CN108410880A - A kind of pellet Bosnia-Herzegovena soybean citrate transporter protein gene and its application - Google Patents

A kind of pellet Bosnia-Herzegovena soybean citrate transporter protein gene and its application Download PDF

Info

Publication number
CN108410880A
CN108410880A CN201810075636.0A CN201810075636A CN108410880A CN 108410880 A CN108410880 A CN 108410880A CN 201810075636 A CN201810075636 A CN 201810075636A CN 108410880 A CN108410880 A CN 108410880A
Authority
CN
China
Prior art keywords
gmmate75
herzegovena
bosnia
soybean
leu
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
CN201810075636.0A
Other languages
Chinese (zh)
Other versions
CN108410880B (en
Inventor
李昆志
郝竞
郝竞一
吴远双
陈丽梅
徐慧妮
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Kunming University of Science and Technology
Original Assignee
Kunming University of Science and Technology
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Kunming University of Science and Technology filed Critical Kunming University of Science and Technology
Priority to CN201810075636.0A priority Critical patent/CN108410880B/en
Publication of CN108410880A publication Critical patent/CN108410880A/en
Application granted granted Critical
Publication of CN108410880B publication Critical patent/CN108410880B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
    • C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
    • C12N15/8261—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield
    • C12N15/8271—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance

Landscapes

  • Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Biophysics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • Biochemistry (AREA)
  • Wood Science & Technology (AREA)
  • General Health & Medical Sciences (AREA)
  • General Engineering & Computer Science (AREA)
  • Biotechnology (AREA)
  • Biomedical Technology (AREA)
  • Botany (AREA)
  • Physics & Mathematics (AREA)
  • Cell Biology (AREA)
  • Plant Pathology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Microbiology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Medicinal Chemistry (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

The invention discloses a kind of red Bosnia-Herzegovena soybean citrate transporter protein geneGmMATE75, nucleotide sequence such as SEQ ID NO:Shown in 1, coding such as SEQ ID NO:Amino acid sequence shown in 2 applies red Bosnia-Herzegovena soybean citrate transporter protein gene in improving Aluminum Tolerance in Plants ability；Experiments have shown that red Bosnia-Herzegovena soybean citrate transporter protein geneGmMATE75Have the function of improving tobacco plant Aluminum toxicity；GmMATE75The application value that gene has in acid soil Genes For Plant Tolerance Al toxicity stress can be used for Aluminum Tolerance in Plants molecular mechanism research and crop aluminum-resistant poison field of molecular breeding.

