CN112457380A - Protein for regulating and controlling content of fruit shape and/or fruit juice of plant, related biological material and application thereof - Google Patents

Abstract

The invention discloses a protein for regulating the fruit shape and/or fruit juice content of a plant and its related biological materials and applications. The present invention firstly discloses the application of the following proteins in regulating the fruit shape and/or juice content of plants: A1) a protein consisting of the amino acid sequence shown in SEQ ID NO: 3; A2) N-terminal or/and C-terminal connection of the protein shown in SEQ ID NO: 3 A fusion protein obtained by a protein tag; A3) The amino acid sequence shown in SEQ ID NO: 3 is obtained by substitution and/or deletion and/or addition of one or several amino acid residues, which is more than 90% identical to the protein shown in A1) proteins that are both sexual and functional. The present invention further discloses a method for growing transgenic plants of round fruit and/or juicy fruit. The invention introduces the coding gene of the protein S1FS8.1 into the plant, and successfully cultivates the plant with round juicy fruit, the operation is simple, the cost is low, the breeding process is accelerated, and the invention has wide application prospects.

Description

Protein for regulating and controlling content of fruit shape and/or fruit juice of plant, related biological material and application thereof

Technical Field

The invention belongs to the field of plant genetic engineering, and particularly relates to a protein for regulating and controlling the content of plant fruit shape and/or fruit juice, a related biological material and application thereof.

Background

Tomatoes are widely planted in the world as vegetables and fruits, and are popular among consumers in various countries. The fruit type of tomato is an important economic trait. At present, most fresh tomato fruits are round juicy fruits, because the round shape is the most favored character by consumers, and the juicy fruits have better mouthfeel. The rapid and accurate improvement of tomato fruit type is always one of the important targets of Chinese breeding.

At present, the conversion from rectangular fruit to round fruit is mainly realized by traditional means of hybridization and backcross in production, and the method has a long period, generally needs 5-10 years and is limited by factors such as linkage drag and the like.

Disclosure of Invention

The technical problem to be solved by the invention is to provide a simple and efficient method for regulating and controlling the fruit shape and/or fruit juice content of a plant.

In order to solve the problems, the invention provides a protein SlFS8.1 which is derived from tomato (Solanum lycopersicum) and is a protein shown in any one of the following items:

A1) a protein consisting of an amino acid sequence shown in a sequence 3 in a sequence table;

A2) a fusion protein obtained by connecting labels to the N end or/and the C end of the amino acid sequence shown in the sequence 3 in the sequence table;

A3) the protein which is obtained by substituting and/or deleting and/or adding one or more amino acid residues in the amino acid sequence shown in the sequence 3 in the sequence table, has more than 90 percent of identity with the protein shown in A1), and has the same function.

Wherein, the sequence 3 consists of 315 amino acid residues.

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

Among the above proteins, 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 above proteins, identity refers to the identity of amino acid sequences. The identity of the amino acid sequences can be determined using homology search sites on the Internet, such as the BLAST web pages of the NCBI home website. For example, in the advanced BLAST2.1, by using blastp as a program, setting the value of Expect to 10, setting all filters to OFF, using BLOSUM62 as a Matrix, setting Gap existence cost, Per residual Gap cost, and Lambda ratio to 11, 1, and 0.85 (default values), respectively, and performing a calculation by searching for the identity of a pair of amino acid sequences, a value (%) of identity can be obtained.

In the above protein, the 90% or more identity may be at least 91%, 92%, 95%, 96%, 98%, 99% or 100% identity.

The invention also provides the application of the protein SlFS8.1 in any one of the following steps:

B1) regulating and controlling the shape of the plant fruits;

B2) preparing a product for regulating and controlling the shape of the plant fruit;

B3) changing the shape of the plant fruit from a rectangular fruit into a round fruit;

B4) preparing a product which enables the shape of the plant fruit to be changed from a rectangular fruit into a round fruit;

B5) regulating and controlling the content of the plant juice;

B6) preparing a product for regulating and controlling the content of the plant juice;

B7) the content of the plant juice is changed from a less juice fruit to a more juice fruit;

B8) preparing a product having a reduced juice content to a higher juice content;

B19) regulating and controlling the fruit shape and the fruit juice content of the plant;

B10) preparing a product for regulating and controlling the fruit shape and the fruit juice content of the plant;

B11) the fruit of the plant is changed from a rectangular low-juice fruit into a round high-juice fruit;

B12) the method can convert the fruit of plant from rectangular low-juice fruit into round high-juice fruit.

Protein SlFS8.1 related biomaterials are also within the scope of the invention.

The biological material related to the protein SlFS8.1 also belongs to the protection scope of the invention, and the invention also provides a new application of the biological material related to the protein SlFS8.1.

