala Lys Leu Glu Pro Val Pro His Asn Thr Thr Phe Gly

545 550 555 560

Leu Thr Pro Gln Asn Asn Pro Pro Pro Pro Pro Pro Pro Ile Met Ala

565 570 575

Gln Pro Glu Gln Gln Trp Pro Ile Pro Gln Asn Gln Leu His Gln Gln

580 585 590

Asn Arg Asp His His His Asp Asn Glu Ser Asp Ser Met Asp His Asp

595 600 605

Leu Glu Glu Asp Glu Asp Glu Asp Glu Glu Asp Glu Gly Asn Gly Tyr

610 615 620

Glu Ile Ile Ile Thr Asn Lys Gln Gln Ser Ser Ser Met Ala Ala Thr

625 630 635 640

Pro Val Thr Thr Thr Thr Ser Ala Ala Ala Val

645 650

<210> 3

<211> 1965

<212> DNA

<213> tomato (Lycopersicon esculentum)

<400> 3

atgctgggcg tttccggctt agtaagtagt gaaggtggtg gtgataatcc agaaagcggt 60

ggaggagctg gaagcggagg gagtagtgag attggattag gcggtggaag tggcggcggt 120

ggaggtagta gcggtggatt catgacggaa gatggagaaa gaaattcagg tggaaataga 180

tggccaagac aagaaacaat tgctttgctg aaaataaggt ctgaaatgga tgttattttt 240

agagactcaa gtcttaaagg acctttatgg gaagaagttt ccaggaaaat ggcagacctt 300

gggttccaca gaagttccaa gaaatgcaag gagaagttcg aaaatgtata caaatatcac 360

aagagaacca aggatggccg agcatcgaaa gcggatggaa agaattatag gtttttcgag 420

caattggaag ccctggagaa cattacatct catcattctc taatgccagt accgtcgtct 480

aatacgcgtc ctccaccccc tccgttggaa gctactccaa taaatatggc tatgccaatg 540

gcatcatcaa atgtacaagt cacggcttca caaggtacta ttcctcatca tgttactatt 600

tcatcagcac caccgccacc gaatagcctt tttgctcctt ctcatcaaaa tgctccgtca 660

agttcacccg tgccactacc accaccgcca tcacagcaac catcaccgca gccagctgtc 720

aatccgatta ataatattcc tcaacaagtg aacgcttcag caatgtcgta ttcaacttct 780

tcgtctactt cctcggatga ggatatacaa agaaggcata agaagaagag gaaatggaag 840

gattattttg agaagttcac caaggatgtg attaataagc aggaggaatc gcacaggagg 900

ttcttggaga agcttgagaa gcgggaacat gatcggatgg ttcgagaaga agcatggaaa 960

gtagaggaaa tggcaaggat gaatagggag catgatcttt tagttcaaga aagagcaatg 1020

gcggcagcca aggatgcagc tgttatttct tttttacaaa agataactga acagcaaaac 1080

attcaaattc caaatagtat caacgttggc cctccatcag cacaagtaca aatacaattg 1140

cctgaaaacc cactatccgc gcctgtacca acacaaatac aaccgacgac tgttacagca 1200

gcagcaccac ctcaaccagc accggtccca gtatcgttgc cagtaacaat accagctcca 1260

gtaccagcat taataccatc attgtcgcta ccactgacac caccagtgcc atccaagaac 1320

atggagttag taccaaaaag cgataacgga ggtgatagtt acagtccagc aagctcttca 1380

aggtggccaa aagcagaagt tgaagcattg attaaacttc gtacaaattt agatgtcaaa 1440

taccaagaga acggacctaa aggtccactt tgggaagaga tatcatctgg aatgaagaaa 1500

attggataca atcggaatgc aaagagatgc aaagaaaaat gggaaaacat caacaaatac 1560

ttcaagaagg tgaaggagag caacaaaaaa cgacccgaag attccaaaac ttgcccatat 1620

ttccaccagc tcgatgcact gtacaaggag aaagccaaaa accccgaaac agcttcttca 1680

acgtcttcgt tcaatccttc attcgcttta aaccccgata acaaccaaat ggctcccatc 1740

atggctcgtc cagaacagca atggccactt ccacaacacc atgaaagcac cacccgtatc 1800

gaccacgaaa acgagagcga caacatggat gaagatgatc acgatgatga ggaggatgaa 1860

gatgacgagg acgaaaacaa cgcttatgag atagtagcaa acaagcaaca atcctcaatg 1920

gcggccgcaa acaccactac cagcaccgca acaacaacag tttga 1965

<210> 4

<211> 1956

<212> DNA

<213> tomato (Lycopersicon esculentum)