Description

A kind of pellet Bosnia-Herzegovena soybean citrate transporter protein gene and its application
Technical field
The present invention relates to molecular biology and genetic engineering correlative technology fields, are related to a kind of red Bosnia-Herzegovena soybean citric acid turn Transport protein geneGmMATE75And its application in improving plant aluminum resistance.
Background technology
Acid-Al stress is one of the most important factor that plant growth is influenced on acid soil.Aluminium as content in the earth's crust the most One of abundant mineral element is distributed widely among soil.Under normal circumstances, aluminium does not have larger shadow to the growth of crop It rings, but with the reduction of pH, aluminium can be present in toxic ionic forms in soil.As Al in acid soil3+It is dense When degree reaches micro-molar concentration, suction of the crop to moisture and mineral nutrient will be influenced by way of inhibiting crop root growth It receives, to inhibit the growth of crop, leads to dropping in production over a large area for crop on acid soil.
In traditional liming and the method for acid soil can alleviate the acidity of soil to a certain extent, to reduce Aluminium poison is poisoned.But this method cost is higher, and the acidity that can not implement on a large scale, and can only neutralize surface soil plays temporarily Effect, and cannot fundamentally solve the problems, such as aluminium poison.Therefore, from the machine that aluminium poison is poisoned on molecular level analysis acid soil Reason, cultivating aluminum-resistant poison crops with genetic engineering the relevant technologies has important theory value and practice significance.
The unifacial leaf of most of resistance to aluminium and dicotyledon can discharge organic acid by root system and cope with Acid-Al stress；Plant The organic acid of root secretion can be with the Al in apoplast3+It is combined into Chelating state, and is discharged it, to reduce or exclude Al Toxicity；More organic acids are usually secreted than aluminium poison sensitive strain by the drugs of the resistance to aluminium system of same species.
Citrate transporter albumen, that is, MATE albumen belongs to a big gene man in prokaryotes and eucaryote Race transports secondary metabolite and toxin by ion electrochemical gradient.The SbMATE genes of sorghum correspond to the master of aluminum-resistant poison Want the sites QTL AltSB, it is the aluminum-resistant virus gene of the first MATE family obtained through map based cloning, the mistake in arabidopsis It can cause Al when amount expression3+It is arranged outside the citric acid of activation, while with the increase of aluminum-resistant poison ability.In addition, HvAACT1 is also compiled Code MATE albuminoids, in the root tip position constitutive expression of aluminum-resistant virus gene type barley, Al3+It can activate and be arranged outside HvAACT1 In addition citric acid has the secretory volume positive correlation of the ability and its root system citric acid of 10 different barley strain aluminum-resistant poison, explanation HvAACT1 is as Al3+The citrate transporter albumen of activation plays an important role in terms of barley aluminum-resistant poison.One in wheat The expression of the express sequence tag of MATE genes is related to the phenotype of this segregating population citrate exudation, illustrates in wheat There is also the similar mechanism that Al toxicity stress is fought by secreting citric acid.
Invention content
The object of the present invention is to provide a kind of red Bosnia-Herzegovena soybean citrate transporter protein geneGmMATE75, nucleotides sequence Row such as SEQ ID NO:Shown in 1, full length gene 1288bp, coding such as SEQ ID NO:Amino acid sequence shown in 2.
The present invention obtains citrate transporter protein gene from the soybean of Aluminum toxicity pellet Bosnia-HerzegovenaGmMATE75, special by PCR The complete segment of the gene is amplified；Recovery purifying GmMATE75 genetic fragments, are connected on pMD18-T carriers, turn Change bacillus coli DH 5 alpha, extraction plasmid obtains recombinant vector pMD18-T-GmMATE75 after digestion detects；Pass through digestion PMD18-T-GmMATE75 carriers are simultaneously attached with carrier pENTR-2B, convert bacillus coli DH 5 alpha, and extract plasmid acquisition Entry clones carrier pENTR-2B-GmMATE75；It is reacted by the LR of Gateway technologies and GmMATE75 genes is subcloned into plant In object expression vector pK2GW7, plant expression vector pK2-35S-GmMATE75 is obtained, which contains enhanced promoter, can The overexpression target gene in recipient plant.
