CN108676803B - Plant medicago truncatula floral organ stigma exsertion gene and encoded protein and application thereof - Google Patents

Abstract

The invention discloses a plant Medicago truncatula flower organ stigma exposed gene and its encoded protein and application. The gene is a DNA molecule described in any one of (a1)-(a4): (a1) a DNA molecule with a coding region shown in SEQ ID NO.1; (a2) a DNA molecule shown in SEQ ID NO.3; (a3) a DNA molecule that hybridizes to the DNA sequence defined in (a1) or (a2) under stringent conditions and encodes the protein of claim 1; (a4) is derived from Medicago truncatula and is combined with (a1) or (a2) or ( a3) DNA molecules with more than 90% homology in the defined DNA sequence. The gene of the invention can control plant stigma exposure and petal fusion; it lays a theoretical foundation for studying the regulation of CABL2 gene in plant stigma exposure.

Description

Plant medicago truncatula floral organ stigma exsertion gene and encoded protein and application thereof

Technical Field

The invention belongs to the technical field of genetic engineering, and particularly relates to a plant medicago truncatula floral organ stigma exsertion gene, and a protein coded by the same and application thereof.

Background

The flower is the specific reproductive organ of angiosperm, the normal development of the flower organ is directly related to the species multiplication, and leguminous plants are different from arabidopsis thaliana and other radiation symmetrical flowers and are typical sphenoideae plants. The floral organs of the Papilionaceae plant are typically bilaterally symmetrical, and the corolla is butterfly. The flower petal is composed of 5 petals with different sizes and shapes, wherein the bigger and unpaired petal is a flag petal; two sides below the flag flap are respectively provided with a flap which is in a wing shape and is called as a wing flap; the petal is fused into a boat shape and is covered outside the gynoecium and the androecium, which is called as a keel petal.

Heterosis has been exploited in many species and significantly improves crop yield. Alfalfa and soybean have small flower type and complex structure, and the artificial emasculation and pollination is time-consuming, labor-consuming and high in working strength, so that the difficulty and cost of crop crossbreeding are increased. Meanwhile, the cross breeding yield is low and the cost is high due to the low cross breeding rate of the sterile lines, so that the industrialization process of the alfalfa and soybean hybrid is restricted. Therefore, the study of the flower type of leguminous plants has been receiving attention from broad breeders. Therefore, the development of the genetic mechanism analysis of the development of the specific floral organs of the Papilionaceae plants has important theoretical value for the elucidation of the molecular mechanism regulation of the development of the floral organs of higher plants.

Disclosure of Invention

In view of the above, the invention provides a medicago truncatula floral organ stigma exsertion gene, and a protein coded by the gene and application of the gene.

In order to solve the technical problem, the invention discloses a medicago truncatula floral organ stigma exsertion gene of a plant, which is a DNA molecule as shown in any one of the following (a1) - (a 4):

(a1) the coding region is a DNA molecule shown as SEQ ID NO. 1;

(a2) a DNA molecule shown as SEQ ID NO. 3;

(a3) a DNA molecule which hybridizes with the DNA sequence defined in (a1) or (a2) under stringent conditions;

(a4) a DNA molecule which is derived from medicago truncatula and has more than 90% homology with the DNA sequence defined by (a1) or (a2) or (a 3).

The invention also discloses a protein coded by the gene.

Optionally, the protein is as follows (b1) or (b 2):

(b1) a protein consisting of an amino acid sequence shown in SEQ ID No. 2;

(b2) the protein which is obtained by substituting and/or deleting and/or adding one or more amino acid residues of the amino acid sequence shown in SEQ ID NO.2 and is derived from the SEQ ID NO.2 and has the same function.

The invention also discloses a recombinant expression vector, an expression cassette, a transgenic cell line or a recombinant bacterium containing the gene.

The invention also discloses an application of the gene or the protein in regulating and controlling plant stigma exsertion.

The invention also discloses a method for controlling the exposure of the stigma of the plant and the fusion of the petals, which comprises the following steps: inhibiting the activity of the gene and/or the expression level of the protein in the target plant to obtain the transgenic plant with exposed stigma and fused petals.