<400> 4

atgcttggtg tttcttcaag tttaatagct agcagtaata ctagtattac tgctggtgct 60

gcaggtgatg gagctgccat ttcggcagct ccatcacagt tagcaccgcc accacaagaa 120

gctccggaga gtggtgggag tagtgaaggt ggtggcggtg gaggagattt gtcgattggc 180

ggtgaagatg gagaaaggaa ctcaggtgga aatcgatggc caaggcaaga aactttagct 240

ttactgaaaa ttagatcgga aatggatgtt gttttcaaag attcaagtct taaaggaccg 300

ttatgggaag aagtttccag aaaactcgcg gagttgggtt atcatcgaag tgctaagaaa 360

tgtaaagaga aattcgagaa tgtttacaag tatcacagga gaaccaaaga tggtcgtgct 420

tcgaaagcag atggaaaaac ttatcgattc tttgatcagt tacaggcttt ggaaaacaat 480

ccatcttctc attctaacat accgccacct ccattagcag caacacccat aacaatggca 540

atgccaatgc gatcaggaaa caattcagca aatcctccaa tgccgacgcc aacgccaact 600

ccacaaaatc ataatcattt ttttagtgtt tcgcagaaaa gtgttgtgac aggagcagcg 660

cagcctgctg ttatgactgc acctgcgctg ccactgtcac aagtgccgat aggtaataat 720

aacttgaacc agatgcatcg gcctcaaggt aatactacta ctacaaaaac aagtttcctg 780

tcgaattcaa cttcatcatc atcttcaact tcgtcggatg aggatataca aaggaggcag 840

atgaagaagc ggaaatggaa ggaattcttt gagagtttaa tgaaggatgt gattgagaag 900

caagaggaat tgcagaagaa gtttttggaa acgctcgaga agcgcgagag ggataggttg 960

atgagagagg aggcatggag agtgcaagag atggctagat tgaataggga acatgatctt 1020

ttagtccaag agagatcaat ggcagcagct aaagacgcaa caatcatcgc cttcttgcaa 1080

aaaataactg aacagcaaaa cacacaaacc ccgaatagta caaataacac ttctccttct 1140

ccttttccaa ttgctcaaat tcaattaaaa ttgtccgaaa agccattcag tacaccacca 1200

caaccacaac cacaaccatc agctaccgcg gtatcactgc caatgacaat acatacacca 1260

acaccagcac caccacagac actgacatta cctgtagtat catcaaaatc acttgaacct 1320

ccaaaatccg ataatggtgg tgagaatttc tctccagcaa gctcgtcaag atggccgaaa 1380

gaagaaatcg aagcattgat aagtctccga acctgtttag atctaaaata ccaagaaaat 1440

ggaccgaaag gaccactgtg ggaagaaatt tcatctggaa tgagaaagat aggatacaac 1500

aggaatgcaa agagatgcaa ggaaaaatgg gagaacatca acaagtactt caagaaggta 1560

aaagaaagca acaaaaaaag accagaagat tccaaaactt gcccatattt ccaccagctg 1620

gaagcactgt acaaagaaaa agccaagctc gaacctgtac cacacaacac taccttcgga 1680

ttaacacccc aaaacaatcc tcctcctcct cctcctccca tcatggctca acccgagcaa 1740

caatggccaa ttcctcaaaa tcaacttcac cagcaaaatc gtgatcatca tcacgataat 1800

gaaagcgaca gcatggatca cgatttggaa gaggacgagg atgaggacga agaagatgaa 1860

ggtaatggct atgaaataat aatcacaaat aaacaacaat catcatcaat ggcggctacc 1920

ccagtaacaa caacaacttc tgctgctgca gtttaa 1956

<210> 5

<211> 2717

<212> DNA

<213> tomato (Lycopersicon esculentum)