Using agrobacterium tumefaciens-mediated transformation, pK2-35S-GmMATE75 carriers are transferred to Agrobacterium competent cell first, Target gene is transferred in receptor tobacco plant with dip method later, overexpression is obtained using the method for Plant Tissue Breeding The transgenic plant parts of the gene, and verify whether the gene has the spy for improving plant aluminum-resistant poison ability by further experiment Property；The result shows that the tobacco of the overexpression gene has the stronger poison of resistance to aluminium characteristic relative to wild-type tobacco.
Resistance of the plant to Acid-Al stress can be improved in gene provided by the present invention, while the invention can be used for the plant aluminum-resistant side of body Compel the research of molecule mechanism and the field of molecular breeding of aluminum-resistant stress plant.
Description of the drawings
Fig. 1 is the electrophoretogram that GmMATE75 genetic fragments are expanded from aluminium treated red Bosnia-Herzegovena soybean, and M is DNA in figure Marker, 1 is GmMATE75 genetic fragments；
Fig. 2 is root specific elongation rate measurement result schematic diagram of the GmMATE75 types tobacco under Acid-Al stress；
Fig. 3 is GmMATE75 types tobacco soluble sugar content measurement result schematic diagram in cell after aluminium is handled, and wherein A figures are leaf Polyoses content in piece, B figures are root polyoses content；
Fig. 4 is GmMATE75 types tobacco content of hydrogen peroxide measurement result schematic diagram in cell after aluminium is handled, and wherein A figures are leaf Content of hydrogen peroxide in piece, B figures are root content of hydrogen peroxide.
Specific implementation mode
Test method in embodiment described below is conventional method unless otherwise specified, and agents useful for same is such as without special Explanation is conventional commercial reagent and the reagent that configures according to a conventional method.
Embodiment 1：The structure of GmMATE75 transgene tobacco strains
Red Bosnia-Herzegovena soybean water planting seedling is taken, with the AlCl of 50 μM of pH4.53Solution treatment for 24 hours, takes its blade guanidinium isothiocyanate Method extract total serum IgE, use reverse transcriptase M-MLV (promega) using total serum IgE as the first chains of templated synthesis cDNA, reaction system with Operating process is：5 μ g Total RNA are taken, 50 ng oligo are sequentially added（dT）, 2 μ L dNTP（2.5mM each）、 DEPC water to reaction volume is 14.5 μ L；It is rapid after 70 DEG C of 5 min of heat denatured to cool down 5 min on ice after mixing, then Sequentially add 4 μ L 5 × First-stand buffer, 0.5 μ L RNasin（200U）、1 μL M-MLV（200U）, mixing And centrifuge in short-term, 42 DEG C of 1.5 h of warm bath, 70 DEG C of 10 min of heating, terminate reaction after taking-up；The first chains of cDNA synthesis be placed on- 20 DEG C save backup.
PCR amplification GmMATE75 genes are carried out using the cDNA of synthesis as template with GmMATE75 specific primers；
It is as follows to design specific primer：
MA75-F：CGGGATCCCGATGGACGAGAATAGAAGTTCCAAC, GGATCC are BamH I restriction enzyme sites；
MA75-R：CCGCTCGAGCGGTCATTTGCAGTGTCCTTGTTGCTG, CTCGAG are XhoI restriction enzyme sites；
PCR reaction conditions：95℃4 min；95 DEG C of 30 s, 54 DEG C of 30 s, 72 DEG C of 1min, 30 cycles；72℃10 min；Reaction System（20 μL）For 1 μ L cDNA, 2 10 × Advantage of μ L, 2 PCR Buffer, 1.8 μ 50 × dNTP of L Mix （10mM each）, 0.2 μ L forward primers（10 μM）, 0.2 μ L reverse primers（10 μM）、0.2 μL Advantage 2 PCR Polymerase Mix、14.6μL PCR-Grade water；After PCR, take 5 μ L for agarose gel electrophoresis, with inspection The specificity and size for surveying amplified production, are as a result consistent with target gene size（See Fig. 1）.
The DNA fragmentation is obtained using Ago-Gel absorption method, is connected to pMD18T cloning vectors, the kit used is pMD18-T vector kit（The precious biology in Dalian）, reaction system and operating process are：1.5 μ L PCR products are taken, are sequentially added 1 μL pMD18-T vector（50ng/μL）With 2.5 μ L 2 × Ligation solution I, mixing is placed on 16 DEG C of mistakes Night reacts；Connection product is transferred in bacillus coli DH 5 alpha using heat-shock transformed method；Using containing ampicillin （Ampicillin, Amp）LB solid medium screening positive clones, extract plasmid after bacterium colony PCR detection is correct.