The invention also discloses application of the gene, the protein or the method in plant breeding.

Compared with the prior art, the invention can obtain the following technical effects:

the invention finds a stigma exsertion Tnt1 insertion mutant from a medicago truncatula Tnt1 insertion mutant library, and clones a control gene CABL2 of the stigma exsertion phenotype through flanking sequence analysis. To demonstrate that CABL2 is a control gene with exposed stigma, we screened a mutant library for another insertion mutant of CABL2 gene and found that the mutant exhibited a phenotype consistent with the previous mutant. The invention discloses a theoretical basis for researching that CABL2 gene regulates and controls stigma exsertion in plants.

Of course, it is not necessary for any one product in which the invention is practiced to achieve all of the above-described technical effects simultaneously.

Drawings

The accompanying drawings, which are included to provide a further understanding of the invention and are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and together with the description serve to explain the invention and not to limit the invention. In the drawings:

FIG. 1 is a phenotypic analysis of the cabled 2 mutant of the present invention; wherein, A-D is a phenotype graph of flowers of different wild flowers in different flower development periods; F-H is a floral phenotype diagram of the cable 2 mutant flowers at different periods of development; forming normal fruit pods after D-E wild type flower pollination is completed; I-J cablel 2 mutant;

FIG. 2 is a fertility analysis of the cabled 2 of the present invention;

FIG. 3 shows the gene structure diagram of CABL2 and the detection of CABL2 gene expression level in the mutant of the present invention.

Detailed Description

The following embodiments are described in detail with reference to the accompanying drawings, so that how to implement the technical features of the present invention to solve the technical problems and achieve the technical effects can be fully understood and implemented.

Medicago truncatula R108 (wild type): the nobelfoundation.

cable 2 mutant: the nobelfoundation. cab 2-1(NF19675) cab 2-2(NF 14424).

Example 1

Acquisition and phenotypic analysis of first, cabled 2 mutants

The mutant NF19675 with stigma exserted, petal fold and fusion and stamen wrapped in the petal is obtained by screening Medicago truncatula Tnt1 and inserting into mutant library, and is named as cabl2(cabbage-like)。

The observation of the floral phenotype of the cabl2 mutant and medicago truncatula R108 (wild type) is shown in FIG. 1. In FIG. 1, A-D are phenotypic graphs of flowers during different flower development stages of the wild type; F-H is a floral phenotype diagram of the cable 2 mutant flowers at different periods of development; forming normal fruit pods after D-E wild type flower pollination is completed; the I-J cablel 2 mutant has stamens wrapped due to petal fusion, and can not complete normal pollination and finally can not form normal fruit pods.

Second, mutant fertility analysis

To study fertility of mutant stigma due to mutant stigma exposure, we hybridized the mutants as shown in fig. 2: a is selecting cabl2 buds with a constant growth state, and awarding wild pollen to one of the buds; after B4 days, the pollinated buds begin to grow a pod spiral structure, and buds which are not pollinated with pollen are dried; after 15 days C, the pollinated bracts grow into normal fruit pods which are the same as the wild type, and the non-pollinated bracts wither and die. Indicating that the stigma of the cable 2 mutant still has biological activity. Provides convenience for plant hybridization.

Example 2 cloning of CABL2 Gene

Corresponding primers are designed according to the flanking sequence of NF19675 published by Tnt1 website (https:// media a go-mutant. noble. org/mutant /), and the linkage of the mutant and the gene is detected. Finally, a gene CABL2 linked with the mutant is discovered;

NF19675-F TCATGGACGTTAGTTGGAAA；

NF19675-R AAGCACATTACCAGACATGA；

the NF19675 mutant has Tnt1 inserted in the second exon of the gene. To demonstrate that the CABL2 gene is the control gene for the mutant, we ordered another mutant of this gene: NF14424, inserted in the third exon. And the previous NF19675 was named cabl2-1 (see FIG. 2A). The phenotype of the cabl2-2 homozygous mutant was observed and found to be identical to the cabl2-1 phenotype. In the CABL2-1 and CABL2-2 homozygous mutants, the full-length expression of CABL2 gene was completely deleted (FIG. 2B). The CABL2 gene was demonstrated to be the control gene for the stigma exserted phenotype.