<400> 5

atgctgggcg tttccggctt agtaagtagt gaaggtggtg gtgataatcc agaaagcggt 60

ggaggagctg gaagcggagg gagtagtgag attggattag gcggtggaag tggcggcggt 120

ggaggtagta gcggtggatt catgacggaa gatggagaaa gaaattcagg tggaaataga 180

tggccaagac aagaaacaat tgctttgctg aaaataaggt ctgaaatgga tgttattttt 240

agagactcaa gtcttaaagg acctttatgg gaagaagttt ccaggtaatt aaattcaatt 300

tcattattcc aatttcttca cctgaccttc tcaatcatta ttaagctgca ccagtttcat 360

catgcataaa taaaaattga tagaaatgga atctttattt aatttttttt ttcaatttct 420

acttttggga aaaaaataat taatagaatg atttttattt tttgggaaat gaaaagatag 480

atctatggat cagaattcca ttgatttatt gcttttttga ttaaaagggt tattgttttt 540

cagttcattt cactacaaac aatacaacaa aaatacaatt gttgaggaaa ttcagattcc 600

ctccttccgg gttttgagcc aaattcagtt ttgctttttt ggcgtttttt ctttctctgc 660

caattccagc aacaaatttt ggaaactaat ttactcatct tttttgtatt agagttccaa 720

ctttatgaac tacctttttt taaatttagc aaataaataa gtttggtaat catcaaatct 780

aataattaag caagtaaaaa aacaagattt atgattgaga aaaatgtggt ttccatagag 840

tgtttcaatt gtctcctact tgtttaatta attgatttct taattacctt aatcttgatt 900

aataatctca tttttatttt atgtggtgaa tagtatttta ctattgaatt caattaccaa 960

ggatttaaat tattgtactt gtttatttac taccattttt tctaatactt atgccaactg 1020

ttgttatcat gagcaggaaa atggcagacc ttgggttcca cagaagttcc aagaaatgca 1080

aggagaagtt cgaaaatgta tacaaatatc acaagagaac caaggatggc cgagcatcga 1140

aagcggatgg aaagaattat aggtttttcg agcaattgga agccctggag aacattacat 1200

ctcatcattc tctaatgcca gtaccgtcgt ctaatacgcg tcctccaccc cctccgttgg 1260

aagctactcc aataaatatg gctatgccaa tggcatcatc aaatgtacaa gtcacggctt 1320

cacaaggtac tattcctcat catgttacta tttcatcagc accaccgcca ccgaatagcc 1380

tttttgctcc ttctcatcaa aatgctccgt caagttcacc cgtgccacta ccaccaccgc 1440

catcacagca accatcaccg cagccagctg tcaatccgat taataatatt cctcaacaag 1500

tgaacgcttc agcaatgtcg tattcaactt cttcgtctac ttcctcggat gaggatatac 1560

aaagaaggca taagaagaag aggaaatgga aggattattt tgagaagttc accaaggatg 1620

tgattaataa gcaggaggaa tcgcacagga ggttcttgga gaagcttgag aagcgggaac 1680

atgatcggat ggttcgagaa gaagcatgga aagtagagga aatggcaagg atgaataggg 1740

agcatgatct tttagttcaa gaaagagcaa tggcggcagc caaggatgca gctgttattt 1800

cttttttaca aaagataact gaacagcaaa acattcaaat tccaaatagt atcaacgttg 1860

gccctccatc agcacaagta caaatacaat tgcctgaaaa cccactatcc gcgcctgtac 1920

caacacaaat acaaccgacg actgttacag cagcagcacc acctcaacca gcaccggtcc 1980

cagtatcgtt gccagtaaca ataccagctc cagtaccagc attaatacca tcattgtcgc 2040

taccactgac accaccagtg ccatccaaga acatggagtt agtaccaaaa agcgataacg 2100

gaggtgatag ttacagtcca gcaagctctt caaggtggcc aaaagcagaa gttgaagcat 2160

tgattaaact tcgtacaaat ttagatgtca aataccaaga gaacggacct aaaggtccac 2220

tttgggaaga gatatcatct ggaatgaaga aaattggata caatcggaat gcaaagagat 2280

gcaaagaaaa atgggaaaac atcaacaaat acttcaagaa ggtgaaggag agcaacaaaa 2340

aacgacccga agattccaaa acttgcccat atttccacca gctcgatgca ctgtacaagg 2400

agaaagccaa aaaccccgaa acagcttctt caacgtcttc gttcaatcct tcattcgctt 2460

taaaccccga taacaaccaa atggctccca tcatggctcg tccagaacag caatggccac 2520

ttccacaaca ccatgaaagc accacccgta tcgaccacga aaacgagagc gacaacatgg 2580

atgaagatga tcacgatgat gaggaggatg aagatgacga ggacgaaaac aacgcttatg 2640

agatagtagc aaacaagcaa caatcctcaa tggcggccgc aaacaccact accagcaccg 2700

caacaacaac agtttga 2717

<210> 6

<211> 2364

<212> DNA

<213> tomato (Lycopersicon esculentum)