Using a small amount of extraction agent boxes of SanPrep pillar Plasmid DNA（Give birth to work in Shanghai）Plasmid is extracted, takes 1 μ L for fine jade Sepharose electrophoresis is to detect the integrality and concentration level of extracted plasmid；Use restriction enzyme（TaKaRa）To plasmid pMD-18T-GmMATE75Double digestion is carried out with pENTR-2B（100 μ L systems）, reaction system and operating process are：Take 20 μ L pMD-18T-GmMATE75With pENTR-2B plasmids, sequentially add 10 μ L enzyme cutting buffering liquids, each 5 μ L of corresponding restriction endonuclease, 60 μ L ddH2O centrifuges after mixing, is placed in 37 DEG C of reaction overnights in short-term；All digestion products points are subjected to electrophoresis in Ago-Gel, Then rightGmMATE75Segment and pENTR-2B carrier segments carry out glue recycling respectively, and whole process uses SanPrep pillars DNA Plastic recovery kit（Give birth to work in Shanghai）；Take 1 μ L recovery products by agarose gel electrophoresis detect recycling segment size and Concentration is placed in -20 DEG C and saves backup.
Utilize T4 DNA Ligase（TaKaRa）, by recyclingGmMATE75Segment is connected with pENTR-2B carrier segments Get up, reaction system（20 μL）It is with operating process：Take 10 μ LGmMATE75 DNA fragmentation sequentially adds 2 μ L pENTR- 2B carrier DNAs, 2 μ L 10 × T4 DNA Ligase Buffer, 1 μ L T4 DNA Ligase, 5 μ L ddH2O is short after mixing When centrifuge, then 16 DEG C of water-bath reaction overnights；Then connection product is transferred in bacillus coli DH 5 alpha using heat-shock transformed method, is used Contain 50mg/L kanamycins（Kanamycin, Km）Solid medium screening positive clone；Picking individual colonies shake bacterium, to PCR is detected, and selection detects correct sample and is sequenced.
Correct bacterium solution is sequenced and carries out plasmid extraction, carrying out LR with over-express vector pK2GW7 reacts, and is weighed after reaction The plasmid of group carrier pK2-35S-GmMATE75, Transformed EcoliDH5 α, with the solid culture containing 100mg/L spectinomycins Base culture, screening positive clone simultaneously carry out bacterium colony PCR detections；It extracts plasmid and converts Agrobacterium pMP90 competent cells.
The recipient plant that this experiment uses is wild type（WT）Tobacco（Nicotiana L）, acquisition methods are by tobacco seed Son impregnates 30s with 75% alcohol, with the HgCl with 0.1% after sterile water washing28min is impregnated, if then again with sterile water washing It dry time, is seeded on MS culture mediums, 28 DEG C of 6 d of light culture go to illumination box after germination（25 DEG C, 16h/d illumination）, with Monthly use MS culture medium subcultures primary afterwards.WT tobacco leafs are taken, with agriculture bacillus mediated leaf disc transformation method transformation of tobacco blade.Turn Blade after change, which is placed on, to be co-cultured on solid medium MS1, is protected from light culture for 24 hours.Blade is moved into bud later and induces solid culture It is induced to sprout on base MS4, selection markers are kanamycins.After 1 month or so, young shoot is grown.Clip length is 1-2cm's Bud is moved on MS culture mediums and is cultivated, until it grows up to complete tobacco seedling.Blade is taken from cultured tobacco, is carried Total DNA is taken, and PCR identifications are carried out using it as template with the specific primer of GmMATE75, and identifies the tobacco of successful conversion Plant continues to cultivate the tobacco of successful conversion and carries out subsequent experimental.
Embodiment 2：GmMATE75 transgenic tobacco plant Aluminum toxicities are assessed