The protein SlFS8.1 related biological material of the invention can be applied to any one of the following biological materials:

B1) regulating and controlling the shape of the plant fruits;

B2) preparing a product for regulating and controlling the shape of the plant fruit;

B3) changing the shape of the plant fruit from a rectangular fruit into a round fruit;

B4) preparing a product which enables the shape of the plant fruit to be changed from a rectangular fruit into a round fruit;

B5) regulating and controlling the content of the plant juice;

B6) preparing a product for regulating and controlling the content of the plant juice;

B7) the content of the plant juice is changed from a less juice fruit to a more juice fruit;

B8) preparing a product having a reduced juice content to a higher juice content;

B19) regulating and controlling the fruit shape and the fruit juice content of the plant;

B10) preparing a product for regulating and controlling the fruit shape and the fruit juice content of the plant;

B11) the fruit of the plant is changed from a rectangular low-juice fruit into a round high-juice fruit;

B12) the production of a product is made such that the fruit of the plant is transformed from a rectangular low-juice fruit to a round high-juice fruit.

In the above application, the related biomaterial is any one of the following C1) -C10):

C1) a nucleic acid molecule encoding a protein slfs8.1;

C2) an expression cassette comprising the nucleic acid molecule of C1);

C3) a recombinant vector comprising the nucleic acid molecule of C1), or a recombinant vector comprising the expression cassette of C2);

C4) a recombinant microorganism containing C1) the nucleic acid molecule, or a recombinant microorganism containing C2) the expression cassette, or a recombinant microorganism containing C3) the recombinant vector;

C5) a transgenic plant cell line comprising C1) the nucleic acid molecule, or a transgenic plant cell line comprising C2) the expression cassette, or a transgenic plant cell line comprising C3) the recombinant vector;

C6) transgenic plant tissue comprising C1) the nucleic acid molecule, or transgenic plant tissue comprising C2) the expression cassette, or transgenic plant tissue comprising C3) the recombinant vector;

C7) a transgenic plant organ containing C1) said nucleic acid molecule, or a transgenic plant organ containing C2) said expression cassette, or a transgenic plant organ containing C3) said recombinant vector;

C8) a transgenic plant containing C1) the nucleic acid molecule, or a transgenic plant containing C2) the expression cassette, or a transgenic plant containing C3) the recombinant vector;

C9) a tissue culture produced from regenerable cells of the transgenic plant of C8);

C10) protoplasts produced from the tissue culture of C9);

C11) expressing nucleic acid molecules with normal level or higher content and/or activity of protein SlFS8.1.

Wherein the nucleic acid molecule may be DNA, such as cDNA, genomic DNA or recombinant DNA; the nucleic acid molecule may also be RNA, such as mRNA or hnRNA, etc.

In the related biological material, the nucleic acid molecule C1) is represented by the following D1) or D2) or D3) or D4):

D1) DNA molecule shown in sequence 1 in the sequence table;

D2) the coding sequence is a DNA molecule shown in a sequence 2 in a sequence table;

D3) transcribing a DNA molecule of RNA shown as a sequence 5 in a sequence table;

D4) a DNA molecule which hybridizes with the DNA molecule defined by D1) or D2) or D3) under strict conditions and codes for a protein SlFS8.1.

In the related biological material, the nucleic acid molecule C11) is represented by the following E1) or E2) or E3):

E1) RNA molecule shown in sequence 5 in the sequence table;

E2) transcribing a DNA molecule of RNA shown as a sequence 5 in a sequence table;

E3) an RNA molecule or a DNA molecule which hybridizes with the nucleotide sequence limited by E1) or E2) under strict conditions and has the same function.

Wherein, the sequence 1 in the sequence table is composed of 1057 nucleotides, the coding sequence is shown as the sequence 2 and is composed of 948 nucleotides, RNA shown as the sequence 5 in the transcription sequence table, and protein shown as the coding sequence 3.

The stringent conditions are hybridization and washing of the membrane 2 times 5min at 68 ℃ in a solution of 2 XSSC, 0.1% SDS and 2 times 15min at 68 ℃ in a solution of 0.5 XSSC, 0.1% SDS.

In the above-mentioned related biological materials, the expression cassette described in C2) refers to DNA capable of expressing the protein slfs8.1 in a host cell, which DNA may include not only a promoter that initiates transcription of the slfs8.1 gene, but also a terminator that terminates transcription of the slfs8.1 gene. Further, the expression cassette may also include an enhancer sequence. Promoters useful in the present invention include, but are not limited to: the SlFS8.1 gene promoter, constitutive promoter, tissue, organ and development specific promoter and inducible promoter. Examples of promoters include, but are not limited to: the constitutive promoter of cauliflower mosaic virus 35S; the wound-inducible promoter from tomato, leucine aminopeptidase ("LAP", Chao et al (1999) Plant Physiol 120: 979-992); chemically inducible promoter from tobacco, pathogenesis-related 1(PR1) (from salicylic acid and BTH (benzothiadiazole-7-carbothioic acid S)-methyl ester) induction); tomato proteinase inhibitor II promoter (PIN2) or LAP promoter (both inducible with jasmonic acid ester); heat shock promoters (us patent 5,187,267); tetracycline inducible promoters (U.S. Pat. No. 5,057,422); seed-specific promoters, such as the millet seed-specific promoter pF128(CN101063139B (Chinese patent 200710099169.7)), seed storage protein-specific promoters (e.g., the promoters of phaseolin, napin, oleosin, and soybean beta conglycin (Beachy et al (1985) EMBO J.4: 3047-Bus3053). they can be used alone or in combination with other plant promoters. all references cited herein are incorporated herein in their entirety suitable transcription terminators include, but are not limited to, the terminator of the SlFS8.1 gene itself, the Agrobacterium nopaline synthase terminator (NOS terminator), the cauliflower mosaic virus CaMV 35S terminator, the tml terminator, the pea rbcS E9 terminator, and the nopaline and octopine synthase terminators (see, e.g., Odell et al (I patent 200710099169.7) ("see⁹⁸⁵) Nature 313: 810; rosenberg et al (1987) Gene, 56: 125; guerineau et al (1991) mol.gen.genet, 262: 141, a solvent; proudfoot (1991) Cell, 64: 671; sanfacon et al Genes dev., 5: 141, a solvent; mogen et al (1990) Plant Cell, 2: 1261; munroe et al (1990) Gene, 91: 151, and (b); ballad et al (1989) Nucleic Acids Res.17: 7891; joshi et al (1987) Nucleic Acid Res, 15: 9627).