<400> 6

atgcttggtg tttcttcaag tttaatagct agcagtaata ctagtattac tgctggtgct 60

gcaggtgatg gagctgccat ttcggcagct ccatcacagt tagcaccgcc accacaagaa 120

gctccggaga gtggtgggag tagtgaaggt ggtggcggtg gaggagattt gtcgattggc 180

ggtgaagatg gagaaaggaa ctcaggtgga aatcgatggc caaggcaaga aactttagct 240

ttactgaaaa ttagatcgga aatggatgtt gttttcaaag attcaagtct taaaggaccg 300

ttatgggaag aagtttccag gtactgtttt tttttggtca ttttgattaa ctctttcatc 360

atcatcatat gcatagaatt cagaaaatta aaagatctat tatatttgga attaaaattc 420

gtatttgatg gaaagtgtta ttttttttta gttttgttgt tataccattt tctgctgatt 480

ccagcaagaa atttaggatg agtttaattt ctctactcat cttcaacact ttttgtgctt 540

ttccctattt tccagaaaat tcaagctaag atgttgatga ttgatggttt attatgtttt 600

attttatgta tataaaaata gtatgagatt ttgttttttt attactaatg aatgatgaat 660

atgagatgag ataattaaga caaggtgttt ttctttttgt atccattttg aactttgttg 720

tttatcagaa aactcgcgga gttgggttat catcgaagtg ctaagaaatg taaagagaaa 780

ttcgagaatg tttacaagta tcacaggaga accaaagatg gtcgtgcttc gaaagcagat 840

ggaaaaactt atcgattctt tgatcagtta caggctttgg aaaacaatcc atcttctcat 900

tctaacatac cgccacctcc attagcagca acacccataa caatggcaat gccaatgcga 960

tcaggaaaca attcagcaaa tcctccaatg ccgacgccaa cgccaactcc acaaaatcat 1020

aatcattttt ttagtgtttc gcagaaaagt gttgtgacag gagcagcgca gcctgctgtt 1080

atgactgcac ctgcgctgcc actgtcacaa gtgccgatag gtaataataa cttgaaccag 1140

atgcatcggc ctcaaggtaa tactactact acaaaaacaa gtttcctgtc gaattcaact 1200

tcatcatcat cttcaacttc gtcggatgag gatatacaaa ggaggcagat gaagaagcgg 1260

aaatggaagg aattctttga gagtttaatg aaggatgtga ttgagaagca agaggaattg 1320

cagaagaagt ttttggaaac gctcgagaag cgcgagaggg ataggttgat gagagaggag 1380

gcatggagag tgcaagagat ggctagattg aatagggaac atgatctttt agtccaagag 1440

agatcaatgg cagcagctaa agacgcaaca atcatcgcct tcttgcaaaa aataactgaa 1500