Example 3 obtaining of CABL2 Gene and its encoded protein CABL2

1. RNA from alfalfa R108 (wild-type) flourishing flowers was extracted and reverse transcribed into cDNA.

2. And (3) taking the cDNA obtained in the step (1) as a template, and amplifying by adopting a primer pair consisting of a primer CABL2-F and a primer CABL2-R to obtain a PCR amplification product.

CABL2-F：5’-caccATGGAGATAGAGCAAGACAATGCC-3’；

CABL2-R：5’-ATGAAATGGAGAGTTGAGACCCTCC-3’。

3. Sequencing the PCR amplification product obtained in the step 2 to obtain a coding region sequence of the target gene, such as SEQ ID NO: 1, SEQ ID NO: 2. Consists of 689 amino acid residues. The CDS of the gene is compared with a Medicago truncatula website (http:// blast.jcvi. org/Medicago-Blast /), so as to obtain a genome sequence of CABL2 gene, such as SEQ ID NO: 3, respectively. The sequence of SEQ ID NO: the protein shown in 2 was designated as CABL2, and the gene encoding CABL2 was designated as CABL gene.

While the foregoing description shows and describes several preferred embodiments of the invention, it is to be understood, as noted above, that the invention is not limited to the forms disclosed herein, but is not to be construed as excluding other embodiments and is capable of use in various other combinations, modifications, and environments and is capable of changes within the scope of the inventive concept as expressed herein, commensurate with the above teachings, or the skill or knowledge of the relevant art. And that modifications and variations may be effected by those skilled in the art without departing from the spirit and scope of the invention as defined by the appended claims.

Sequence listing

<110> institute of biotechnology of Chinese academy of agricultural sciences

<120> a plant medicago truncatula floral organ stigma exsertion gene and coded protein and application thereof

<130> 2018

<160> 7

<170> SIPOSequenceListing 1.0

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<211> 2070

<212> DNA

<213> Medicago truncatula (Medicago truncatula)

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ccaaactttg ggaatggaca tacgagaaga ttgcttaatg gtttgaatgg atatgttgaa 180

ccaaacagaa tcatggctat tatgggtcca tctggttctg gaaaatctac tcttcttgat 240

gctttggcag gtagactctc tggaaatgtt atcatgtctg gtaatgtgct tttaaatgga 300

aagaaaagga gattagacta tggggttgta gcttatgtga cacaagaaga catattgttg 360

ggaacactaa cagtgagaga gacaatatct tactcagcaa atttaaggct tccagctaaa 420

atgacaaaag aagaggtaaa tgagattgta gaaggaacaa taatggagat gggtcttcaa 480

gattgtgctg atagacttat aggaaattgg cacttaagag gtataagtgg tggagaaaag 540

aaaagaacta gcatagcact tgaaattctt actaggccat gtctcttgtt ccttgatgag 600

ccaacaagtg gtttggacag tgcctcagca tattttgttg ctcaaacttt gagaaacatt 660

gctcatgatg ggaaaactgt tatttcttca attcatcaac caagtagtga ggtttttgca 720

ttatttgatg atctatttct actttctggg ggtcaaacaa tttactttgg accagctgaa 780

aatgcagttg agttctttgg caaagcagga ttcccatgtc caagtagaag aaacccttca 840

gatcatttcc tacgttgtat taattcagac tttgatactg tcacaactac tatgatgtcc 900

tccgggagaa cacatgagcc aaaaacattg gcatcaccaa caatgaactt atcaacagca 960

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gcaagaatca aacaaatctc aaattttgaa gtacatattt atgaaagtaa aagcaagagc 1080

caagccaaat ggttgaagca actatcaact ttgactcata gatcttttgt caacatgtct 1140

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ggaacaatct tttatgacat tggatcaagt tatacagcca ttttagcaag aggagcatgt 1260

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tcatacatca attatggagc atggggtttg cagggagctt tcaagaatga tatgattggg 1740