In the related biological material, the recombinant vector C3) can contain a DNA sequence which is shown in a sequence 1 or a sequence 2 in a sequence table and is used for encoding a protein SlFS8.1.

The recombinant vector containing the protein SlFS8.1 coding gene or the expression cassette of the protein SlFS8.1 coding gene can be constructed by using the existing plant expression vector. The plant expression vector can be a Gateway system vector or a binary agrobacterium vector and the like, such as pGWB411, pGWB412, pGWB405, pBin438, pCAMBIA1302, pCAMBIA2300, pCAMBIA2301, pCAMBIA1301, pCAMBIA1300, pBI121, pCAMBIA1391-Xa or pCAMBIA 1391-Xb. When SlFS8.1 is used for constructing a recombinant vector, any one of enhanced, constitutive, tissue-specific or inducible promoters, such as a cauliflower mosaic virus (CAMV)35S promoter, a ubiquitin gene Ubiqutin promoter (pUbi) and the like, can be added in front of a transcription initiation nucleotide, and can be used alone or combined with other plant promoters; in addition, when the gene of the present invention is used to construct plant expression vectors, enhancers, including translational or transcriptional enhancers, may be used, and these enhancer regions may be ATG initiation codon or initiation codon of adjacent regions, etc., but must be in the same reading frame as the coding sequence to ensure proper translation of the entire sequence. The translational control signals and initiation codons are widely derived, either naturally or synthetically. The translation initiation region may be derived from a transcription initiation region or a structural gene.

In order to facilitate the identification and screening of transgenic plant cells or plants, plant expression vectors to be used may be processed, for example, by adding a gene encoding an enzyme or a luminescent compound which can produce a color change (GUS gene, luciferase gene, etc.), an antibiotic marker having resistance (gentamicin marker, kanamycin marker, etc.), or a chemical-resistant marker gene (e.g., herbicide-resistant gene), etc., which can be expressed in plants.

In the related biological material, the recombinant microorganism C4) can be yeast, bacteria, algae and fungi; the bacterium can be Agrobacterium LBA4404 strain.

In the above-mentioned related biological materials, C7) the transgenic plant organ may be a root, a stem, a leaf, a flower, a fruit, and a seed of the transgenic plant.

In the above-mentioned related biomaterials, C9) the tissue culture may be derived from roots, stems, leaves, flowers, fruits, seeds, pollen, embryos and anthers.

In the related biological material, the transgenic plant cell line, the transgenic plant tissue and the transgenic plant organ do not comprise propagation materials.

The application of the protein SlFS8.1 or the related biological materials thereof in cultivating transgenic plants of round fruits and/or juicy fruits is also within the protection scope of the invention; the application of the protein SlFS8.1 or related biological materials thereof in plant breeding is also within the protection scope of the invention.

Among the above applications, the plant breeding application can be specifically that a plant containing the protein slfs8.1 or the related biological material (such as the gene slfs8.1 encoded by the protein slfs8.1) is crossed with other plants to carry out plant breeding.

The invention also provides a method for changing the content of the fruit shape or the fruit juice of the plant.

The method for changing the fruit shape and/or fruit juice content of the plant comprises the step of changing the expression and/or content and/or activity of protein SlFS8.1 in a target plant, so that the fruit shape and/or fruit juice content of the target plant is changed.

The invention further provides products for modifying the fruit shape and/or juice content of plants, which contain the protein SlFS8.1 or the related biological materials.

The product can take the protein SlFS8.1 or the related biological materials as active ingredients, and can also take the protein SlFS8.1 or the related biological materials and substances with the same functions as the active ingredients.

The invention further relates to a method for cultivating transgenic plants of round fruits and/or juicy fruits.

The method for cultivating the transgenic plant with round fruits and/or juicy fruits comprises the steps of improving the expression and/or content and/or activity of protein SlFS8.1 in the target plant with rectangular fruits and/or juicy fruits to obtain the transgenic plant; the transgenic plant is round fruit and/or juicy fruit compared with the target plant.

In the method, the method for improving the expression and/or content and/or activity of the protein SlFS8.1 in the target plants of the rectangular fruits and/or the low-juice fruits is to express or over-express the protein SlFS8.1 in the target plants of the rectangular fruits and/or the low-juice fruits.

In the method, the expression or overexpression method is to introduce a coding gene of the protein SlFS8.1 into a target plant of rectangular fruits and/or juicy fruits.