This experiment mainly assesses the Aluminum toxicity of tobacco in terms of 3:Opposite root elongation under aluminium processing, after aluminium processing Soluble sugar content in plant cell and hydrogen peroxide（H2O2）Content.Due under acid condition aluminium poison to the main of plant Harm is to inhibit the growth of its root, therefore degree of injury of the aluminium poison to plant can be indicated by measuring opposite root elongation.It is soluble The solute that sugar alleviates osmotic pressure as plant cell will produce soluble sugar to alleviate cellular damage when plant cell is damaged. Before reaching cell and bearing the upper limit, cellular damage degree can be considered positive correlation with the amount for generating soluble sugar.In plant tissue The H of accumulation2O2It is by some oxidizing ferment（Mainly superoxide dismutase SOD）The super positive anion generation oxidation reaction of catalysis and It is formed, content can reflect the extent of damage of membrane lipids of plant cell indirectly.
（1）Tobacco measures with respect to root elongation
Take the GmMATE75 transgenic tobaccos that young root length is about 1cm（MAC）, the solid medium that root is washed with water is placed in After cultivating 3d in tap water, it is put into the CaCl of 0.5mM pH4.5212h is pre-processed in solution；The root long of tobacco is measured later, so AlCl is added in culture solution afterwards3It is respectively 0 μM, 50 μM, 100 μM, 200 μM and 400 μM to be allowed to concentration；Exist respectively later For 24 hours, tobacco root depth is measured when 48h and 72h, as a contrast with WT types tobacco, every group of experiment carries out parallel laboratory test three times.
Root extends（cm）=this time measures the upper time measurement root long of root long-；
Opposite root elongation（%）=（Root extends/0 μM of aluminium chloride processing root elongation）100%；
By measurement result it is found that with the growth of aluminium processing time and the raising of concentration for the treatment of, the opposite root of tobacco extends Rate is on a declining curve；And no matter GmMATE75 transgenics tobacco with respect to root extends at any period under concentration for the treatment of Rate is obviously higher than corresponding WT types tobacco（Fig. 2）, it can thus be appreciated that GmMATE75 type tobaccos root is presented compared with WT type tobaccos Go out higher to Acid-Al stress resistance.
（2）Tobacco cell soluble sugar content measures
The GmMATE75 type tobaccos for taking 1 month or so after taking root after identifying are placed in water planting in clear water and after a week, 0.5 mM are added The CaCl of pH4.52Solution pre-processes 12 h, then is separately added into 0 μM, 50 μM, 100 μM, 200 μM and 400 μM of AlCl3Processing 24, it after 48 and 72h, takes root and leaf sample, every part of 1g of sample to add liquid nitrogen grinding, the 1 M Tris-HCl (pH of 1.5 mL is added 7.4) it, is transferred in EP pipes, 12000 rpm centrifuge 15 min, and transfer supernatant to new EP is managed, and soluble sugar and H are used for2O2Content It measures；Each sample takes 3 parts of measurement, as a contrast with WT types tobacco.
The content of soluble sugar is measured using anthrone colorimetry, with a μm olg-1FW is indicated.20 μ L supernatants are added Enter to a concentration of 1 mgmL of 1 mL-1Anthrone solution in, add 180 μ L ddH2After O, 10 min postcoolings of boiling water bath To room temperature；Soluble sugar content is calculated after substituting into standard curve y=0.485x+0.0118 after measurement light absorption value at 625 nm（Standard Curve is measured by glucose standards solution）；
By measurement result it is found that tobacco soluble sugar occurs mainly in blade cell after aluminium processing；Pass through comparison, WT type cigarettes Grass is higher than the blade and root cell of GmMATE75 type tobaccos by the polysaccharide generated in root after Acid-Al stress and leaf cell（Figure 3）, it may thus be appreciated that compared to WT type tobaccos, GmMATE75 types tobacco is influenced smaller by aluminium poison.
（3）Tobacco cell H2O2Assay
Use step（2）In obtained sample extracting solution measure H2O2Content；H2O2It is measured using xylenol orange method, with μ mol·g-1FW is indicated.Use ddH2O water reagent preparations A (3.3 mM FeSO4, 3.3 mM (NH4)2SO4, 412.5 mM H2SO4) and reagent B (165 μM of xylenol orange, 165 mM sorbierites).Using preceding reagent A and reagent B according to 1:10 ratio is mixed It closes and constitutes working reagent.This working reagent and H2O2Prepare liquid is according to 2:1 ratio mixes, and after 30 DEG C of 30 min of metal bath, measures OD560Value substitutes into standard curve y=0.26544x-0.0523 and calculates H2O2Changes of contents（With H2O2Titer makes standard curve）.