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cctttccttc acagtatcta cacaaagcaa acactgcaac gtatcaagaa gcgggcatca 1980

ttcaggaaaa cgccatcgtt cgcttccaag agacaccaac cattgcatcc attgtcttct 2040

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Trp Glu Asp Leu Ser Val Val Ile Pro Asn Phe Gly Asn Gly His Thr

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Arg Arg Leu Leu Asn Gly Leu Asn Gly Tyr Val Glu Pro Asn Arg Ile

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Met Ala Ile Met Gly Pro Ser Gly Ser Gly Lys Ser Thr Leu Leu Asp

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

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Leu Leu Asn Gly Lys Lys Arg Arg Leu Asp Tyr Gly Val Val Ala Tyr

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

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Ile Ser Tyr Ser Ala Asn Leu Arg Leu Pro Ala Lys Met Thr Lys Glu

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Glu Val Asn Glu Ile Val Glu Gly Thr Ile Met Glu Met Gly Leu Gln

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Asp Cys Ala Asp Arg Leu Ile Gly Asn Trp His Leu Arg Gly Ile Ser

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Gly Gly Glu Lys Lys Arg Thr Ser Ile Ala Leu Glu Ile Leu Thr Arg

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Pro Cys Leu Leu Phe Leu Asp Glu Pro Thr Ser Gly Leu Asp Ser Ala

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Ser Ala Tyr Phe Val Ala Gln Thr Leu Arg Asn Ile Ala His Asp Gly

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Lys Thr Val Ile Ser Ser Ile His Gln Pro Ser Ser Glu Val Phe Ala

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Leu Phe Asp Asp Leu Phe Leu Leu Ser Gly Gly Gln Thr Ile Tyr Phe

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Gly Pro Ala Glu Asn Ala Val Glu Phe Phe Gly Lys Ala Gly Phe Pro

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Cys Pro Ser Arg Arg Asn Pro Ser Asp His Phe Leu Arg Cys Ile Asn

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Ser Asp Phe Asp Thr Val Thr Thr Thr Met Met Ser Ser Gly Arg Thr

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His Glu Pro Lys Thr Leu Ala Ser Pro Thr Met Asn Leu Ser Thr Ala

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Glu Ile Lys Ala Ile Leu Ile Glu Lys Tyr Arg Trp Ser Glu Tyr Ala

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Thr Cys Ala Arg Ala Arg Ile Lys Gln Ile Ser Asn Phe Glu Val His

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Ile Tyr Glu Ser Lys Ser Lys Ser Gln Ala Lys Trp Leu Lys Gln Leu

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Ser Thr Leu Thr His Arg Ser Phe Val Asn Met Ser Arg Asp Val Gly

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Gly Thr Ile Phe Tyr Asp Ile Gly Ser Ser Tyr Thr Ala Ile Leu Ala

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Arg Gly Ala Cys Gly Ala Phe Ile Ser Gly Phe Met Thr Phe Met Ser

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Ile Gly Gly Phe Pro Ser Phe Ile Glu Glu Met Lys Val Phe Tyr Lys

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Glu Arg Leu Asn Gly Tyr Tyr Gly Ile Ala Val Tyr Ile Leu Ser Asn

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Phe Leu Ser Ser Phe Pro Phe Ile Ala Val Met Ser Phe Ala Thr Gly

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Ser Ile Thr Tyr Tyr Met Val Lys Phe Arg Ser Glu Phe Ser His Leu

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Leu Tyr Ile Cys Leu Asp Leu Leu Gly Cys Ile Ala Val Val Glu Ser

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Ser Met Met Ile Ile Ala Ala Leu Val Pro Asn Phe Leu Met Gly Leu

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Ile Ile Gly Ala Gly Tyr Ile Gly Leu Met Met Met Thr Ala Gly Tyr

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Phe Arg Gln Ile Arg Asp Leu Pro Lys Phe Phe Trp Arg Tyr Pro Ile

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Ser Tyr Ile Asn Tyr Gly Ala Trp Gly Leu Gln Gly Ala Phe Lys Asn

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Asp Met Ile Gly Met Glu Phe Asp Pro Glu Asn Pro Gly Glu Pro Lys

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His Arg Val Ile Phe Phe Phe Ile Leu Lys Phe Lys Glu Arg Val Val