The gene encoding the protein SlFS8.1 can be introduced into a target plant through a plant expression vector carrying the gene SlFS8.1. The plant expression vector carrying the gene SlFS8.1 can transform plant cells or tissues by using Ti plasmids, Ri plasmids, plant virus vectors, direct DNA transformation, microinjection, conductance, agrobacterium mediation and other conventional biological methods, and culture the transformed plant cells or tissues into plants.

The plant expression vector carrying the gene SlFS8.1 can be pCAMBIA 1300-SlFS8.1. Specifically, pCAMBIA1300-SlFS8.1 utilizes restriction enzymes SalI and KpnI to insert a DNA molecule shown in a sequence 4 into a pCAMBIA1300 vector to obtain a SlFS8.1 gene expression vector pCAMBIA1300-SlFS8.1 capable of transcribing mRNA shown in a sequence 5.

In the method, the nucleotide sequence of the coding gene of the protein SlFS8.1 is a DNA molecule shown as a sequence 2 in a sequence table.

In the present invention, the plant is a dicotyledonous plant or a monocotyledonous plant;

the dicot may be any one of F1) -F4):

F1) tubular plants of the order florida;

F2) a plant of the Solanaceae family;

F3) a plant of the genus Lycopersicon;

F4) tomato.

The invention introduces the coding gene of the protein SlFS8.1 into a target plant by utilizing a transgenic method, realizes the conversion from rectangular few-juice fruits to round many-juice fruits and successfully cultivates the round many-juice fruit plants. The invention can obtain a new plant variety with the transformation from a rectangular low-juice fruit to a round high-juice fruit within 1 year. The method of the invention has the advantages of simple operation, low cost, greatly quickening the breeding process and wide application prospect. In addition, the rectangular tomato with less juice is converted into the round tomato with more juice, although the transgenic technology is utilized, the widely applied genes are only transformed among varieties, and the safety problem is completely avoided.

Drawings

FIG. 1 shows T of wild type tomato M82, empty vector pCAMBIA1300 of rectangular low-juice fruit₀Transgenic tomato plant and round succulent fruitDue to the fruit phenotype of homozygous tomatoes.

Detailed Description

The following examples are given to facilitate a better understanding of the invention, but do not limit the invention. The experimental procedures used in the following examples are all conventional procedures unless otherwise specified. Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.

The tomatoes used in the following examples were a rectangular, less juicy variety tomato M82 (also called wild-type tomato below) and a round, juicy variety tomato AC (Ailsa Craig), which were derived from the American tomato genetic resource center (TGRC, http:// TGRC. ucdavis. edu. /).

The vector pCAMBIA1300 used in the following examples was purchased from Addgene vector libraryhttp：// www.addgene.org/)。

Coli Top10, Agrobacterium LBA4404 strain used in the examples below was purchased from Beijing Huayue Yangtze Biometrics, Inc.

The inventor researches and discovers a gene SlFS8.1 related to the shape, the juice content and the like of tomatoes from round succulent tomatoes AC, the sequence of the gene is shown as a sequence 1 and consists of 1057 nucleotides, and the sequences 393 to 501 are introns; the coding sequence is shown as sequence 2 and consists of 948 nucleotides, and the protein shown as coding sequence 3 is named as protein SlFS8.1.

Example 1 tomato transformation of rectangular Low-juice fruit into round high-juice fruit Using transgenic method

First, construction of recombinant vector

And (3) carrying out PCR amplification on the genome DNA of the wild tomato AC plant by using a primer pair consisting of a forward primer (5'-GGGGTACCGTCCACTGAACTCACAGTGTCCC-3') and a reverse primer (5'-ACGCGTCGACTCAGGTTTCGTCTTCCTTTATTTTGA-3') to obtain a PCR product of a sequence 4 in a sequence table, taking the 9 th site to the 2862 th site as a promoter, and recovering the PCR product for later use.

The PCR product and the vector pCAMBIA1300 were digested with SalI and KpnI, and ligated with T4 ligase to obtain a ligation product.

And transforming the ligation product into the competence of an escherichia coli Top10 strain, extracting a plasmid, sequencing, and identifying a clone with a correct sequence to obtain a recombinant plasmid pCAMBIA 1300-SlFS8.1. The recombinant plasmid pCAMBIA1300-SlFS8.1 contains SlFS8.1 gene shown in a sequence 1 in a sequence table, can transcribe mRNA shown in a sequence 5 in the sequence table, and further expresses protein shown in a sequence 3 in the sequence table.

Second, obtaining and identifying transgenic tomato

One), obtaining transgenic tomato:

and (3) introducing the recombinant plasmid pCAMBIA1300-SlFS8.1 obtained in the step one into an agrobacterium LBA4404 strain to obtain a recombinant strain pCAMBIA1300-SlFS8.1/LBA 4404.

