CN112010956B

CN112010956B - Wheat booting stage root depth related gene TaVSR1-B and encoding protein and application thereof

Info

Publication number: CN112010956B
Application number: CN202010993871.3A
Authority: CN
Inventors: 王景一; 景蕊莲; 李龙; 毛新国; 李超男
Original assignee: Institute of Crop Sciences of Chinese Academy of Agricultural Sciences
Current assignee: Institute of Crop Sciences of Chinese Academy of Agricultural Sciences
Priority date: 2020-09-21
Filing date: 2020-09-21
Publication date: 2022-07-19
Anticipated expiration: 2040-09-21
Also published as: CN112010956A

Abstract

The invention discloses a protein coded by a wheat booting stage root depth related gene TaVSR1-B and application thereof. The invention provides the application of any substance of the following 1) to 3) in at least one of the following a) to e); 1) protein TaVSR 1-B; 2) a nucleic acid molecule encoding the protein TaVSR 1-B; 3) a recombinant vector, an expression cassette or a recombinant bacterium containing a nucleic acid molecule encoding the protein TaVSR 1-B; a) regulating and controlling the growth of plant roots; b) breeding plant varieties with different root depths; c) cultivating plants with increased root depth; d) cultivating the plant with increased root weight; e) cultivating plants with changed root system configurations; the TaVSR1-B encoded protein provided by the invention has important significance in regulating and controlling the growth of plant roots and plays an important role in cultivating plant varieties with different root depths.

Description

Wheat booting stage root depth related gene TaVSR1-B and encoding protein and application thereof

Technical Field

The invention belongs to the field of plant genetic engineering, and relates to a wheat booting stage root depth related gene TaVSR1-B and a coding protein and application thereof.

Background

Wheat is an important food crop for human beings, but stress conditions such as drought, high temperature and the like seriously restrict the production of wheat. The root system is not only an important organ for crops to absorb water and nutrients, but also an important organ for substance synthesis and transformation, and is closely related to the yield and stress resistance of crops. However, the root system is not easy to observe, and the research for systematically explaining the genetic mechanism of the wheat root depth from the gene level has been reported rarely.

Wheat is a typical fibrous root system crop, the root system of the wheat consists of seed roots and adventitious roots, and the root depth is an important aspect of the root system configuration and plays an important role in absorbing water and nutrients in the deep layer of soil. The booting period is one of the key periods of wheat yield formation, and the root system configuration directly determines the absorption of soil moisture and nutrients by wheat, so that the grain number and thousand grain weight of a single plant per ear are influenced. Therefore, the root depth of the wheat in the booting stage is an important factor influencing the yield of the wheat. The development of the wheat root depth related gene in the booting stage has important theoretical value and also has very important practical significance for improving the stress resistance and the yield potential.

Disclosure of Invention

The invention aims to provide a wheat booting stage root depth related gene TaVSR1-B and application of a protein coded by the gene.

The invention provides the application of any substance in the following 1) to 3) in at least one of the following a) to e);

1) protein TaVSR 1-B;

2) a nucleic acid molecule encoding the protein TaVSR 1-B;

3) a recombinant vector, an expression cassette or a recombinant bacterium containing a nucleic acid molecule encoding the protein TaVSR 1-B;

a) regulating and controlling the growth of plant roots;

b) breeding plant varieties with different root depths;

c) cultivating plants with increased root depth;

d) cultivating the plant with increased root weight;

e) cultivating plants with changed root system configurations;

the protein TaVSR1-B is (1), (2) or (3) as follows:

(1) a protein consisting of an amino acid sequence shown in a sequence 1 in a sequence table;

(2) a protein formed by adding a tag sequence at the tail end of an amino acid sequence shown in a sequence 1 in a sequence table;

(3) and (b) protein which is derived from the protein (1) or (2) and has the same function, wherein the protein 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 1 in the sequence table.

In order to facilitate the purification of the protein shown in (1) above, a tag as shown in the following Table 1 may be attached to the amino terminus or the carboxyl terminus of the protein consisting of the amino acid residue sequence of SEQ ID No. 1 in the sequence Listing.

Table 1 sequences of tags

Label (R)	Residue(s) of	Sequence of
			Poly-Arg	5-6 (typically 5)	RRRRR
Poly-His	2-10 (generally 6)	HHHHHH
			FLAG	8	DYKDDDDK
Strep-tag II	8	WSHPQFEK
			c-myc	10	EQKLISEEDL

The protein of (2) above may be synthesized artificially, or may be obtained by synthesizing the coding gene and then performing biological expression. The gene encoding the protein of (2) above can be obtained by deleting one or several codons of amino acid residues from the DNA sequence shown in sequence 2 of the sequence listing, and/or performing missense mutation of one or several base pairs.

The nucleic acid molecule may be DNA, such as cDNA, genomic DNA or recombinant DNA; the nucleic acid molecule can also be an RNA, such as an mRNA, hnRNA, or tRNA, and the like.

In one embodiment of the invention, the nucleic acid molecule encoding the protein TaVSR1-B is a DNA molecule according to any one of the following 1) to 6):

1) the coding region is a DNA molecule shown as a sequence 2 in a sequence table;

2) the coding region is a DNA molecule shown in the 215 th-2158 site of the sequence 2 in the sequence table;

3) the coding region is a DNA molecule shown in the 224-2143 site of the sequence 2 in the sequence table;

4) the coding region is a DNA molecule of a DNA molecule shown in a sequence 4 in a sequence table;

5) a DNA molecule which hybridizes with the DNA sequence defined in any one of 1) to 4) under strict conditions and codes for a protein with the same function;

6) a DNA molecule having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% homology to the DNA sequence defined in any one of 1)1) to 4) and encoding a protein having the same function.

The above stringent conditions may be hybridization at 65 ℃ using a solution of 6 XSSC, 0.5% SDS, followed by washing the membrane once with each of 2 XSSC, 0.1% SDS and 1 XSSC, 0.1% SDS.

Wherein the sequence 2 consists of 2786 nucleotides, the 224-th 2143 th site is ORF and encodes the protein shown in the sequence 1 in the sequence table.

The recombinant vector can be a recombinant expression vector and can also be a recombinant cloning vector.

The recombinant expression vector can be constructed using existing plant expression vectors. The plant expression vector comprises a binary agrobacterium vector, a vector which can be used for plant microprojectile bombardment and the like, such as pGreen0029, pCAMBIA3301, pCAMBIA1391, pBI121, pBin19, pCAMBIA2301, pCAMBIA1301-UBIN or other derivative plant expression vectors. The plant expression vector may also comprise a 3' untranslated region of the foreign gene, i.e., comprising a polyadenylation signal and any other DNA segments involved in mRNA processing or gene expression. The poly A signal can direct the addition of poly A to the 3' end of the mRNA precursor. When the gene is used for constructing a recombinant expression vector, any one of enhanced, constitutive, tissue-specific or inducible promoters, such as a cauliflower mosaic virus (CAMV)35S promoter, a Ubiquitin gene Ubiquitin promoter (pUbi), a stress-inducible promoter rd29A and the like, can be added before the transcription initiation nucleotide, and can be used alone or in combination with other plant promoters; in addition, when the gene of the present invention is used to construct a recombinant expression vector, enhancers, including translational enhancers or transcriptional enhancers, may be used, and these enhancer regions may be ATG initiation codons or initiation codons 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, the recombinant expression vectors used may be processed, for example, by adding genes encoding enzymes or luminescent compounds which produce a color change, antibiotic markers having resistance or chemical resistance marker genes, etc., which are expressed in plants. Or directly screening the transformed plants in a stress environment without adding any selective marker gene.