By result it is found that generating H in the root cell of tobacco after aluminium processing2O2It is more；It is thin by WT type tobaccos known to comparison H in born of the same parents2O2Content is apparently higher than GmMATE75 type tobaccos（Fig. 4）, it may thus be appreciated that compared to WT type tobaccos, GmMATE75 type cigarettes Grass cell membrane lipid damage under Acid-Al stress is smaller.
Sequence table
<110>Kunming University of Science and Technology
<120>A kind of pellet Bosnia-Herzegovena soybean citrate transporter protein gene and its application
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1288
<212> DNA
<213>Red Bosnia-Herzegovena soybean (Glycine max)
<400> 1
atggacgaga atagaagttc caacgaaccc aacaagtgga agatgcctct cttcgttttc 60
ttcaaaggtg caaggaatgt tttcaagctg gatgctctat ctcgggagat attagggatt 120
gcactcccct cagcactggc cgtttctgct gatccaattg cttctctcat agacacagca 180
ttcataggcc gtttaggacc ggtggaactt gcagctgctg gagtttccat ttctttgttt 240
aaccaagctt cgaggattac catattccct ctggtcaaca ttaccacttc ctttgtggct 300
gaagaagata ccattcaaaa actgaacacc aaagcagctg aaaatggtaa tagtaaggcc 360
aagttcggcg aaacaattat gccagaggat catatgcttc aagacatgga aaaaggtacc 420
cccaaagtga tgaatactga cgctccaaca gaatttaggg aagaaaaaga tgaatcaaag 480
gaatataatg ctaccggcaa caatgacaca aatattggag atggagccaa tacaatatgc 540
aagttttcat cggttactag cagtaagaag agtaaggaca aagttggaaa gaaaaagcga 600
cttattgctt cagcatcaac agcactactt tttggcacaa tccttggtct cattcaagct 660
gcagttctta tatttgcaac caaacctctg ttaggtgtta tgggtgtcaa acgagattct 720
cctatgctaa aacctgcaga gagctactta agattgagat cgttcggggc accagcagta 780
cttctctcct tggccatgca aggcatcttt cgagggttca aggatacaac aactccttta 840
tatgtcatcg tttcgggata tgcattgaat gtcatattgg acccaatttt tattttcaca 900
ttgaagttag gcatcaaagg tgcagccatt gcacatgttc tctcccagta catgatggca 960
ttcactctct tattgatatt aatgaaaaaa gtgcatctcc tacctccaag aataaaggat 1020
ttacagattt tccgatttct taaaaatgga ggattgttga tgctaaaggt aatagcagtc 1080
acattttgtg ttaccttagc aacatcattg gctgcaaggc taggttcaat tcccatggct 1140
gcatttcaaa catgcctaca ggtctggatg acatcgtccc ttcttgcaga tggtttggct 1200
gttgctgttc aggcaattct agcttgttct tttactgaga aagactataa gaaagcaaca 1260
gcagcagcaa caaggacact gcaaatga 1288
<210> 2
<211> 429
<212> PRT
<213>Red Bosnia-Herzegovena soybean (Glycine max)
<400> 2
Met Asp Glu Asn Arg Ser Ser Asn Glu Pro Asn Lys Trp Lys Met Pro
1 5 10 15
Leu Phe Val Phe Phe Lys Gly Ala Arg Asn Val Phe Lys Leu Asp Ala
20 25 30
Leu Ser Arg Glu Ile Leu Gly Ile Ala Leu Pro Ser Ala Leu Ala Val
35 40 45
Ser Ala Asp Pro Ile Ala Ser Leu Ile Asp Thr Ala Phe Ile Gly Arg
50 55 60
Leu Gly Pro Val Glu Leu Ala Ala Ala Gly Val Ser Ile Ser Leu Phe
65 70 75 80
Asn Gln Ala Ser Arg Ile Thr Ile Phe Pro Leu Val Asn Ile Thr Thr
85 90 95
Ser Phe Val Ala Glu Glu Asp Thr Ile Gln Lys Leu Asn Thr Lys Ala
100 105 110
Ala Glu Asn Gly Asn Ser Lys Ala Lys Phe Gly Glu Thr Ile Met Pro
115 120 125
Glu Asp His Met Leu Gln Asp Met Glu Lys Gly Thr Pro Lys Val Met
130 135 140
Asn Thr Asp Ala Pro Thr Glu Phe Arg Glu Glu Lys Asp Glu Ser Lys
145 150 155 160
Glu Tyr Asn Ala Thr Gly Asn Asn Asp Thr Asn Ile Gly Asp Gly Ala
165 170 175
Asn Thr Ile Cys Lys Phe Ser Ser Val Thr Ser Ser Lys Lys Ser Lys
180 185 190
Asp Lys Val Gly Lys Lys Lys Arg Leu Ile Ala Ser Ala Ser Thr Ala
195 200 205
Leu Leu Phe Gly Thr Ile Leu Gly Leu Ile Gln Ala Ala Val Leu Ile
210 215 220
Phe Ala Thr Lys Pro Leu Leu Gly Val Met Gly Val Lys Arg Asp Ser
225 230 235 240
Pro Met Leu Lys Pro Ala Glu Ser Tyr Leu Arg Leu Arg Ser Phe Gly
245 250 255
Ala Pro Ala Val Leu Leu Ser Leu Ala Met Gln Gly Ile Phe Arg Gly
260 265 270
Phe Lys Asp Thr Thr Thr Pro Leu Tyr Val Ile Val Ser Gly Tyr Ala
275 280 285
Leu Asn Val Ile Leu Asp Pro Ile Phe Ile Phe Thr Leu Lys Leu Gly
290 295 300
Ile Lys Gly Ala Ala Ile Ala His Val Leu Ser Gln Tyr Met Met Ala
305 310 315 320
Phe Thr Leu Leu Leu Ile Leu Met Lys Lys Val His Leu Leu Pro Pro
325 330 335
Arg Ile Lys Asp Leu Gln Ile Phe Arg Phe Leu Lys Asn Gly Gly Leu
340 345 350
Leu Met Leu Lys Val Ile Ala Val Thr Phe Cys Val Thr Leu Ala Thr
355 360 365
Ser Leu Ala Ala Arg Leu Gly Ser Ile Pro Met Ala Ala Phe Gln Thr
370 375 380
Cys Leu Gln Val Trp Met Thr Ser Ser Leu Leu Ala Asp Gly Leu Ala
385 390 395 400
Val Ala Val Gln Ala Ile Leu Ala Cys Ser Phe Thr Glu Lys Asp Tyr
405 410 415
Lys Lys Ala Thr Ala Ala Ala Thr Arg Thr Leu Gln Met
420 425
<210> 3
<211> 34
<212> DNA
<213>Artificial sequence (Artificial)
<400> 3
cgggatcccg atggacgaga atagaagttc caac 34
<210> 4
<211> 36
<212> DNA
<213>Artificial sequence (Artificial)
<400> 4
ccgctcgagc ggtcatttgc agtgtccttg ttgctg 36