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ccaaactttg ggaatggaca tacgagaaga ttgcttaatg gtttgaatgg atatgttgaa 180

ccaaacagaa tcatggctat tatgggtcca tctggttctg gaaaatctac tcttcttgat 240

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cttgcattct atgtatatga tttttatttt tatttttagg tcattcatgt tggtttttgt 360

attaaataac gcacttcaaa ttatatatat atatagacag acatctattc tctctaacta 420

tttgattaaa aaataaataa aaactctata tctctttttt actaactaca ttatttttaa 480

aatgtataca tctcttcaaa tgaactaagt cttctctatc atcttgttcc cttttgtata 540

ttgcaaagtg gggtgagtta tataatattc ttgttgctat gaggggttgt gtgcttcaaa 600

tcaccctata actgaaggaa ggcctaagta aacgtgatta ctgaagaaaa aagaatatat 660

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tgtggctttt acgttagaaa tatcttggga agattatttt tttaaaaaaa tcttgggaag 1020

tgggtgcaaa catgagtcta ttggcgcaga tgtgaaacca aacacataat aagcaattgt 1080

tgcaattgtt ttcttctcac gaccacaaaa tctatgagga tttttatttg gtgtccgaaa 1140

caataatgtt atgttgacac ccttttttcc ctacatccat gtgaaaggac attttaataa 1200

tttcacacca tgtaaaagag agtgttaggt gggatggtga gggtcacata gccacgtgtc 1260

acatttatgt gtggtggtaa aagagatatg tgccacctat gatggtcaac ttatcaccac 1320

tctatctaaa agcatttcgg tacgaaattg agctaataca aagcggttat aatttttctt 1380

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cccaactttt cggattacta gtgttcatcc tctataaatc ggatgattaa caacaacaaa 1500