In the invention, the promoter for starting the gene transcription in the recombinant expression vector is specifically the promoter of the wheat TaVSR1-B gene.

More specifically, the recombinant expression vector is a recombinant plasmid obtained by inserting the gene promoter and CDS into the multiple cloning site of the pCAMBIA1391 vector.

The pCAMBIA1391-TaVSR1-B is characterized in that the construction method of the TaVSR1-B vector is as follows: pCAMBIA1391 is used as an original vector, and a promoter and CDS of a TaVSR1-B gene are inserted into a multiple cloning site of the pCAMBIA1391-TaVSR1-B vector, namely TaVSR1-B vector.

Wherein, the promoter sequence of the TaVSR1-B gene is specifically a sequence 3 in a sequence table.

The expression cassette consists of a promoter capable of driving expression of the gene, and a transcription termination sequence.

In a), the regulating the root growth of the plant is specifically promoting the root growth of the plant, and in the embodiment of the invention, the root depth of the plant is increased, and/or the root weight of the plant is increased, and in the embodiment of the invention, the root weight is the dry root weight.

In b), the breeding of plant varieties with different root depths is breeding of plant varieties with increased root depths or breeding of plant varieties with decreased root depths.

The invention also provides a method for promoting the growth of plant roots, which is 1) or 2) as follows:

1) the method comprises the following steps: the content and/or activity of the protein TaVSR1-B in the target plant are/is improved, and the growth of plant roots is promoted;

2) the method comprises the following steps: the expression of the nucleic acid molecule of the coding protein TaVSR1-B in the target plant is improved, and the growth of plant roots is promoted;

the protein TaVSR1-B is (1), (2) or (3) as follows:

(2) a protein formed by adding a tag sequence to the tail end of an amino acid sequence shown in a sequence 1 in a sequence table;

In the method, the promotion of the plant root growth is embodied in increasing the root depth and/or the root weight of the plant.

It is still another object of the present invention to provide a method for cultivating transgenic plants having increased root depth and/or increased root weight, which is 1) or 2) as follows:

1) the method comprises the following steps: improving the content and/or activity of the protein TaVSR1-B in the target plant to obtain a transgenic plant;

2) the method comprises the following steps: improving the expression of a nucleic acid molecule of the coding protein TaVSR1-B in the target plant to obtain a transgenic plant;

the transgenic plant has deeper root and/or bigger root than the target plant;

the protein TaVSR1-B is (1), (2) or (3) as follows:

(1) a protein consisting of an amino acid sequence shown as a sequence 1 in a sequence table;

In the above method, the increasing of the content and/or activity of protein TaVSR1-B in the plant of interest or the increasing of the expression of the nucleic acid molecule encoding protein TaVSR1-B in the plant of interest is carried out by introducing the nucleic acid molecule encoding protein TaVSR1-B into the plant of interest.

The gene can be specifically introduced into the receptor plant through any one of the recombinant expression vectors to obtain the transgenic plant. Specifically, the recombinant expression vector can be transformed into plant cells or tissues by using conventional biological methods such as Ti plasmid, Ri plasmid, plant viral vector, direct DNA transformation, microinjection, conductance, agrobacterium mediation, gene gun, etc., and the transformed plant tissues can be cultivated into plants.

In the above application or method, the plant may be a monocotyledon or a dicotyledon; such as: wheat, rice, tobacco or Arabidopsis, etc. In the embodiment of the invention, the plant is rice or wheat, in particular rice Kitaake, wheat Fielder.

Experiments prove that the T obtained by transforming rice with a recombinant expression vector capable of expressing DNA molecules shown as a sequence 2 in a sequence table₃Compared with wild plants under the same conditions, the generation homozygous transgenic plants have 4.50cm increased root depth, namely 14.8% deepened root; the dry root weight increased by 0.108g, i.e., 11.8%. T obtained by transforming wheat with recombinant expression vector capable of expressing DNA molecule shown in sequence 2 in sequence table₃Compared with wild plants under the same conditions, the generation homozygous transgenic plants have 6.51cm increased root depth, namely 8.42% deepened root; the dry weight of the root is increased by 0.049g, namely the rootThe dry weight increased by 12.3%. The TaVSR1-B gene and the coding protein thereof provided by the invention have important significance in regulating and controlling root depth, and can play an important role in cultivating plant varieties with different root system configurations.

Drawings

FIG. 1 shows that the relative expression of TaVSR1-B gene in different tissues of wheat is detected by real-time fluorescent quantitative PCR.

FIG. 2 shows the results of plant root phenotype identification of Arabidopsis thaliana and wheat tetraploid Kronos mutant vsr 1; wherein, A is a plant phenotype photo of an arabidopsis vsr1 mutant, B is a root growth histogram of each strain of an arabidopsis vsr1 mutant, C is a plant phenotype photo of a wheat mutant vsr1, and D is a root depth histogram of each strain of the wheat mutant vsr 1.

FIG. 3 is the information related to Arabidopsis mutant vsr 1; wherein, A is the insertion site information of the Arabidopsis vsr1 mutant, B is the identification result of the Arabidopsis vsr1 mutant, and C is the AtVSR1 gene expression result of the Arabidopsis vsr1 mutant.

FIG. 4 is the T-DNA region structure diagram of recombinant vector pCAMBIA1391-TaVSR1-B TaVSR 1-B.

FIG. 5 shows the results of the identification of transgenic lines; wherein, A is the identification result of the transgenic rice, B is the expression condition of the TaVSR1-B gene in the transgenic rice, C is the identification result of the transgenic wheat, and B is the expression condition of the TaVSR1-B gene in the transgenic wheat.

FIG. 6 shows the results of phenotypic identification of transgenic rice plants.

FIG. 7 shows the results of phenotypic identification of transgenic wheat plants.

Detailed Description

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.

Wheat (Triticum aestivum L.) variety dry selection No. 10: drought-tolerant varieties are derived from a national germplasm bank (serial number: ZM009279) and are recorded in the text of' Wangzhenghai, Wuxianshan, Changxueping and the like, analysis on chlorophyll content of wheat flag leaves and gray correlation degree of fluorescence kinetic parameters and yield, the crop academy, 2010-02, and the invention is obtained by the applicant and is only limited to be used for repeating the invention.

pCAMBIA1391 vector: product of Biovector Science Lab, Inc.

Agrobacterium tumefaciens EHA105(Agrobacterium tumefaciens Strain EHA 105): product of Biovector Science Lab, Inc.

Arabidopsis thaliana Columbia type 0 (Clo-0, hereinafter also referred to as wild type Arabidopsis thaliana): described in "Mayueli. influence of exogenous NO on Arabidopsis callus respiration intensity and mitochondrial Complex I protein". A Master paper, 2007, Lanzhou university, publicly available from the Applicant, is limited to use in repeating the present invention.

Arabidopsis mutants vsr1-7(SALK-000723) and vsr1-8(SALK-040506) were purchased from the American SALK center, and only T-DNA insertion occurred for TaVSR1-B gene compared to wild-type Columbia type 0 (Clo-0), and the other sequences were unchanged.

Wheat mutants Kronos M1, M2, M3(TILLING mutants) were purchased from UC Davis usa and backcrossed twice with wild-type Kronos, identifying lines with mutations in the TaVSR1-B gene for phenotypic analysis.

Example 1 obtaining of TaVSR1-B Gene and its encoded protein and study of Gene expression level

Firstly, TaVSR1-B gene and obtaining of coding protein thereof

Seeds of wheat (Triticum aestivum L.) variety No. 10 are selected by dry method, placed in a culture dish with diameter of 9cm, and placed at 22 deg.C with light intensity of 150 μmol · m^-2·s^-1Adding deionized water for culturing under the conditions of 12h illumination, 12h darkness and 70% relative humidity.