cagcaaaaca cacaaacccc gaatagtaca aataacactt ctccttctcc ttttccaatt 1560

gctcaaattc aattaaaatt gtccgaaaag ccattcagta caccaccaca accacaacca 1620

caaccatcag ctaccgcggt atcactgcca atgacaatac atacaccaac accagcacca 1680

ccacagacac tgacattacc tgtagtatca tcaaaatcac ttgaacctcc aaaatccgat 1740

aatggtggtg agaatttctc tccagcaagc tcgtcaagat ggccgaaaga agaaatcgaa 1800

gcattgataa gtctccgaac ctgtttagat ctaaaatacc aagaaaatgg accgaaagga 1860

ccactgtggg aagaaatttc atctggaatg agaaagatag gatacaacag gaatgcaaag 1920

agatgcaagg aaaaatggga gaacatcaac aagtacttca agaaggtaaa agaaagcaac 1980

aaaaaaagac cagaagattc caaaacttgc ccatatttcc accagctgga agcactgtac 2040

aaagaaaaag ccaagctcga acctgtacca cacaacacta ccttcggatt aacaccccaa 2100

aacaatcctc ctcctcctcc tcctcccatc atggctcaac ccgagcaaca atggccaatt 2160

cctcaaaatc aacttcacca gcaaaatcgt gatcatcatc acgataatga aagcgacagc 2220

atggatcacg atttggaaga ggacgaggat gaggacgaag aagatgaagg taatggctat 2280

gaaataataa tcacaaataa acaacaatca tcatcaatgg cggctacccc agtaacaaca 2340

acaacttctg ctgctgcagt ttaa 2364

<210> 7

<211> 97

<212> RNA

<213> Artificial Sequence (Artificial Sequence)

<400> 7

gggugauaau ccagaaagcg gguuuuagag cuagaaauag caaguuaaaa uaaggcuagu 60

ccguuaucaa cuugaaaaag uggcaccgag ucggugc 97

<210> 8

<211> 97

<212> RNA

<213> Artificial Sequence (Artificial Sequence)

<400> 8

gaagaagcuc cggagagugg uguuuuagag cuagaaauag caaguuaaaa uaaggcuagu 60

ccguuaucaa cuugaaaaag uggcaccgag ucggugc 97

<210> 9

<211> 28

<212> PRT

<213> Artificial Sequence (Artificial Sequence)

<400> 9

Met Leu Gly Val Ser Gly Leu Val Ser Ser Glu Gly Gly Gly Asp Asn

1 5 10 15

Pro Glu Ser Arg Trp Arg Ser Trp Lys Arg Arg Glu

20 25

<210> 10

<211> 79

<212> PRT

<213> Artificial Sequence (Artificial Sequence)