Claims (2)

1. a kind of pellet Bosnia-Herzegovena soybean citrate transporter protein geneGmMATE75, it is characterised in that：Its nucleotide sequence such as SEQ ID NO:Shown in 1, coding such as SEQ ID NO:Amino acid sequence shown in 2.
2. application of the pellet Bosnia-Herzegovena soybean citrate transporter protein gene described in claim 1 in improving Aluminum Tolerance in Plants ability.
CN201810075636.0A 2018-01-26 2018-01-26 Danbo black soybean citric acid transport protein gene and application thereof Active CN108410880B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201810075636.0A CN108410880B (en) 2018-01-26 2018-01-26 Danbo black soybean citric acid transport protein gene and application thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201810075636.0A CN108410880B (en) 2018-01-26 2018-01-26 Danbo black soybean citric acid transport protein gene and application thereof

Publications (2)

Publication Number Publication Date
CN108410880A true CN108410880A (en) 2018-08-17
CN108410880B CN108410880B (en) 2021-10-22

Family

ID=63126243

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201810075636.0A Active CN108410880B (en) 2018-01-26 2018-01-26 Danbo black soybean citric acid transport protein gene and application thereof

Country Status (1)

Country Link
CN (1) CN108410880B (en)

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111826383A (en) * 2020-07-16 2020-10-27 昆明理工大学 Application of Danbo black soybean superoxide dismutase gene in improving plant aluminum tolerance
CN114045294A (en) * 2021-11-22 2022-02-15 昆明理工大学 Lipid transport protein gene and application thereof
CN116694675A (en) * 2023-06-19 2023-09-05 东北农业大学 Application of soybean GmGST gene in improving aluminum toxicity stress resistance of plants

Citations (14)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008038705A1 (en) * 2006-09-29 2008-04-03 Kobayashi Pharmaceutical Co., Ltd. Glycation inhibitor
CN102154318A (en) * 2011-01-30 2011-08-17 浙江大学 Snailflower citric acid transporter gene VuMATE and use thereof
CN102952822A (en) * 2012-11-06 2013-03-06 昆明理工大学 Plant expression vector of Tanba black soybean C2H2 zinc finger protein gene STOP1 and application thereof
EP2155219B1 (en) * 2007-05-17 2014-02-19 The United States of America, as represented by the Secretary of Agriculture The sorghum aluminum tolerance gene, sbmate
KR101506938B1 (en) * 2014-11-10 2015-03-30 문석우 Producing method of pork cultet type soybean processed foods using dried fruits
CN104542033A (en) * 2014-12-18 2015-04-29 昆明理工大学 Application of vanadate to improvement of soil cultivated plant drought resistant stress
CN104561026A (en) * 2013-10-29 2015-04-29 中国农业大学 Application of peanut AhFRDL1 gene in improving aluminum toxicity stress resistance of plants
CN104871847A (en) * 2015-05-14 2015-09-02 昆明理工大学 Application of ascorbic acid in plant nitrate nitrogen adsorption under increased aluminum stress
CN105861459A (en) * 2016-05-17 2016-08-17 昆明理工大学 Application of AtPrx64 gene in improving aluminum tolerance of plants
CN106047919A (en) * 2016-06-02 2016-10-26 昆明理工大学 Application of AtPrx64 in improving plant nitrate nitrogen absorption under aluminum stress
WO2017042099A1 (en) * 2015-09-09 2017-03-16 Basilea Pharmaceutica Ag Efflux-pump inhibitors and therapeutic uses thereof
WO2017093157A1 (en) * 2015-11-30 2017-06-08 Basilea Pharmaceutica Ag Piperidine, pyrrolidine and 2-oxo-1,3-oxazinane derivatives as inhibitors of bacterial efflux-pumps for the treatment of microbial infections
CN107488668A (en) * 2017-08-28 2017-12-19 广西大学 A kind of sorghum SbNrat1 genes and its application
US10561148B2 (en) * 2015-05-06 2020-02-18 Snipr Technologies Limited Altering microbial populations and modifying microbiota