Taking 10 dry-selected seedling leaves growing to one leaf and one heart, quickly freezing the seedling leaves by liquid nitrogen, and storing the seedling leaves at the temperature of minus 80 ℃ for later use. Total RNA was extracted from TRIZOL, and first strand cDNA (Invitrogen) was synthesized using M-MLV reverse transcription kit, and PCR was performed using the cDNA as a template and primers F1 and R1.

F1: 5'-GTGATCGAGATGGGAGGGAGATCG-3' (same as in 215 th-238 of sequence 2);

r1: 5'-GAACTCCTTGGTGCTTCAGATGTCG-3' (complementary to 2134-2158 of SEQ ID NO: 2).

And (3) carrying out agarose gel electrophoresis on the amplification product, recovering the purified fragment and sequencing, wherein the sequence length of the fragment is 1917bp, as shown in the 215 th 2158 th site of the sequence 2 in the sequence table. The gene shown in sequence 2 of the sequence table is named TaVSR1-B, and the 224 th and 2143 th genes in the sequence 2 are CDS regions. The protein TaVSR1-B coded by the gene shown in the sequence 2 of the sequence table is composed of 639 amino acid residues and is shown in the sequence 1 of the sequence table.

Secondly, real-time fluorescent quantitative PCR analysis of the expression quantity of the TaVSR1-B gene in different development periods of wheat

The following materials were taken at the following times, respectively:

1) seedling stage material: no. 10 wheat variety with 2 weeks is selected on dry land, and roots and leaves are respectively taken.

2) Booting period material: no. 10 wheat varieties planted in the fields are selected in a dry mode, and roots, stems, leaves and ears are respectively taken.

Total RNA of the liquid nitrogen preserved samples is respectively extracted by TRIZOL, first-strand cDNA (Invitrogen) is synthesized by an M-MLV reverse transcription kit, and the expression level of the gene TaVSR1-B in different development periods of wheat is detected by a Real-time Quantitative PCR (Real-time Quantitative PCR) method. Using a constitutively expressed GAPDH gene as an internal control, primer sequences were designed as follows:

the primer sequence for detecting the expression of the gene TaVSR1-B is as follows:

TaVSR1-B-RT-F：5’-GACATCTGAAGCACCAAGGAGTTCTAAGTAC-3’；

TaVSR1-B-RT-R：5’-CTTGAAGTTTCAGAAAGAGAGAAAAGAGAGAGTAC-3’；

the primer sequences for detecting the expression of the gene GAPDH are as follows:

GAPDH-RT-F：5’-CTGCATCATACGATGACATC-3’；

GAPDH-RT-R：5’-TGTCACCGACAAAGTCAGTG-3’。

according to the method for estimating the relative expression amount of a target gene proposed by Livak and Schmittgen (Livak, K.J.; Schmittgen, T.D. analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta Delta Delta Delta Delta Delta Delta C (T)) method, methods, 2001,25, 402-408, doi: 10.1006/method, 2001.1262),calculating that the expression level of the TaVSR1-B gene in each tissue at different development stages is N times of the expression level of the TaVSR1-B gene in the stem at the booting stage, wherein N is 2^－ΔΔCT，ΔΔC_T＝(C_T,Target–C_T,Tubulin)_DT–(C_T,Target–C_T,Tubulin)_S. Wherein DT (differential tissues) in the formula represents a certain tissue in a certain development period, and S (stem) represents a tissue (stem in the invention) with the lowest expression level in the same development period as DT.

The experiment was repeated 3 times, and the results are shown in fig. 1 (expression in stem at the booting stage is taken as reference, and expression levels in other developmental stages and other tissues are taken as relative expression), and it can be seen that the expression levels of TaVSR1-B are higher in root and ear at the seedling stage and the booting stage.

Wherein, C_TThe meaning of the values is: the number of cycles that the fluorescence signal in each reaction tube has undergone to reach a set threshold. PCR cycles at arrival C_TThe number of cycles at which the value is at, just before the true exponential amplification phase (log phase), where the minor error has not yet been amplified, so C_TThe value reproducibility is excellent, i.e. C obtained by amplification of the same template at different times or in different tubes at the same time_TThe value is constant.

Example 2 functional study of TaVSR1-B Gene

Preparation of TaVSR1-B transgenic plant

1. Construction of recombinant expression vectors

1) Cloning of CDS region of TaVSR1-B Gene

A primer pair (F1 and R1) is designed according to the sequence of the TaVSR1-B gene, the TaVSR1-B gene (215-.

F1: 5'-GTGATCGAGATGGGAGGGAGATCG-3' (same as in 215 th-238 of sequence 2);

r1: 5'-GAACTCCTTGGTGCTTCAGATGTCG-3' (complementary to 2134 th and 2158 of SEQ ID NO: 2).

2) Cloning of promoter region of TaVSR1-B Gene

Designing a primer pair (F3 and R3) according to the sequence of the TaVSR1-B gene, taking DNA of No. 10 dry choice wheat variety as a template, carrying out PCR amplification on a promoter (sequence 3) of the TaVSR1-B gene, carrying out agarose gel electrophoresis on a PCR amplification product, and recovering and purifying a band.

F2: 5'-CGGTGGGAGTACTGGTTAGAACACTTATG-3' (identical to positions 1-29 of SEQ ID NO: 3);

r2: 5'-CAGCACCGACACGAGAAGAAGAAGAAG-3' (same as sequence 3 at position 2424-2451).

3) Connection of TaVSR1-B gene promoter region and CDS region

Using the recovered TaVSR1-B gene promoter region obtained in 2) and the CDS region fragment obtained in 1) as templates, amplifying by using primers F2 and R1, carrying out agarose gel electrophoresis on the PCR amplification product, and recovering a purified band.

4) Promoter region and CDS region of TaVSR1-B gene increase enzyme cutting sites EcoR I and Spe I

A primer set (F3 and R3) was designed based on the TaVSR1-B gene sequence, PCR amplification was performed using the above fragments as templates (sequence 4), and the PCR amplification product was subjected to agarose gel electrophoresis to recover a purified band.

F3: 5'-CTAGGAATTCCGGTGGGAGTACTGGTTAGAACACTTATG-3' (the italic part is the EcoR I enzyme cutting site, the orthobody part is the same as the 1 st to 29 th position of the sequence 4);

r3: 5'-TGACACTAGTGAACTCCTTGGTGCTTCAGATGTCG-3' (the italic part is the Spe I cleavage site and the plus part is complementary to the 4280-4304 th site of SEQ ID NO: 4).

5) Construction of recombinant expression vectors

Digesting the purified PCR product recovered in the step 4 by using restriction enzymes EcoR I and Spe I, and recovering a digested product;

secondly, the vector pCAMBIA1391 is cut by restriction enzymes EcoR I and Spe I, and the vector framework is recovered;

connecting the enzyme digestion product of the step I with the carrier skeleton of the step II;

fourthly, the ligation product in the third step is converted into an escherichia coli DH5 alpha strain by electric shock, the strain is cultured overnight at 37 ℃, and positive clones are picked for sequencing; sequencing results show that the recombinant vector pCAMBIA1391-TaVSR1-B TaVSR1-B is obtained.

The recombinant vector pCAMBIA1391-TaVSR1-B has the structure description that TaVSR1-B is as follows: the DNA molecule (sequence 4 in the sequence table) consisting of the promoter of TaVSR1-B and the CDS of TaVSR1-B is inserted between the EcoR I and Spe I enzyme cutting sites of the pCAMBIA1391 vector to obtain the recombinant plasmid.