aattgtacaa aaaaaaaaaa aagaaatatt taagtccaaa tattttttat aatgttttga 1560

actgtataaa atctattgtt tctcgatcgt tgttaagtat taatattttt aaaatattaa 1620

catatgttat tagagcactt gttaatatat gttaatagtg cacttgttag catactaaaa 1680

atagaaattt tgtcagtgtt tctcgaacga tttgttgaaa ggattcaaac ctcttgtcac 1740

cagaagcaat ccattaatgt cggtggttga ttgacattct attcgttggt tcacttgaac 1800

catttccaag ccaacatcca tctaccatca actcatagtt agatatgttg aaaaatataa 1860

tataactgaa atggaaatgg aaatgttaag cctcccaatt tataattatg gcttttacta 1920

tatatttttt attctataac attttattat tatatttttc tacatagaag aatagaattt 1980

atagctttat atttcatctc attcattttt taaattatag aatgataatt ttgttaaatt 2040

tcattctaaa attttgaaat aatatgacca aactttttat ttcataaaat aaaatcaaaa 2100

tattgtttta tttcactttc ttctgccgca tttttttatc tttctaaaaa ccattttcaa 2160

atccacacct taatttatta agaacaattc cattccgttt cattccattt tattccgttc 2220

atcacattca ttttgctcat tatagtcatt tcattatgtc catatttagc tagaaagcca 2280

ttttattttt tattcatata ttagcttttg cattccctta gcattttatc cacagtagac 2340

tagattaaaa gttcatacta aatttgccgg tacaaggttg aaatcaaatg atgtagattc 2400

atcatcaaat gtaaaatgat gagcacatga agccattaaa ttgtacctgc attattcttc 2460

aattatttat tttcttcaaa agaatggagc aaaagcatgg tatttttttt aagatcgata 2520

aaaattagca gaatattata tatatatata tatatatata tagcgacata catacactgt 2580

ttatttattt caagatgtga attgggaata aagaaaaatg catttgtggg ttcataaaca 2640

tctataccac acccaacaaa caagacctac tcacactcct atgacccatg ttactttatc 2700

tatcctataa aagataaata aagttcaatt attcagcggc agttcataac taacatagta 2760

ctaacatcta ctcttgacag tgagaattta atatttgatt tcattgtcat ggacgttagt 2820

tggaaagaaa aaaaaaaatg catgtataat aatatatgag caaaatgcat gtacttgtac 2880

atttagtgtt gtccaatcca aacaaagcaa aatgcatgta tatatatgtt tgaatatatg 2940

tactataaca aaaattggtg ggtgaatgct aattaataat ctaaagtcct agaacctaag 3000

ccgttatttt ggacataagg taatgaacaa attaagaatt ttataactag attagtttat 3060

gttattgaga tttcttatga acaaattacc aatgctatcg atttcgattt caacattgga 3120

tatataattg agatttgttg catttcaggt agactctctg gaaatgttat catgtctggt 3180

aatgtgcttt taaatggaaa gaaaaggaga ttagactatg gggttgtagt aagtaactaa 3240

catcttacat acatataatc cactttttaa gtcattatat tcttttataa atttatgata 3300

atgagtatta accagcaaag aataaattaa caatgtccta tatataataa tatgttttta 3360

ttatgtatga aacactatat aggcttatgt gacacaagaa gacatattgt tgggaacact 3420

aacagtgaga gagacaatat cttactcagc aaatttaagg cttccagcta aaatgacaaa 3480

agaagaggta aatgagattg tagaaggaac aataatggag atgggtcttc aagattgtgc 3540

tgatagactt ataggaaatt ggcacttaag aggtataagt ggtggagaaa agaaaagaac 3600

tagcatagca cttgaaattc ttactaggcc atgtctcttg ttccttgatg agccaacaag 3660

tggtttggac agtgcctcag catattttgt tgctcaaact ttgagaaaca ttgctcatga 3720

tgggaaaact gttatttctt caattcatca accaagtagt gaggtttttg cattatttga 3780

tgatctattt ctactttctg ggggtcaaac aatttacttt ggaccagctg aaaatgcagt 3840

tgaggtacat tttttaatat caatctctcc ctctcatatt tactttcatt atttattcca 3900

attttcaatt aaaaatcact gctgttttaa agaccaaaat atcattgtac cctggaccgt 3960

tatcaagtca ccatatattt gcaactactt tgatataaag attttttttt tcttagaggt 4020

ttaataaatg aagataacat tgagtaaaac gtaataaata ttctatttgt attttaaaat 4080

aacaaataat acttatcaga cagtggaata tttctaatgt gacacacgaa atattcattt 4140

atttattcat gtgttgggaa ctatggtcgg tccctcttgt tcccttcaaa aaaaaaacta 4200

tggcccctct tgttagcttc tatgattagt gctttaatta agggaggatg cattccaaac 4260

tcaaagtttt gggatttgaa tatctcacga gtcatgtgac aaaattcaca tgaaggggtc 4320

aagtgcaatt aaaatttata actagattaa tttcttgcaa tctagttata aattttacta 4380

cattaaaatg tacatacaag tttgatgttt ttctcctata acgaatctta atcatataaa 4440

cactgacata atttatatat ccatgtattt acatgcagtt ctttggcaaa gcaggattcc 4500

catgtccaag tagaagaaac ccttcagatc atttcctacg ttgtattaat tcagactttg 4560

atactgtcac aactactatg atgtcctccg ggagaacaca tgtatataac aatttctcat 4620

atttattatt aaccaattct ttgtatctca ctatctccac tattaattct tttgaatgca 4680

ggagccaaaa acattggcat caccaacaat gaacttatca acagcagaaa tcaaagcaat 4740