Infecting a bacterial solution of a recombinant bacterium pCAMBIA1300-SlFS8.1/LBA4404 with a rectangular succulent variety tomato M82 explant, and then co-culturing for 48 hours in an MS solid culture medium (Beijing Huayu ocean organisms) containing 1mg/L indoleacetic acid, 1.75mg/L zeatin nucleoside and pH of 5.8 at 25 +/-1.5 ℃ and illumination intensity of 100-; then transferring the obtained product into an MS solid culture medium containing 1.0mg/L indoleacetic acid, 1.75mg/L zeatin, 200mg/L timentin and 75mg/L kanamycin and having the pH value of 5.8, and culturing the obtained product under the conditions of 25 +/-1.5 ℃, 16h/d illumination, 8h/d darkness and 1200lx illumination intensity until a regeneration bud grows out; cutting the regeneration bud when the regeneration bud grows to 2-3cm, transferring into MS solid culture medium containing 200mg/L timentin and 50mg/L kanamycin and having pH of 5.8, culturing at 25 + -1.5 deg.C under conditions of illumination of 16h/d, darkness 8h/d and illumination intensity of 800-₀And (5) tomato plant generation.

II) obtaining of empty vector tomato

Replacing the recombinant plasmid pCAMBIA1300-SlFS8.1 with the vector pCAMBIA1300 according to the method of the step one) and repeating the step one) to obtain the T of the empty vector pCAMBIA1300₀And (5) tomato plant generation.

Thirdly) identification of the transgenic tomato:

on MS culture medium containing 150mg/L hygromycin, the seedlings capable of growing roots are transgenic positive seedlings. The T0 generation tomatoes and the wild type tomatoes are inserted into an MS culture medium containing 150mg/L hygromycin, compared with the wild type tomatoes, the T0 generation tomatoes grow roots, the wild type tomatoes do not root, and finally the transgenic positive tomatoes which are transferred with the SlFS8.1 gene are obtained.

Extracting tomato leaf DNA, carrying out PCR amplification by using a primer pair consisting of a forward primer (5'-GCGGAGCATATACGCCCGGAG-3') and a reverse primer (5'-TGCGGCCGATCTTAGCCAGACG-3'), and detecting the amplified 436bp DNA fragment, namely the transgenic tomato. The results of the detection of the transgenic positive tomatoes and the tomatoes M82 show that 436bp DNA fragments are amplified by the transgenic positive tomatoes, and DNA fragments with corresponding lengths are not amplified by the tomatoes M82.

Planting transgenic tomato in greenhouse, and selfing to obtain T₁Transgenic tomato seeds are generated. Will T₁The transgenic tomato seeds are sown on 1/2MS culture medium containing 800mg/L hygromycin, and 10 normal-growing seedlings are selected and transplanted into the soil. Collecting T for single plant₂Seeds are generated, 30 seeds are sown on 1/2MS culture medium of 800mg/L hygromycin, and the single plant with all the seeds having resistance is the transgenic homozygous tomato material.

IV), fruit observation

The plant to be tested is the T of the transgenic homozygous tomato material obtained in the third step and the empty vector pCAMBIA1300₀Tomato plant generation and wild type tomato M82 (as control plant).

And (3) growing seedlings of the plants to be detected in a greenhouse, and culturing the seedlings under the conditions of 25 ℃ and 16h illumination/8 h dark for 20 d. 10 seedlings with consistent growth vigor are respectively selected from the transgenic homozygous tomato material and the wild tomato M82 and transplanted into a greenhouse, the plant spacing and the row spacing are more than 60cm, and the two materials are randomly distributed. Normal water and fertilizer management ensures that the water and fertilizer conditions of all plants are basically consistent. After the fruits turned completely red, the fruits of the comparative transgenic homozygous tomato material and the control plant M82 were observed and photographed for record.

The results are shown in FIG. 1, comparing the T of the control plant M82 (wild type tomato M82) and the empty vector pCAMBIA1300₀The volume of fruit juice of a single fruit of transgenic homozygous tomato material is that of wild type tomato M82 or transgenic empty tomato plants (indicated as "transgenic empty vector T0 generation tomato" in the figure) compared to round, juicy fruits of transgenic homozygous tomato materialT of somatic pCAMBIA1300₀About 2 times of the generation tomato plant.

The results show that the SlFS8.1 gene and the protein coded by the gene can regulate and control the shape and the juice content of tomatoes, and can convert tomato materials of rectangular low-juice fruits into tomato materials of round high-juice fruits.

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, national academy of sciences, Qingdao City institute of agricultural science

<120> protein for regulating and controlling content of fruit shape and/or fruit juice of plant and related biological material and application thereof

<130> GNC191936

<160> 5

<170> PatentIn version 3.5

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Ile His Ser Leu Pro Ile Thr Gln His Phe Phe Pro Tyr Gln His Ala

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His Tyr Arg Ser Met Ser Glu Glu Ile Arg Val Asp Gln Ser Gln Thr