The structure of the recombinant vector insert is schematically shown in FIG. 4.

2. Obtaining plants transformed with TaVSR1-B

1) And 2. mu.L of pCAMBIA1391-TaVSR1-B prepared in the above 1, TaVSR1-B was added to 20. mu.L of Agrobacterium tumefaciens EHA105 competent cells, and heat shock transformation was performed at 28 ℃.

2) The transformed Agrobacterium was identified by colony PCR using pCAMBIA1391 vector primer set (F4 and R4) (the colony with a PCR product of 4349bp was positive), and named recombinant Agrobacterium EHA105/pCAMBIA1391-TaVSR1-B:: TaVSR 1-B.

F4：5’-CTTGGAGTAGACGAGAGTGTCGTGC-3’；

R4：5’-GGTTGGGGTTTCTACAGGACGTAAAC-3’。

Recombinant Agrobacterium EHA105/pCAMBIA1391-TaVSR1-B TaVSR1-B was inoculated into 5mL YEB liquid medium (containing 50mg/L kanamycin and 50mg/L rifampicin), and cultured overnight at 28 ℃ under shaking at 250 rpm.

3) The recombinant agrobacterium EHA105/pCAMBIA1391-TaVSR1-B is used for infecting the callus of the leaf blade of a wild rice Kitaake (hereinafter referred to as wild rice) by using an agrobacterium infection method, and then the T0 generation TaVSR1-B rice is obtained by sequentially carrying out co-culture, screening culture (the screening antibiotic is hygromycin with the concentration of 50mg/L) and rooting culture.

The recombinant agrobacterium EHA105/pCAMBIA1391-TaVSR1-B is used for infecting callus of embryo of wild type wheat Fielder (hereinafter referred to as wild type wheat) by using an agrobacterium infection method, and then co-culture, screening culture (the antibiotic used for screening is hygromycin with the concentration of 50mg/L) and rooting culture are sequentially carried out to obtain T0 generation transgenic TaVSR1-B wheat.

The empty vector pCAMBIA1391 is respectively transferred into wild type wheat and wild type rice according to the method to respectively obtain T0 generation empty vector transferred wheat and T0 generation empty vector transferred rice.

And breeding the T0 generation plants to obtain each T3 generation plant.

3. Identification of TaVSR1-B transgenic plants

1) Identification of positive transgenic plants

The pCAMBIA1391 vector primer pair (F4 and R4) is used for detecting positive transgenic plants, and the plants with the PCR products of 4349bp are the positive transgenic plants.

Primer F4 has sequence 5'-CTTGGAGTAGACGAGAGTGTCGTGC-3';

primer R4 has the sequence 5'-GGTTGGGGTTTCTACAGGACGTAAAC-3'.

Respectively taking strains of T3-generation TaVSR1-B rice, T3-generation TaVSR1-B wheat, wild rice and wild wheat in booting stage.

RNA of each strain root was extracted using TRIZOL as a template, and PCR amplification was performed using F4 and R4.

The results are shown in FIG. 5, wherein FIG. 5A shows the identification result of transgenic rice and FIG. 5C shows the identification result of transgenic wheat, compared with the wild type variety, the transgenic plant has 4349bp target segment, and is positive transgenic plant, and positive T3 generation transformed TaVSR1-B rice OE1, OE3, OE7, OE8 and positive T3 generation transformed TaVSR1-B wheat OE3, OE5, OE7 and OE89 are obtained.

2) Real-time fluorescent quantitative PCR detection of expression condition of TaVSR1-B gene of transgenic plant

The specific operation is as follows:

respectively taking strains of a booting stage material T3 generation transformed TaVSR1-B rice, T3 generation transformed TaVSR1-B wheat, T3 generation transformed empty carrier wheat, T3 generation transformed empty carrier rice, wild rice and wild wheat.

Total RNA of each strain root is extracted by TRIZOL, first strand cDNA (Invitrogen) is synthesized by an M-MLV reverse transcription kit, and the expression level of the gene TaVSR1-B is detected by a Real-time Quantitative PCR (Real-time Quantitative PCR) method. Using a constitutively expressed GAPDH gene as an internal control, primer sequences were designed as follows:

TaVSR1-B-RT-F：5’-GACATCTGAAGCACCAAGGAGTTCTAAGTAC-3’；

TaVSR1-B-RT-R：5’-CTTGAAGTTTCAGAAAGAGAGAAAAGAGAGAGTAC-3’；

GAPDH-RT-F：5’-CTGCATCATACGATGACATC-3’；

GAPDH-RT-R：5’-TGTCACCGACAAAGTCAGTG-3’。

as shown in FIGS. 5B and 5D, in FIG. 5B, the results of the identification of transgenic rice and in FIG. 5D, the results of the identification of transgenic wheat show that the expression levels of TaVSR1-B gene in the roots of TaVSR1-B rice (OE-7, OE-8) at T3 and TaVSR1-B wheat (OE-7, OE-9) at T3 were increased as compared with those of the wild type variety.

The results of the empty vector plants of each T3 generation were not significantly different from those of the corresponding wild-type plants.

Second, phenotype of individual plant mutants

1. Root phenotype of Arabidopsis mutants

1) Arabidopsis mutant-related assays

The insertion sites of the Arabidopsis thaliana mutants are shown in FIG. 3A. PCR amplification was performed using two pairs of primers (L + R and LB + R), and as shown in FIG. 3B, a PCR amplification product was obtained using the wild type Arabidopsis thaliana material as a template and using the primer pair (L + R), whereas a PCR amplification product was not obtained using the primer pair (LB + R). The arabidopsis mutant material is used as a template, a PCR amplification product cannot be obtained by using the primer pair (L + R), and the PCR amplification product can be obtained by using the primer pair (LB + R).

L：5’-ATTTGATGAATGTGGTGGTGG-3’；

R：5’-ACCTCCACGATCGATAAGGAC-3’；

LB：5’-ATTTTGCCGATTTCGGAAC-3’。

Detection of expression of genes in mutants:

total RNAs of wild type Arabidopsis thaliana (clo-0), vsr1-7(AtVSR1-7) and vsr1-8(AtVSR1-8) are respectively extracted by TRIZOL, first strand cDNA (Invitrogen) is synthesized by an M-MLV reverse transcription kit, and the expression amount of the gene AtVSR1 is detected by a Real-time Quantitative PCR (Real-time Quantitative PCR) method. Using the constitutive expression Actin2 gene as reference, the designed primer sequence is as follows:

RT-AtVSR1-F2：5’-GTGAGTGCCCTACTGTTCAAGGTG-3’；

RT-AtVSR1-R1：5’-GTTGGGTGGTTGACTTTCCAATGGC-3’；

the primer sequences for detecting the expression of the gene Actin2 are as follows:

RT-AtActin2-F：5’-AGCACTTGCACCAAGCAGCATG-3’；

RT-AtActin2-R：5’-ACGATTCCTGGACCTGCCTCATC-3’。

as a result, as shown in FIG. 3C, the expression level of AtVSR1 gene was decreased in the mutant as compared with the wild type plant.

2) Root phenotype of Arabidopsis mutants

Seeds of Arabidopsis thaliana Columbia type 0 (Clo-0) (wild type control), vsr1-7 and vsr1-8 mutants were sterilized with 10% (v/v) aqueous NaClO solution to which 0.01% (v/v) Triton X-100 was added for 15min, and then washed 6 times with sterilized water in a clean bench. Then the seeds are sowed in MS culture medium added with 3.0 percent (w/v) of sucrose and 0.8 percent (w/v) of agar powder for germination culture. The germination culture is firstly processed at 4 ℃ for 2 days, and then transferred to an incubator at 22 ℃ for 12h illumination for culture for 5 days. Then transplanting to MS culture medium for vertical culture for 7 d. The root growth phenotype was observed.