<400> 10

Met Leu Gly Val Ser Ser Ser Leu Ile Ala Ser Ser Asn Thr Ser Ile

1 5 10 15

Thr Ala Gly Ala Ala Gly Asp Gly Ala Ala Ile Ser Ala Ala Pro Ser

20 25 30

Gln Leu Ala Pro Pro Pro Gln Glu Ala Pro Glu Arg Val Val Gly Val

35 40 45

Val Lys Val Val Ala Val Glu Glu Ile Cys Arg Leu Ala Val Lys Met

50 55 60

Glu Lys Gly Thr Gln Val Glu Ile Asp Gly Gln Gly Lys Lys Leu

65 70 75

<210> 11

<211> 1966

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 11

atgctgggcg tttccggctt agtaagtagt gaaggtggtg gtgataatcc agaaagtcgg 60

tggaggagct ggaagcggag ggagtagtga gattggatta ggcggtggaa gtggcggcgg 120

tggaggtagt agcggtggat tcatgacgga agatggagaa agaaattcag gtggaaatag 180

atggccaaga caagaaacaa ttgctttgct gaaaataagg tctgaaatgg atgttatttt 240

tagagactca agtcttaaag gacctttatg ggaagaagtt tccaggaaaa tggcagacct 300

tgggttccac agaagttcca agaaatgcaa ggagaagttc gaaaatgtat acaaatatca 360

caagagaacc aaggatggcc gagcatcgaa agcggatgga aagaattata ggtttttcga 420

gcaattggaa gccctggaga acattacatc tcatcattct ctaatgccag taccgtcgtc 480

taatacgcgt cctccacccc ctccgttgga agctactcca ataaatatgg ctatgccaat 540

ggcatcatca aatgtacaag tcacggcttc acaaggtact attcctcatc atgttactat 600

ttcatcagca ccaccgccac cgaatagcct ttttgctcct tctcatcaaa atgctccgtc 660

aagttcaccc gtgccactac caccaccgcc atcacagcaa ccatcaccgc agccagctgt 720

caatccgatt aataatattc ctcaacaagt gaacgcttca gcaatgtcgt attcaacttc 780

ttcgtctact tcctcggatg aggatataca aagaaggcat aagaagaaga ggaaatggaa 840

ggattatttt gagaagttca ccaaggatgt gattaataag caggaggaat cgcacaggag 900

gttcttggag aagcttgaga agcgggaaca tgatcggatg gttcgagaag aagcatggaa 960

agtagaggaa atggcaagga tgaataggga gcatgatctt ttagttcaag aaagagcaat 1020

ggcggcagcc aaggatgcag ctgttatttc ttttttacaa aagataactg aacagcaaaa 1080

cattcaaatt ccaaatagta tcaacgttgg ccctccatca gcacaagtac aaatacaatt 1140

gcctgaaaac ccactatccg cgcctgtacc aacacaaata caaccgacga ctgttacagc 1200

agcagcacca cctcaaccag caccggtccc agtatcgttg ccagtaacaa taccagctcc 1260

agtaccagca ttaataccat cattgtcgct accactgaca ccaccagtgc catccaagaa 1320

catggagtta gtaccaaaaa gcgataacgg aggtgatagt tacagtccag caagctcttc 1380

aaggtggcca aaagcagaag ttgaagcatt gattaaactt cgtacaaatt tagatgtcaa 1440

ataccaagag aacggaccta aaggtccact ttgggaagag atatcatctg gaatgaagaa 1500

aattggatac aatcggaatg caaagagatg caaagaaaaa tgggaaaaca tcaacaaata 1560

cttcaagaag gtgaaggaga gcaacaaaaa acgacccgaa gattccaaaa cttgcccata 1620

tttccaccag ctcgatgcac tgtacaagga gaaagccaaa aaccccgaaa cagcttcttc 1680

aacgtcttcg ttcaatcctt cattcgcttt aaaccccgat aacaaccaaa tggctcccat 1740

catggctcgt ccagaacagc aatggccact tccacaacac catgaaagca ccacccgtat 1800

cgaccacgaa aacgagagcg acaacatgga tgaagatgat cacgatgatg aggaggatga 1860

agatgacgag gacgaaaaca acgcttatga gatagtagca aacaagcaac aatcctcaat 1920

ggcggccgca aacaccacta ccagcaccgc aacaacaaca gtttga 1966

<210> 12

<211> 1958

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 12