65 70 75 80

Ala Glu Leu Val Ala Phe Pro Ala Ser Glu Ile Arg Gly Gly Gln Glu

85 90 95

Asp Ala Leu Ile Arg Gly Ser Glu Arg Tyr Cys Thr Gln Pro Arg Gln

100 105 110

Thr Cys Val Ala Val Trp Gln Asn Gln Glu Asp Ser Ala Met Lys Gln

115 120 125

Pro Val Trp Lys Gly Glu Tyr Ser Asn Gly Asn Ala Ile Glu Lys Asn

130 135 140

Lys Gln Glu Glu Asp Glu Glu Leu Tyr Ser Leu Glu Glu Thr Asn Lys

145 150 155 160

Arg Arg Val Val Phe Gly Glu Leu Glu Ala Ile Cys Ile Arg Gly Ile

165 170 175

Ala Ser Glu Ser Ala Asp Asn Leu Pro Thr Asn His Asn Val Thr Phe

180 185 190

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

195 200 205

Met Gly Lys Phe His Lys Arg Lys Arg Gly Lys Asp Glu Trp Gly Arg

210 215 220

Phe Phe Lys Ser Leu Val Lys Lys Leu Ala Asn His Gln Glu Asp Leu

225 230 235 240

Gln Arg Ser Leu Met Glu Thr Met Glu Arg Leu Asp Gln Glu Arg Lys

245 250 255

Glu Arg Glu Glu Leu Trp Arg Glu Lys Glu Leu Glu Lys Leu Gln Asn

260 265 270

Glu Glu Ala Ala Arg Ala His Glu Arg Arg Leu Ala Ser Thr Arg Glu

275 280 285

Ala Ala Leu Val Ser Cys Leu Glu Lys Leu Thr Gly Gln Lys Ile Asp

290 295 300

Phe Gln Thr Phe Lys Ile Lys Glu Asp Glu Thr

305 310 315

<210> 4

<211> 3929

<212> DNA

<213> tomato (Solanum lycopersicum)