As shown in FIGS. 2A and 2B, the relative root growth (mutant root length/wild-type root length) of vsr1-7(atvsr1-7) and vsr1-8(atvsr1-8) was about 85% of that of the wild type, as compared with the wild type, where Col is the Columbia type 0 (Clo-0) plant of Arabidopsis thaliana as a wild-type control. The root length of the plant with TaVSR1-B gene mutation is smaller than that of the wild type.

2. Wheat Kronos mutant root phenotype identification

1) Wheat Kronos mutant

The mutation information of the tetraploid Kronos mutant of wheat is shown in table 2 below:

TABLE 2 mutation information for tetraploid wheat Kronos mutant

2) Wheat Kronos mutant root phenotype

And (3) assembling the plastic hoses with the diameter of 1m into a soil column, and pre-embedding the soil column into a plastic hard pipe in the field. The Kronos mutants M1, M2 and M3 and wild-type Kronos seeds were sown therein and cultured normally in the field. By the stage of booting, the soil column was removed from the tube and observed for root phenotype.

As shown in FIGS. 2C and 2D, the root depth of Kronos mutant was about 10-12cm shorter than that of wild type. In the figure, M1, M2 and M3 are Kronos mutants; WT is wild-type Kronos. The root depth of the plant with TaVSR1-B gene mutation is smaller than that of the wild type.

Third, phenotype identification of transgenic rice

Will T₃TaVSR1-B rice (OE-8) seeds were transferred and planted in plastic boxes of 90X 35X 30 cm. In the booting stage, washing roots, and observing the root system and the plant phenotype; taking roots, drying and measuring the dry weight of the roots. Wild type rice (WT) and empty vector rice from the T3 generation were used as controls. Each strain was tested for 10 strains, which were repeated three times, and the results averaged.

The results are shown in FIG. 6; wherein, A is a plant phenotype picture, B is a root depth column statistical chart, C is a root dry weight column statistical chart, and T can be seen₃Compared with wild rice, the transgenic TaVSR1-B rice has 4.50cm increased root depth, namely 14.8% increased root depth; the dry root weight increased by 0.108g, i.e., 11.8%.

In FIG. 6, OE-8 is a positive T₃Generation homozygous lines; WT is Kitaake plant as wild type control.

The results of wild rice (WT) and the rice with the empty carrier transferred from the T3 generation have no significant difference.

Fourth, phenotype identification of transgenic wheat

And (3) assembling the plastic hoses with the diameter of 1m into a soil column, and pre-embedding the soil column into a plastic hard pipe in the field. Handle T₃Transferring TaVSR1-B wheat (OE-9) seeds, sowing them, and culturing in field. Until the booting stage, the female parent is reached,the soil column was removed from the tube and observed for root phenotype. And meanwhile, taking roots, drying and measuring the dry weight of the roots. Wild type Wheat (WT) and wheat with empty vector transferred from T3 were used as controls.

The results are shown in FIG. 7, where A is the plant phenotype picture, B is the root depth histogram, C is the root dry weight histogram, T is compared to wild type wheat plants₃The root depth of the transgenic TaVSR1-B wheat plant is increased by 6.51cm, namely the root depth is increased by 8.42%; the dry root weight was increased by 0.049g, i.e., 12.3%.

In FIG. 7, OE-9 is a positive T₃Generation homozygous lines; WT was the Fielder plant as wild type control.

SEQUENCE LISTING

<110> institute of crop science of Chinese academy of agricultural sciences

<120> wheat booting stage root depth related gene TaVSR1-B and coding protein and application thereof