atgcttggtg tttcttcaag tttaatagct agcagtaata ctagtattac tgctggtgct 60

gcaggtgatg gagctgccat ttcggcagct ccatcacagt tagcaccgcc accacaagaa 120

gctccggaga gagtggtggg agtagtgaag gtggtggcgg tggaggagat ttgtcgattg 180

gcggtgaaga tggagaaagg aactcaggtg gaaatcgatg gccaaggcaa gaaactttag 240

ctttactgaa aattagatcg gaaatggatg ttgttttcaa agattcaagt cttaaaggac 300

cgttatggga agaagtttcc agaaaactcg cggagttggg ttatcatcga agtgctaaga 360

aatgtaaaga gaaattcgag aatgtttaca agtatcacag gagaaccaaa gatggtcgtg 420

cttcgaaagc agatggaaaa acttatcgat tctttgatca gttacaggct ttggaaaaca 480

atccatcttc tcattctaac ataccgccac ctccattagc agcaacaccc ataacaatgg 540

caatgccaat gcgatcagga aacaattcag caaatcctcc aatgccgacg ccaacgccaa 600

ctccacaaaa tcataatcat ttttttagtg tttcgcagaa aagtgttgtg acaggagcag 660

cgcagcctgc tgttatgact gcacctgcgc tgccactgtc acaagtgccg ataggtaata 720

ataacttgaa ccagatgcat cggcctcaag gtaatactac tactacaaaa acaagtttcc 780

tgtcgaattc aacttcatca tcatcttcaa cttcgtcgga tgaggatata caaaggaggc 840

agatgaagaa gcggaaatgg aaggaattct ttgagagttt aatgaaggat gtgattgaga 900

agcaagagga attgcagaag aagtttttgg aaacgctcga gaagcgcgag agggataggt 960

tgatgagaga ggaggcatgg agagtgcaag agatggctag attgaatagg gaacatgatc 1020

ttttagtcca agagagatca atggcagcag ctaaagacgc aacaatcatc gccttcttgc 1080

aaaaaataac tgaacagcaa aacacacaaa ccccgaatag tacaaataac acttctcctt 1140

ctccttttcc aattgctcaa attcaattaa aattgtccga aaagccattc agtacaccac 1200

cacaaccaca accacaacca tcagctaccg cggtatcact gccaatgaca atacatacac 1260

caacaccagc accaccacag acactgacat tacctgtagt atcatcaaaa tcacttgaac 1320

ctccaaaatc cgataatggt ggtgagaatt tctctccagc aagctcgtca agatggccga 1380

aagaagaaat cgaagcattg ataagtctcc gaacctgttt agatctaaaa taccaagaaa 1440

atggaccgaa aggaccactg tgggaagaaa tttcatctgg aatgagaaag ataggataca 1500

acaggaatgc aaagagatgc aaggaaaaat gggagaacat caacaagtac ttcaagaagg 1560

taaaagaaag caacaaaaaa agaccagaag attccaaaac ttgcccatat ttccaccagc 1620

tggaagcact gtacaaagaa aaagccaagc tcgaacctgt accacacaac actaccttcg 1680

gattaacacc ccaaaacaat cctcctcctc ctcctcctcc catcatggct caacccgagc 1740

aacaatggcc aattcctcaa aatcaacttc accagcaaaa tcgtgatcat catcacgata 1800

atgaaagcga cagcatggat cacgatttgg aagaggacga ggatgaggac gaagaagatg 1860

aaggtaatgg ctatgaaata ataatcacaa ataaacaaca atcatcatca atggcggcta 1920

ccccagtaac aacaacaact tctgctgctg cagtttaa 1958

Claims

1. A protein, comprising the steps of: the protein is SlGT-2 and/or a homologous protein SlGTL1 thereof;

the SlGT-2 is the protein of the following A1), A2) or A3):

the SlGTL1 is the protein of A4), A5) or A6) as follows:

a5 Protein which is obtained by substituting and/or deleting and/or adding more than one amino acid residue of an amino acid sequence shown in any one sequence 2 in a sequence table, is derived from the protein shown in A4) and is related to the shape of the plant fruit;