<400> 4

ggggtaccgt ccactgaact cacagtgtcc ccttatgaaa attattcctc tctagtatcc 60

gaggtttgat ttagaatatg acctcccagg gtaaaatgat ctcaatcacc agagtataga 120

taccaaaaac tccggtgtca gcgaaccact gaaaggcagt aaactacact tagaatacta 180

gatttagtag ttgaagaaga agtccatgaa ttctatatta aaatgagagg aaatccctca 240

atttatagaa aacaaagggt agtgcgaaaa agttcttatt gtgccttacc ggaaaggtca 300

cacaccttag gaaaagtcat aatcttttaa aaaggttgtc acctttcata aaagtcacaa 360

cttaccataa aattcacaat ttttcataaa actcgcaact tttcataaaa gttataactt 420

ttcataaaag tcccaacatt tcataaaagt cacaactctt tataaaagtt ataactcttc 480

atttttcgtt cacagctttt taaaaatcca acaatgacct cgagcttgag aaattcatca 540

aaagaaaaaa gaatgaacaa gagtgtgaac catatagaca aggttatggt tcaataagaa 600

gttgaagcaa tgtcttaacc acaacacaaa aactactaaa aaggacaaaa taatattaat 660

atgcaaaagg caagaaaagg accatgagga tcgtcacact aactcatcta caatatcaaa 720

atactttaaa caaatcatgt cattttgatt cgtattatat gttcatgttg tctatagaat 780

aacacgtggc tacttttgat tcatattata tatatatata tatatatata tatatatata 840

tatatatata attttttaag gtttttttat accctcctta atattttcga tatatatctt 900

tttaacatct catttaatta atgttgaact aataaaaaca ttaaaaaata tatacttctt 960

ttttttaaaa ataatctcgt actcatctaa aatgtcaaat tcggtaaaac aaagtgagtt 1020

atttttattc aaattatatg ttttaattga actcacctta aaaggacatt taatagtttc 1080

tatgaagaaa agtcacttaa ttataacact acaccaattg aaacaataag aaataagctt 1140

ctttttttta aaaaaaaaga aagataatct cattattcat ctaaattgtc aaatatttct 1200

aaatcaaaac aagttgtttt tatttatatt ctatattttt ttaaaagatt tttatactca 1260

atttaaataa aatagattta atattaattc tgacgtatat ctctttaaaa tctcatttaa 1320

ttaacactat attaacttaa tataataaga aatgtgtttt aatttgtttt aaaggtaatg 1380

tcttaactct tacacatcta aaatgtgaaa caaatgtagt tatcaaaaga attaatcttt 1440

aggatcaaag atagagtaga attaatggct tttattcata gaaatagaat tcaaaaagaa 1500

gatatagata gttgatgaag gtggtcctag acacaaaaat taatgaagat gagggtgggg 1560

gtgggggaca cacacatctg acataggagg aggacaatag cacacagtaa tacattcacg 1620

tgcacttctt caattgtatc cttccaaaat cctaatatca gaatgtccct acacccctac 1680

ccctcactta ctaaagtcag cacactgcaa ttttttttta acatacatct ctctctgtgt 1740

gtagcagcct tctcccacac gtgtcccttt gttctcttct gtattctcca atggtaatgc 1800

atctcagatc ttctgcacca gttatccttg ccttgttttt attaaatcca atctttgctt 1860

tggaatggca tctcctcttt gctatatgtt tatacctttc acactttcaa taatagggag 1920

tacatctcct gcatagacac atgcgtccta cgtaatgcat ttaaagcaac atgttatgaa 1980

acatacatta acatgtgtac gatgttatat tggatatatg tctttttatt taattttgta 2040

caaatttaaa tatgtgtttg tataaattca aagttgaaag gtgtaacgat gaaaacatat 2100

atatgtttga tcgaatgcta gttgacgcta cttctatcta attaatgata tgaagctacc 2160

aataaaggtt gtatgatgga ataatatagt agtccttcat tcttaatcaa aggtttcaga 2220

ttcggatata tatatgaagg ttagagttaa tgaaggaaat ctcaaggcaa ttggtgtaat 2280

catgtacatg atatagtgtt ctattgttgg ctgatgttaa ctaaaaatat ggtaaataag 2340

taaaatagaa aaagaaaatg tttgttggaa tttggggcat tcatattttt tcaaagtaac 2400

aatcttttta tttgaaaaaa aaaaaaccta tatataattt agtaacatta atgacattgt 2460

ggttggcaaa ataattctac cgttagaaac aaagtgatca caaatgtttt tagtccgtga 2520

ggcgctgcta ttattgttta gtattgattt taaatcatta caaaatgatg agaattaaga 2580

aatttagaag tataaataaa tacccaaaaa aaaaaagtat aaatagtacg ggtcatggat 2640

aatataaaag cgagtaaccc ggcccataac tctctttagc cgcacttcct gcttgctaag 2700

tagaacatac aattcatttt tcttttctct ctaaaacagt ttttgcccat ctttcaaact 2760

cctccgccaa aaaaaaacct cctttccttc tcattagggt ttcctttgaa aaagaaagta 2820

ttgattgata tgttgtagta tttctagatc tgaaacgaag ctatgcaatc agattatgaa 2880

atgggtggga tccatcaaga gtgtgaagat gagcctagat tcatggtgga aaataatgct 2940

tccacttctt ccccatttta cccaaactat cacttccctc cgacgtctca tcctattctc 3000

caacaaatcc acagtttacc gattactcaa cattttttcc cttatcaaca tgctcattac 3060

agatctatgt cagaagaaat aagagtagat cagtctcaga cggcggagtt agtggctttt 3120

cctgcaagtg aaataagagg aggacaagaa gatgctttga ttcgtgggag tgagcgatac 3180

tgtacacagc ctcgacaaac ttgtgtagct gtttggcaaa accaagaaga ttctgctatg 3240

aaacaacctg tttggtaatc ttttctgcac tcaatcacat gtaaatctct cccgtacacc 3300

ctaccctatt aattaatata aatattaata ttttattatt attattattt ttgatttatg 3360

taggaaagga gaatattcaa atggaaatgc aattgagaaa aacaaacaag aagaagatga 3420

ggaattgtat agtttagaag aaactaataa acgtagagta gtattcggag agttagaagc 3480

tatttgtatt cgtggaattg catctgaatc tgctgataat ttgccaacca accacaatgt 3540

cacttttcct gaattggctc tcaatgaagc catggttaac aaaatggata atactatggg 3600

taaatttcat aaaaggaaga gagggaaaga tgaatgggga agatttttca agtcattagt 3660

gaagaaactg gcgaaccacc aagaagatct acaaagaagt ttgatggaaa caatggagag 3720

attagatcaa gaaagaaaag aaagagaaga attatggaga gagaaagaat tagaaaaact 3780

ccaaaatgaa gaagctgcta gagcccatga aagaagattg gcttcaacta gagaagctgc 3840

tcttgtttct tgcttagaaa aactcacagg tcagaaaatt gattttcaaa cattcaaaat 3900

aaaggaagac gaaacctgag tcgacgcgt 3929

<210> 5

<211> 948

<212> RNA

<213> tomato (Solanum lycopersicum)

<400> 5

augcaaucag auuaugaaau gggugggauc caucaagagu gugaagauga gccuagauuc 60

augguggaaa auaaugcuuc cacuucuucc ccauuuuacc caaacuauca cuucccuccg 120

acgucucauc cuauucucca acaaauccac aguuuaccga uuacucaaca uuuuuucccu 180

uaucaacaug cucauuacag aucuauguca gaagaaauaa gaguagauca gucucagacg 240

gcggaguuag uggcuuuucc ugcaagugaa auaagaggag gacaagaaga ugcuuugauu 300

cgugggagug agcgauacug uacacagccu cgacaaacuu guguagcugu uuggcaaaac 360

caagaagauu cugcuaugaa acaaccuguu uggaaaggag aauauucaaa uggaaaugca 420

auugagaaaa acaaacaaga agaagaugag gaauuguaua guuuagaaga aacuaauaaa 480

cguagaguag uauucggaga guuagaagcu auuuguauuc guggaauugc aucugaaucu 540

gcugauaauu ugccaaccaa ccacaauguc acuuuuccug aauuggcucu caaugaagcc 600

augguuaaca aaauggauaa uacuaugggu aaauuucaua aaaggaagag agggaaagau 660

gaauggggaa gauuuuucaa gucauuagug aagaaacugg cgaaccacca agaagaucua 720

caaagaaguu ugauggaaac aauggagaga uuagaucaag aaagaaaaga aagagaagaa 780

uuauggagag agaaagaauu agaaaaacuc caaaaugaag aagcugcuag agcccaugaa 840

agaagauugg cuucaacuag agaagcugcu cuuguuucuu gcuuagaaaa acucacaggu 900

cagaaaauug auuuucaaac auucaaaaua aaggaagacg aaaccuga 948

Claims (10)