<160> 4

<170> PatentIn version 3.5

<210> 1

<211> 639

<212> PRT

<213> Triticum aestivum

<400> 1

Met Gly Gly Arg Ser Ser Pro Ala Gly Trp Gly Arg Arg Ala Pro Pro

1 5 10 15

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

20 25 30

Val Glu Gly Arg Phe Val Val Glu Lys Asn Ser Leu Arg Val Thr Ser

35 40 45

Pro Ala Gly Leu Arg Gly Val Tyr Glu Cys Ala Ile Gly Asn Phe Gly

50 55 60

Met Pro Gln Tyr Gly Gly Thr Met His Gly Val Val Val Tyr Pro Lys

65 70 75 80

Ala Asn Thr Lys Ala Cys Lys Pro Phe Ala Asp Phe Gly Leu Ser Phe

85 90 95

Asn Pro Lys Ala Gly Gly Leu Pro Val Phe Leu Leu Val Asp Arg Gly

100 105 110

Asp Cys Tyr Phe Thr Thr Lys Gly Trp Asn Ala Gln Thr Ala Gly Ala

115 120 125

Ala Ala Val Leu Val Ala Asp Asp Arg Ala Glu Pro Leu Ile Thr Met

130 135 140

Asp Thr Pro Glu Ser Ser Gly Lys Glu His Leu Glu Asn Ile Thr Val

145 150 155 160

Pro Ser Ala Leu Val Ser Lys Arg Phe Gly Asp Asp Leu Lys Thr Ala

165 170 175

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

180 185 190

Leu Pro His Pro Asp Glu Arg Val Glu Tyr Glu Phe Trp Thr Asn Ser

195 200 205

Asn Asp Glu Cys Gly Ala Lys Cys Asp Met Gln Met Ser Phe Val Arg

210 215 220

Asp Phe Arg Gly Val Ala Gln Val Leu Glu Gln Arg Gly Tyr Thr Gln

225 230 235 240

Phe Ala Pro His Tyr Ile Thr Trp Tyr Cys Pro Glu Ala Phe Val Leu

245 250 255

Ser Ala Gln Cys Arg Ser Gln Cys Ile Asn His Gly Arg Tyr Cys Ala

260 265 270

Pro Asp Pro Glu Gln Asp Phe Thr Thr Gly Tyr Asp Gly Lys Asp Val

275 280 285

Val Val Gln Asn Leu Ile Gln Ile Cys Leu Phe Lys Val Ala Asn Glu

290 295 300

Ser Arg Lys Pro Trp Leu Trp Trp Asp Tyr Val His Asp Phe Ala Ile

305 310 315 320

Arg Cys Pro Met Lys Glu Lys Lys Tyr Thr Thr Asp Cys Ala His Gly

325 330 335

Val Ile Lys Ser Leu Gly Met Asp Ile Asp Lys Ile Thr Gln Cys Val

340 345 350

Gly Asp Pro Asp Ala Asp Glu Asp Asn Pro Val Leu Lys Ala Glu Gln

355 360 365

Asp Ala Gln Ile Gly His Gly Ala Arg Gly Asp Val Thr Ile Leu Pro

370 375 380

Thr Phe Val Val Asn Asn Arg Gln Tyr Arg Gly Lys Leu Asp Lys Arg

385 390 395 400

Ala Val Leu Arg Ala Ile Cys Ser Gly Phe Glu Glu Thr Thr Glu Pro

405 410 415

Asp Ile Cys Leu Thr Gln Asp Ile Gln Thr Asn Gln Cys Leu Glu Asn

420 425 430

Asn Gly Gly Cys Trp Leu Asp Lys Asn Thr Asn Phe Thr Ala Cys Lys

435 440 445

Asp Thr Phe Arg Gly Arg Val Cys Glu Cys Pro Val Val Asn Gly Val

450 455 460

Lys Phe Val Gly Asp Gly Tyr Thr His Cys Glu Ala Ser Gly Val Gly

465 470 475 480

Arg Cys Gln Ile Asn Asn Gly Gly Cys Trp Lys Glu Thr Arg Asn Gly

485 490 495

Lys Ser Val Ser Ala Cys Ser Asn Glu Gln Ala Lys Gly Cys Lys Cys

500 505 510

Pro Gln Gly Phe Lys Gly Asp Gly Ile His Gly Cys Glu Asp Val Asp

515 520 525

Glu Cys Lys Glu Arg Leu Phe Cys Gln Cys Lys Asp Cys Ser Cys Glu

530 535 540

Asn Thr Trp Gly Ser Tyr Glu Cys Gly Cys Gly Gly Ser Asn Met Leu

545 550 555 560

Tyr Met Arg Glu His Asp Thr Cys Ile Ser Lys Val Ala Thr Ser Ser

565 570 575

Val Gly Trp Gly Phe Met Trp Val Ile Phe Phe Gly Leu Gly Phe Ala

580 585 590

Gly Val Gly Ala Tyr Ala Val Tyr Lys Tyr Arg Leu Arg Ser Tyr Met

595 600 605

Asp Ser Glu Ile Arg Ala Ile Met Ala Gln Tyr Met Pro Leu Glu Asn

610 615 620

Gln Glu Thr Ser Ser His Gln Arg His Val Asp His Ala Asp Ile

625 630 635

<210> 2

<211> 2786

<212> DNA

<213> Triticum aestivum

<400> 2

actgcttctc cctcgtcctc tccctcccaa cccccggtca aatccatccg cccttgccac 60

tcaacccacg ctcgcctttt ccgctcagac gccaccctcc tctcttcccg agatcatcga 120

ttcgtccatc gaccctcctc cgcgcgcggc tcgatcgtgt tcgatcgtcg tgtgcagcag 180

ttggccgggg aaggggatcg gatcaccgag cgaggtgatc gagatgggag ggagatcgtc 240

gccggcgggg tgggggcgac gtgcgccgcc gccgccgctt cttcttcttc tcgtgtcggt 300

gctggcgctc gcgacggtgg tggaggggag gttcgtggtg gagaagaaca gcctgcgggt 360

gacgtcaccg gcggggctgc ggggcgtgta cgagtgcgcc atcggcaact tcgggatgcc 420

gcagtacggg ggcaccatgc acggcgtcgt cgtctacccc aaggccaaca ccaaggcctg 480

caagcccttc gccgacttcg gcctctcctt caaccccaag gccggcggcc tccccgtctt 540

cctcctcgtc gaccgcggag actgctactt cacaaccaag gggtggaacg cgcagacggc 600

gggagcggcg gcggtgctcg tcgccgacga cagggcagag cccctcatca caatggacac 660

cccagagtcc agcggcaagg agcacctgga gaacatcacc gtgccctcag ccctggtctc 720

caagcgcttc ggcgacgacc tgaagaccgc cctcgagaac ggcgacatgg tgaacgtgct 780

cctggactgg cgggaatccc tcccgcaccc ggacgagcgc gtggagtacg agttctggac 840

caacagcaac gacgagtgcg gcgcgaaatg cgacatgcag atgagcttcg tccgggactt 900

ccggggcgtg gcgcaggtgc tggagcagcg cggctacacc cagttcgcgc cgcactacat 960

cacctggtac tgcccggagg ccttcgtcct gagcgcgcag tgcaggtcgc agtgcatcaa 1020

ccacggccgc tactgcgccc ccgacccgga gcaggacttc accaccgggt atgatgggaa 1080

ggacgtcgtg gtgcagaact tgatccagat ctgcctcttc aaggtcgcca acgagagccg 1140

caagccgtgg ctgtggtggg actatgtgca cgacttcgcc atccggtgcc ccatgaagga 1200

gaagaagtac accaccgatt gcgctcatgg tgtcatcaag tcccttggaa tggacattga 1260

caagattacc cagtgcgtcg gagaccccga cgccgatgag gacaacccgg tgctcaaggc 1320

ggagcaagat gctcaaattg gtcatggtgc tcgaggggat gttaccatac tgccgacttt 1380

cgtcgtcaac aacagacagt acagagggaa actggataaa agggctgtgc tccgagcgat 1440

atgttcggga ttcgaggaga cgaccgaacc cgatatctgt ttgactcaag atatacaaac 1500

aaaccagtgc ttggaaaaca atggaggttg ctggctggac aaaaatacta atttcactgc 1560

atgcaaggat accttccgtg ggcgggtttg cgagtgcccg gttgtcaatg gcgtcaagtt 1620

cgttggcgac gggtacaccc actgtgaagc ttctggtgtt ggtcgatgcc aaatcaacaa 1680

cggaggctgc tggaaggaga ccaggaacgg caagtctgtc tccgcgtgct cgaatgagca 1740

agctaagggc tgcaaatgtc cgcaaggctt caagggtgat ggcatacacg gttgcgaaga 1800

tgttgatgag tgcaaagaga ggctcttctg ccagtgcaag gactgcagct gcgagaacac 1860

atggggaagc tacgagtgtg gctgcggtgg tagcaacatg ctgtacatga gagagcatga 1920

cacttgcatc agcaaggtcg cgacttcgtc ggtgggctgg gggttcatgt gggtcatctt 1980

cttcggcctc ggcttcgccg gagtgggagc atacgccgtc tacaaatatc ggctacggag 2040

ctacatggat tcagagatcc gcgcgatcat ggcgcagtac atgccgctgg agaaccagga 2100

gacgtcgagc caccaacggc acgtggacca cgccgacatc tgaagcacca aggagttcta 2160

agtacatacg gatcatcaag cataagagct agtctcaccg cgtccgtggc gcatcggagt 2220