2. The protein of claim 1, wherein: a2 The protein is SlGT-2+T, and the amino acid sequence of the SlGT-2+T is a sequence 9 in a sequence table;

a5 The protein is SlGTL1+ AG, and the amino acid sequence of the SlGTL1+ AG is a sequence 10 in a sequence table.

3. A gene characterized by: the gene is an SlGT-2 gene encoding the SlGT-2 of the SlGT-1 or an SlGTL1 gene encoding the SlGTL1 of the claim 1, or an SlGT-2+T gene encoding the SlGT-2+T of the claim 2 or an SlGTL1+ AG gene encoding the SlGTL1+ AG of the claim 2;

the SlGT-2 gene is a gene shown in the following B1) or B2):

b1 The coding sequence of the coding chain is a cDNA molecule or a DNA molecule of a sequence 3 in a sequence table;

b2 Nucleotide of the coding strand is a DNA molecule of sequence 5 in the sequence table;

the SlGTL1 gene is a gene shown in the following B3) or B4):

b4 Nucleotide of the coding strand is a DNA molecule of sequence 6 in the sequence table;

4. A method of regulating the shape of a plant fruit, comprising: the method comprises the following steps: and (3) inhibiting the expression of the SlGT-2 gene of the claim 3 and the SlGTL1 gene of the claim 3 in the receptor round fruit plant to obtain the target plant of square fruit.

5. The method of claim 4, wherein: the expression of the SlGT-2 gene of claim 3 and the SlGTL1 gene of claim 3 in the recipient cone plant is realized by carrying out gene editing on the SlGT-2 gene and the SlGTL1 gene in a plant, the gene editing is realized by virtue of a CRISPR/Cas9 system, the CRISPR/Cas9 system comprises a plasmid for expressing Cas9 and sgRNA, the sgRNA is sgRNA1 aiming at a target sequence 1 and sgRNA2 aiming at the target sequence 2, the target sequence 1 is the 40 th to 59 th sites of a sequence 5 in a sequence table, and the target sequence 2 is the 116 th to 136 th sites of a sequence 6 in the sequence table.

6. The method of claim 5, wherein: the nucleotide sequence of the sgRNA1 is shown as a sequence 7 in a sequence table, and the nucleotide sequence of the sgRNA2 is shown as a sequence 8 in the sequence table.

7. The method of claim 6, wherein: the expression of the SlGT-2 gene of claim 3 and the expression of the SlGTL1 gene of claim 3 in the receptor-inhibiting roundfruit plants are the following X1 and X2:

8. The method according to any one of claims 4-7, wherein: the plant is any one of F1) to F4):

f1 Tubular plants of the order florida;

f2 Solanaceae plants;

f3 ) plants of the genus Lycopersicon;

f4 ) tomatoes.

9. The reagent for regulating and controlling the fruit type of the plant is characterized in that: the active ingredients of the reagent are substances for inhibiting the expression of the SlGT-2 gene of claim 3 and the SlGTL1 gene of claim 3, reducing the abundance of the SlGT-2 protein of claim 1 and the SlGTL1 protein of claim 1, and/or knocking out the SlGT-2 gene of claim 3 and the SlGTL1 gene of claim 3.

10. Use of the protein of claim 1, the gene of claim 3 or the agent of claim 9 for regulating the shape of a fruit and/or the moisture content of pectin in a plant.

11. Use of the method of claims 4-8 for regulating the water content of plant pectin.