1. Use of a protein in any of the following B1)-B12): B1) regulating plant fruit shape; B2) prepare the product of regulating and controlling the fruit shape of plant; B3) make the plant fruit shape change from rectangular fruit to round fruit; B4) prepare a product that transforms the shape of the plant fruit from a rectangular fruit into a round fruit; B5) regulate and control the content of plant juice; B6) prepare the product of regulating and controlling the content of plant juice; B7) make the plant juice content change from less juicy fruit to juicy fruit; B8) prepare a product that changes the content of plant juice from less juicy fruit to juicy fruit; B19) regulating plant fruit shape and fruit juice content; B10) Preparation of products for regulating and controlling plant fruit shape and fruit juice content; B11) transforming the fruit of the plant from a rectangular less juicy fruit type to a round juicy fruit type; B12) preparing to transform the fruit of the plant from a rectangular less juicy fruit type to a round juicy fruit type product; Said protein is as follows A1) or A2) or A3): A1) a protein consisting of the amino acid sequence shown in Sequence 3 in the Sequence Listing; A2) The fusion protein obtained by linking the N-terminus or/and the C-terminus of the protein shown in Sequence 3 in the sequence listing with a protein tag; A3) The amino acid sequence shown in SEQ ID NO: 3 in the sequence listing is subjected to substitution and/or deletion and/or addition of one or several amino acid residues to obtain a protein with more than 90% identity and the same function as the protein shown in A1). of protein. 2. the application of the relevant biological material of the protein of claim 1 in any one of B1)-B12) described in claim 1, the relevant biological material is any one of the following C1)-C11): C1) a nucleic acid molecule encoding the protein of claim 1; C2) an expression cassette containing the nucleic acid molecule of C1); C3) a recombinant vector containing the nucleic acid molecule described in C1) or a recombinant vector containing the expression cassette described in C2); C4) a recombinant microorganism containing the nucleic acid molecule described in C1), or a recombinant microorganism containing the expression cassette described in C2), or a recombinant microorganism containing the recombinant vector described in C3); C5) a transgenic plant cell line containing the nucleic acid molecule of C1), or a transgenic plant cell line containing the expression cassette of C2), or a transgenic plant cell line containing the recombinant vector of C3); C6) a transgenic plant tissue containing the nucleic acid molecule of C1), or a transgenic plant tissue containing the expression cassette of C2), or a transgenic plant tissue containing the recombinant vector of C3); C7) a transgenic plant organ containing the nucleic acid molecule of C1), or a transgenic plant organ containing the expression cassette of C2), or a transgenic plant organ containing the recombinant vector of C3); C8) a transgenic plant containing the nucleic acid molecule of C1), or a transgenic plant containing the expression cassette of C2), or a transgenic plant containing the recombinant vector of C3); C9) a tissue culture produced from regenerable cells of the transgenic plant described in C8); C10) protoplasts produced by the tissue culture described in C9); C11) A nucleic acid molecule expressing a normal level or higher content and/or activity of the protein described in claim 1. 3. application according to claim 2 is characterized in that: C1) described nucleic acid molecule is as shown in following D1) or D2) or D3) or D4): D1) the DNA molecule shown in sequence 1 in the sequence listing; D2) the coding sequence is the DNA molecule shown in sequence 2 in the sequence listing; D3) transcribe the DNA molecule of RNA shown in sequence 5 in the sequence table; D4) a DNA molecule that hybridizes to a DNA molecule defined in D1) or D2) or D3) under stringent conditions and encodes a protein according to claim 1; C11) The nucleic acid molecule is as shown in the following E1) or E2) or E3): E1) RNA molecule shown in sequence 5 in the sequence listing; E2) DNA molecule of RNA shown in sequence 5 in the transcription sequence table; E3) An RNA molecule or a DNA molecule that hybridizes to the nucleotide sequence defined in E1) or E2) under stringent conditions and has the same function. 4. The use of the protein described in claim 1 or the related biological material described in claim 2 in the cultivation of transgenic plants of round fruit and/or juicy fruit. 5. Use of the protein of claim 1 or the related biological material of claim 2 in plant breeding. 6. a kind of method that changes plant fruit shape and/or fruit juice content, it is characterized in that: comprise changing the expression and/or content and/or activity of the protein described in claim 1 in the purpose plant, make the fruit shape and/or the activity of the purpose plant. / or change in juice content. 7. a kind of method for cultivating the transgenic plant of round fruit and/or juicy fruit, is characterized in that: comprise the expression of protein described in claim 1 and/or in the purpose plant of improving rectangular fruit and/or juicy fruit content and/or activity to obtain a transgenic plant; compared with the target plant, the transgenic plant is a round fruit and/or a juicy fruit. 8. The method according to claim 7, characterized in that: the expression and/or content and/or activity of the protein described in claim 1 in the target plant for improving the rectangular fruit and/or the less juicy fruit are in the rectangular The protein of claim 1 is expressed or overexpressed in the target plant of fruit and/or juicy fruit. 9. The method according to claim 7, characterized in that: the method for expressing or overexpressing is to introduce the encoding gene of the protein described in claim 1 into the target plant of Fructus Fructus and/or Juicy Fructus; specific , the nucleotide sequence of the protein-encoding gene of claim 1 is the DNA molecule shown in SEQ ID NO: 2 in the sequence listing. 10. The use according to any one of claims 1-5, or the method according to any one of claims 6-9, wherein the plant can be a dicotyledonous plant or a monocotyledonous plant.

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