ggccgggttt ggagctgcaa ttttgaattc agaaaagatt ggccggttag ctagcttatg 2280

attctgaaag acaaacgtac tctctctttt ctctctttct gaaacttcaa ggacaggttt 2340

aggatcgatc tcaccggagt tttagccctc atttagtcat aattgtgatg ctcgaggtct 2400

cgttcagcgt gatgacattg tgtaaatacc gttgttcgtt cggactgaat cgtgttgggt 2460

gcgtttaata aagagaccag aagagcattt agcctacttc cccaaaaacc acatgttgct 2520

ttgcaggcag taggcttcag ctatcacagt ggtgaggttc tcttttttca cattttctga 2580

tgttctgatt attttttttt attatttttt tctgatgttc tgattagata gtacgtgctc 2640

ccctcgtagc aaattcatga cgtcctgacc ttaccatgta gcatggtccc aagtagacaa 2700

gcataaactt tccatcgtca atgcataggt atgtcttgat aaaaaagatg attaaatgtt 2760

gtgctctgga aatagtacaa tttgtg 2786

<210> 3

<211> 2451

<212> DNA

<213> Triticum aestivum

<400> 3

cggtgggagt actggttaga acacttatgc agtgtaacag aacctggatg caaagtacag 60

ttccggtttt gcaaaatttt gataggtaat agtatatagt tttaattgtt caatcttggt 120

gacatgaatt aactttcact tcctcctgat agtttctgcc gaggcagctt gatttctttg 180

cctttagctt cacataacta agatgtgcat tctgcctaat agctcctgga aggtctcaaa 240

catgatcaat gtagatcctc tctaaaaagg cacttctgtt gaatatgttt tctggcgtat 300

tccattattt ctgactgtta tgcatgaaaa tttggagcat gtatgtgtaa tttttttaat 360

actgtcatgt atgcttacac tgctatgaac ttagtatcta ctccctccgt tccgaattac 420

ttgtcttgga tttgtctaga tacggatgta tctagactta ttttagtgct agatacctcc 480

gtatctagac aaatctaaga caagtaattc ggaacggagg gagtagttct gtacatactg 540

tttattaggt tttacgggct atgttgccgg ctttaaagtc aaatgttagg aatttctttt 600

ggttgaaact ggtacctact gccttgtgat gcgatggaac tttatttcct ctactctgac 660

aaaaatctaa ggatccattc tgccatagtc aacattcagt ggttgaggca atttctatga 720

cttggcctag ctactttaca tagtgagtga gtctgcctgt ctggtacaaa aattcttcct 780

cgcatctttt gtcccctgtg atgtctggtc tttgctgtat tggtgaaaac ctggaactaa 840

aatcatgatg aagatccatg gtgtgcctat gctgtttgtg ttaactctat ttaaccgatt 900

ccattagttg cttgatcaat tccaagtacc gtcgcacctt gcatctttat ttgccctcct 960

caaagtcata attattgttg ctaacctgtt tttcgtctga attttcttgt catggtcatc 1020

tcgctattag tttcagtttg tcagtctcta cgatactgaa gatccttttg attttgtttg 1080

cagccatttg actggatcaa gaaaaccttc ttcgagaagc ccgagcctga ggcataagtg 1140

tctatggggg tgcacggcta cagcttttga tgcgtacatt ttcaggatga ttctctgagt 1200

ttggacaacc cagccaataa tatacaagac cctcgtgatc ccttgtttga ttttccatcc 1260

agatgtaggt ttcctcttgc gttgcatgta accctcacca tgggagggga atgatcatgt 1320

aaatttattt tccgtcacag aggaaagaaa ctgtgctttt gtttatttct cttgctatgg 1380

tcaccaaaga gtgagctttg gctcgctcag ccgtggtatt ttgtgtcgag cctggtgtcc 1440

cttgttgaac taactaactc atcgcgaaag aagaagccca tgagatttac ctccgccgta 1500

ggtagcctta acaagattct aacaagcttg agatcgaaat cacttgtaga actcgacaaa 1560

gatcataacg ccgtgttctc gccttaaaag gaaaaaggtt ttgagaacga tgtgtaccaa 1620

atcgcagcac acttctccgg tggtagtctg cactcttcag ccagaaaacc gcacctttgt 1680

agtctcagtt acaacaacaa agacggagac gggtaacgca ccgactcgaa ccacatttgt 1740

tgtgcacagt catattccca acaactctgc cgccccaaga tgttgtattc atcaaggaaa 1800

atccgcctgc ccatggtttc gatcgatact gaccgtccac gtacgccccc cattgctaga 1860

tccacacaca cgcaaaggcg cccggtgccg gccggaagcc cctccctcac gcgcaccata 1920

catgcatgca atgcacggca cgctctacgt ctttctcctc ggatgagcat gagaatgagc 1980

gtgacagcgt ccgaaaaagg tgtggctttc cccgtcagcg cgcgcgagca cggacacggc 2040

cacaaacaca caccagaagc aaaaggcggg aggcttggaa gcgttccaac ccccacacga 2100

ggaggccgcc acgtacctcg ccctccccct tcctctctct cactccactg cttctccctc 2160

gtcctctccc tcccaacccc cggtcaaatc catccgccct tgccactcaa cccacgctcg 2220

ccttttccgc tcagacgcca ccctcctctc ttcccgagat catcgattcg tccatcgacc 2280

ctcctccgcg cgcggctcga tcgtgttcga tcgtcgtgtg cagcagttgg ccggggaagg 2340

ggatcggatc accgagcgag gtgatcgaga tgggagggag atcgtcgccg gcggggtggg 2400

ggcgacgtgc gccgccgccg ccgcttcttc ttcttctcgt gtcggtgctg g 2451

<210> 4

<211> 4932

<212> DNA

<213> Triticum aestivum

<400> 4

cggtgggagt actggttaga acacttatgc agtgtaacag aacctggatg caaagtacag 60

ttccggtttt gcaaaatttt gataggtaat agtatatagt tttaattgtt caatcttggt 120

gacatgaatt aactttcact tcctcctgat agtttctgcc gaggcagctt gatttctttg 180

cctttagctt cacataacta agatgtgcat tctgcctaat agctcctgga aggtctcaaa 240

catgatcaat gtagatcctc tctaaaaagg cacttctgtt gaatatgttt tctggcgtat 300

tccattattt ctgactgtta tgcatgaaaa tttggagcat gtatgtgtaa tttttttaat 360

actgtcatgt atgcttacac tgctatgaac ttagtatcta ctccctccgt tccgaattac 420

ttgtcttgga tttgtctaga tacggatgta tctagactta ttttagtgct agatacctcc 480

gtatctagac aaatctaaga caagtaattc ggaacggagg gagtagttct gtacatactg 540

tttattaggt tttacgggct atgttgccgg ctttaaagtc aaatgttagg aatttctttt 600

ggttgaaact ggtacctact gccttgtgat gcgatggaac tttatttcct ctactctgac 660

aaaaatctaa ggatccattc tgccatagtc aacattcagt ggttgaggca atttctatga 720

cttggcctag ctactttaca tagtgagtga gtctgcctgt ctggtacaaa aattcttcct 780

cgcatctttt gtcccctgtg atgtctggtc tttgctgtat tggtgaaaac ctggaactaa 840

aatcatgatg aagatccatg gtgtgcctat gctgtttgtg ttaactctat ttaaccgatt 900

ccattagttg cttgatcaat tccaagtacc gtcgcacctt gcatctttat ttgccctcct 960

caaagtcata attattgttg ctaacctgtt tttcgtctga attttcttgt catggtcatc 1020

tcgctattag tttcagtttg tcagtctcta cgatactgaa gatccttttg attttgtttg 1080

cagccatttg actggatcaa gaaaaccttc ttcgagaagc ccgagcctga ggcataagtg 1140

tctatggggg tgcacggcta cagcttttga tgcgtacatt ttcaggatga ttctctgagt 1200

ttggacaacc cagccaataa tatacaagac cctcgtgatc ccttgtttga ttttccatcc 1260

agatgtaggt ttcctcttgc gttgcatgta accctcacca tgggagggga atgatcatgt 1320

aaatttattt tccgtcacag aggaaagaaa ctgtgctttt gtttatttct cttgctatgg 1380

tcaccaaaga gtgagctttg gctcgctcag ccgtggtatt ttgtgtcgag cctggtgtcc 1440

cttgttgaac taactaactc atcgcgaaag aagaagccca tgagatttac ctccgccgta 1500

ggtagcctta acaagattct aacaagcttg agatcgaaat cacttgtaga actcgacaaa 1560

gatcataacg ccgtgttctc gccttaaaag gaaaaaggtt ttgagaacga tgtgtaccaa 1620

atcgcagcac acttctccgg tggtagtctg cactcttcag ccagaaaacc gcacctttgt 1680

agtctcagtt acaacaacaa agacggagac gggtaacgca ccgactcgaa ccacatttgt 1740

tgtgcacagt catattccca acaactctgc cgccccaaga tgttgtattc atcaaggaaa 1800

atccgcctgc ccatggtttc gatcgatact gaccgtccac gtacgccccc cattgctaga 1860