actcatagag aaataccgtt ggtcagaata tgcaacttgt gcaagagcaa gaatcaaaca 4800

aatctcaaat tttgtaagta gccctttact cataggatat tgttattcaa acatgaacat 4860

tttatatgta aatgtaaagg ccaaggaatc attttaatta caggaagtac atatttatga 4920

aagtaaaagc aagagccaag ccaaatggtt gaagcaacta tcaactttga ctcatagatc 4980

ttttgtcaac atgtctagag atgtagggta ctattggata aggttaacaa tctatgttgc 5040

cttatctttg tgtgtcggaa caatctttta tgacattgga tcaagttata cagccatttt 5100

agcaagagga gcatgtggcg cttttatctc gggtttcatg acattcatgt ccataggagg 5160

attcccatca ttcatagagg aaatgaaggt atcaaatcaa atgctgcgct atcgcaaatt 5220

tcgtcgtctc taacatttag tgatggaatt aaagacatgt tttattctgt ctctaattct 5280

tgtcttatgc aggtcttcta caaagaaagg ctgaatggat attatggaat tgcagtctac 5340

attctgtcaa attttctctc ttcatttcct tttatagcag tgatgtcatt tgctacaggg 5400

tctataacat attacatggt taagtttcgt tctgagtttt cgcatctttt gtatatctgt 5460

ttggatcttt tgggctgtat agctgttgtg gagagctcaa tgatgattat agctgcattg 5520

gttcccaatt ttcttatggg attgataatt ggagcaggtt atattgtaag taccatcagc 5580

ctttatattt tgaggaaaaa aaagatatgt acggtccgtg taaactagtt ttatatcatc 5640

gttcaataga aatagagtat ttaatagtta tctttaaaag atgggtacga gataggcctg 5700

tatggcaaga tggtgagttt ctattggatg acggtgtaaa agtatttaca ctttcagtgc 5760

atatccacta aactcaaatc atcggatcat taaaaacgtt caactttaat cataattatc 5820

tttaaaatcg catatatgat ttactatgat ttgatgacag tgtaaaactt ttgacaataa 5880

cagtgtataa aaattaaact cgattatggt tgctaatgta aaacatatat gtgtacaggg 5940

tcttatgatg atgactgctg gctatttccg gcagattcgt gatcttccca aattcttctg 6000

gcgttaccct atttcataca tcaattatgg agcatggggt ttgcaggtaa attcaagagt 6060

atcaaaattc tcaaacaata taagaaagaa aatctgaagc acacaatcaa aaggatatga 6120

aatggcgact taattatcgc tttgcaaacg ctttttaatt aagaacttat ttaacaggta 6180

cttatcatat attggttcat gtataaatca tgtctactat caaagagaaa atgagggtaa 6240

actcctcata cactttgtat atttaacttc tataattggg agtttataaa aacaaatttt 6300

aagacattac aaaatatttt tgataattgg gagtgtgtga cataacctta gtgattatgg 6360

gcaatgcagg gagctttcaa gaatgatatg attgggatgg agtttgatcc tgaaaaccct 6420

ggtgagccaa aactgaaagg agaaatcatc ctcaaaacat tgcttggaat gaaagttgat 6480

tattcaaaat ggtgggacct agctgtggtc atcttcatcc tcatgttgca tagagttatt 6540

ttctttttca ttctcaagtt taaggagaga gttgtacctt tccttcacag tatctacaca 6600

aagcaaacac tgcaacgtat caagaagcgg gcatcattca ggaaaacgcc atcgttcgct 6660

tccaagagac accaaccatt gcatccattg tcttctcagg agggtctcaa ctctccattt 6720

cattag 6726

<210> 4

<211> 28

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 4

caccatggag atagagcaag acaatgcc 28

<210> 5

<211> 25

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 5

atgaaatgga gagttgagac cctcc 25

<210> 6

<211> 20

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 6

tcatggacgt tagttggaaa 20

<210> 7

<211> 20

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 7

aagcacatta ccagacatga 20

Claims (6)

1. A gene, characterized by: the gene is a DNA molecule as described in any one of (a1) - (a2) below:

(a1) the coding region is a DNA molecule shown as SEQ ID NO. 1;

(a2) DNA molecule shown in SEQ ID NO. 3.

2. The protein encoded by the gene of claim 1, which is represented by (b 1):

(b1) a protein consisting of an amino acid sequence shown in SEQ ID NO. 2.

3. A recombinant expression vector, expression cassette or recombinant bacterium comprising the gene of claim 1.

4. Use of the gene of claim 1, or the protein of claim 2, for modulating the exposure of the stigma of Medicago truncatula.

5. A method for controlling the exposure of the stigma and the fusion of the petals of medicago truncatula is characterized by comprising the following steps: inhibiting the activity of the gene of claim 1 in a plant of interest.

6. Use of the gene according to claim 1 or the protein according to claim 2 or the method according to claim 5 in alfalfa Tribulus terrestris breeding.

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