tccacacaca cgcaaaggcg cccggtgccg gccggaagcc cctccctcac gcgcaccata 1920

catgcatgca atgcacggca cgctctacgt ctttctcctc ggatgagcat gagaatgagc 1980

gtgacagcgt ccgaaaaagg tgtggctttc cccgtcagcg cgcgcgagca cggacacggc 2040

cacaaacaca caccagaagc aaaaggcggg aggcttggaa gcgttccaac ccccacacga 2100

ggaggccgcc acgtacctcg ccctccccct tcctctctct cactccactg cttctccctc 2160

gtcctctccc tcccaacccc cggtcaaatc catccgccct tgccactcaa cccacgctcg 2220

ccttttccgc tcagacgcca ccctcctctc ttcccgagat catcgattcg tccatcgacc 2280

ctcctccgcg cgcggctcga tcgtgttcga tcgtcgtgtg cagcagttgg ccggggaagg 2340

ggatcggatc accgagcgag gtgatcgaga tgggagggag atcgtcgccg gcggggtggg 2400

ggcgacgtgc gccgccgccg ccgcttcttc ttcttctcgt gtcggtgctg gcgctcgcga 2460

cggtggtgga ggggaggttc gtggtggaga agaacagcct gcgggtgacg tcaccggcgg 2520

ggctgcgggg cgtgtacgag tgcgccatcg gcaacttcgg gatgccgcag tacgggggca 2580

ccatgcacgg cgtcgtcgtc taccccaagg ccaacaccaa ggcctgcaag cccttcgccg 2640

acttcggcct ctccttcaac cccaaggccg gcggcctccc cgtcttcctc ctcgtcgacc 2700

gcggagactg ctacttcaca accaaggggt ggaacgcgca gacggcggga gcggcggcgg 2760

tgctcgtcgc cgacgacagg gcagagcccc tcatcacaat ggacacccca gagtccagcg 2820

gcaaggagca cctggagaac atcaccgtgc cctcagccct ggtctccaag cgcttcggcg 2880

acgacctgaa gaccgccctc gagaacggcg acatggtgaa cgtgctcctg gactggcggg 2940

aatccctccc gcacccggac gagcgcgtgg agtacgagtt ctggaccaac agcaacgacg 3000

agtgcggcgc gaaatgcgac atgcagatga gcttcgtccg ggacttccgg ggcgtggcgc 3060

aggtgctgga gcagcgcggc tacacccagt tcgcgccgca ctacatcacc tggtactgcc 3120

cggaggcctt cgtcctgagc gcgcagtgca ggtcgcagtg catcaaccac ggccgctact 3180

gcgcccccga cccggagcag gacttcacca ccgggtatga tgggaaggac gtcgtggtgc 3240

agaacttgat ccagatctgc ctcttcaagg tcgccaacga gagccgcaag ccgtggctgt 3300

ggtgggacta tgtgcacgac ttcgccatcc ggtgccccat gaaggagaag aagtacacca 3360

ccgattgcgc tcatggtgtc atcaagtccc ttggaatgga cattgacaag attacccagt 3420

gcgtcggaga ccccgacgcc gatgaggaca acccggtgct caaggcggag caagatgctc 3480

aaattggtca tggtgctcga ggggatgtta ccatactgcc gactttcgtc gtcaacaaca 3540

gacagtacag agggaaactg gataaaaggg ctgtgctccg agcgatatgt tcgggattcg 3600

aggagacgac cgaacccgat atctgtttga ctcaagatat acaaacaaac cagtgcttgg 3660

aaaacaatgg aggttgctgg ctggacaaaa atactaattt cactgcatgc aaggatacct 3720

tccgtgggcg ggtttgcgag tgcccggttg tcaatggcgt caagttcgtt ggcgacgggt 3780

acacccactg tgaagcttct ggtgttggtc gatgccaaat caacaacgga ggctgctgga 3840

aggagaccag gaacggcaag tctgtctccg cgtgctcgaa tgagcaagct aagggctgca 3900

aatgtccgca aggcttcaag ggtgatggca tacacggttg cgaagatgtt gatgagtgca 3960

aagagaggct cttctgccag tgcaaggact gcagctgcga gaacacatgg ggaagctacg 4020

agtgtggctg cggtggtagc aacatgctgt acatgagaga gcatgacact tgcatcagca 4080

aggtcgcgac ttcgtcggtg ggctgggggt tcatgtgggt catcttcttc ggcctcggct 4140

tcgccggagt gggagcatac gccgtctaca aatatcggct acggagctac atggattcag 4200

agatccgcgc gatcatggcg cagtacatgc cgctggagaa ccaggagacg tcgagccacc 4260

aacggcacgt ggaccacgcc gacatctgaa gcaccaagga gttctaagta catacggatc 4320

atcaagcata agagctagtc tcaccgcgtc cgtggcgcat cggagtggcc gggtttggag 4380

ctgcaatttt gaattcagaa aagattggcc ggttagctag cttatgattc tgaaagacaa 4440

acgtactctc tcttttctct ctttctgaaa cttcaaggac aggtttagga tcgatctcac 4500

cggagtttta gccctcattt agtcataatt gtgatgctcg aggtctcgtt cagcgtgatg 4560

acattgtgta aataccgttg ttcgttcgga ctgaatcgtg ttgggtgcgt ttaataaaga 4620

gaccagaaga gcatttagcc tacttcccca aaaaccacat gttgctttgc aggcagtagg 4680

cttcagctat cacagtggtg aggttctctt ttttcacatt ttctgatgtt ctgattattt 4740

ttttttatta tttttttctg atgttctgat tagatagtac gtgctcccct cgtagcaaat 4800

tcatgacgtc ctgaccttac catgtagcat ggtcccaagt agacaagcat aaactttcca 4860

tcgtcaatgc ataggtatgt cttgataaaa aagatgatta aatgttgtgc tctggaaata 4920

gtacaatttg tg 4932

Claims

1. The use of any one of the following 1) to 3) in at least one of the following a) to e);

1) protein TaVSR 1-B;

2) a nucleic acid molecule encoding the protein TaVSR 1-B;

a) regulating and controlling the growth of plant roots;

b) breeding plant varieties with different root depths;

c) cultivating plants with increased root depth;

d) cultivating the plant with increased root weight;

e) cultivating plants with changed root system configurations;

the protein TaVSR1-B is (1) or (2) as follows:

the plant is wheat or rice.

2. Use according to claim 1, characterized in that:

the nucleic acid molecule encoding the protein TaVSR1-B is a DNA molecule of any one of the following 1) -6):

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

2) the coding region is a DNA molecule shown in the 215-2158 th site of the sequence 2 in the sequence table;

3) the coding region is a DNA molecule shown in the 224-2143 th site of the sequence 2 in the sequence table;

4) the coding region is a DNA molecule of the DNA molecule shown in the sequence 4 in the sequence table.

3. A method for promoting the growth of plant roots comprises the following steps 1) or 2):

1) the method comprises the following steps: the content and/or activity of the protein TaVSR1-B in the target plant is improved, and the growth of plant roots is promoted;

the protein TaVSR1-B is (1) or (2) as follows:

the plant is wheat or rice.

4. The method of claim 3, wherein: the promotion of plant root growth is embodied in increasing the root depth and/or root weight of a plant.

5. A method for breeding transgenic plants with increased root depth and/or increased root weight, which comprises the following steps 1) or 2):

the transgenic plant has deeper root and/or bigger root than the target plant;

the protein TaVSR1-B is (1) or (2)

the plant is wheat or rice.

6. The method of claim 5, wherein:

the method for increasing the content and/or activity of protein TaVSR1-B in a target plant or for increasing the expression of a nucleic acid molecule encoding protein TaVSR1-B in a target plant is carried out by introducing the nucleic acid molecule encoding protein TaVSR1-B into the target plant.