Patent Citations (15)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008038705A1 (en) * 2006-09-29 2008-04-03 Kobayashi Pharmaceutical Co., Ltd. Glycation inhibitor
EP2155219B1 (en) * 2007-05-17 2014-02-19 The United States of America, as represented by the Secretary of Agriculture The sorghum aluminum tolerance gene, sbmate
CN102154318A (en) * 2011-01-30 2011-08-17 浙江大学 Snailflower citric acid transporter gene VuMATE and use thereof
CN102952822A (en) * 2012-11-06 2013-03-06 昆明理工大学 Plant expression vector of Tanba black soybean C2H2 zinc finger protein gene STOP1 and application thereof
CN104561026A (en) * 2013-10-29 2015-04-29 中国农业大学 Application of peanut AhFRDL1 gene in improving aluminum toxicity stress resistance of plants
KR101506938B1 (en) * 2014-11-10 2015-03-30 문석우 Producing method of pork cultet type soybean processed foods using dried fruits
CN104542033A (en) * 2014-12-18 2015-04-29 昆明理工大学 Application of vanadate to improvement of soil cultivated plant drought resistant stress
US10561148B2 (en) * 2015-05-06 2020-02-18 Snipr Technologies Limited Altering microbial populations and modifying microbiota
US10624349B2 (en) * 2015-05-06 2020-04-21 Snipr Technologies Limited Altering microbial populations and modifying microbiota
CN104871847A (en) * 2015-05-14 2015-09-02 昆明理工大学 Application of ascorbic acid in plant nitrate nitrogen adsorption under increased aluminum stress
WO2017042099A1 (en) * 2015-09-09 2017-03-16 Basilea Pharmaceutica Ag Efflux-pump inhibitors and therapeutic uses thereof
WO2017093157A1 (en) * 2015-11-30 2017-06-08 Basilea Pharmaceutica Ag Piperidine, pyrrolidine and 2-oxo-1,3-oxazinane derivatives as inhibitors of bacterial efflux-pumps for the treatment of microbial infections
CN105861459A (en) * 2016-05-17 2016-08-17 昆明理工大学 Application of AtPrx64 gene in improving aluminum tolerance of plants
CN106047919A (en) * 2016-06-02 2016-10-26 昆明理工大学 Application of AtPrx64 in improving plant nitrate nitrogen absorption under aluminum stress
CN107488668A (en) * 2017-08-28 2017-12-19 广西大学 A kind of sorghum SbNrat1 genes and its application

Non-Patent Citations (7)

* Cited by examiner, † Cited by third party
Title
JUGE LIU等: ""Genome-wide analysis of MATE transporters and expression patterns of a subgroup of MATE genes in response to aluminum toxicity in soybean"", 《BMC GENOMICS》 *
LIU,L.等: ""Glycine max multidrug and toxic compound extrusion (MATE) mRNA, complete cds"", 《GENBANK DATABASE》 *
NCBI: ""MATE family protein [Glycine max]"", 《GENBANK DATABASE》 *
YING ZHOU等: ""Functional characterization of three MATE genes in relation to aluminum-induced citrate efflux from soybean root"", 《PLANT SOIL》 *
侯高杰等: ""铝毒胁迫下大豆ALMT和MATE基因的特异性表达"", 《安徽农业科学》 *
柳聚阁等: ""大豆多药及毒性化合物外排转运（MATE）家族的生物信息学分析及耐铝相关MATE 基因的表达分析"", 《江苏省遗传学会2016年学术年会论文摘要集》 *
郝竞一: ""丹波黑大豆bHLH30 转录因子调控GmSTOP1和GmMATE75基因抗铝胁迫研究"", 《中国优秀硕士学位论文全文数据库基础科学辑》 *

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111826383A (en) * 2020-07-16 2020-10-27 昆明理工大学 Application of Danbo black soybean superoxide dismutase gene in improving plant aluminum tolerance
CN111826383B (en) * 2020-07-16 2022-08-02 昆明理工大学 Application of Danbo black soybean superoxide dismutase gene in improving plant aluminum tolerance
CN114045294A (en) * 2021-11-22 2022-02-15 昆明理工大学 Lipid transport protein gene and application thereof
CN114045294B (en) * 2021-11-22 2023-03-24 昆明理工大学 Lipid transport protein gene and application thereof
CN116694675A (en) * 2023-06-19 2023-09-05 东北农业大学 Application of soybean GmGST gene in improving aluminum toxicity stress resistance of plants
CN116694675B (en) * 2023-06-19 2024-05-03 东北农业大学 Application of soybean GmGST gene in improving aluminum toxicity stress resistance of plants

Also Published As

Publication number Publication date
CN108410880B (en) 2021-10-22

Similar Documents

Publication Publication Date Title
CN107058328A (en) A kind of method for improving plant amylose content and application
CN108410880A (en) A kind of pellet Bosnia-Herzegovena soybean citrate transporter protein gene and its application
CN103194457B (en) Lilium regale germin-like protein gene LrGLP2 and application thereof
CN104878019B (en) Yangbi bulla walnut class sprouts fibroin gene JsGLP1 and application
CN113646326A (en) Gene for resisting plant diseases
CN106636134A (en) Maize zinc-iron regulation transporter ZmZIP2 gene and application thereof
CN104558128A (en) Protein relating to resistance to fusarium graminearum schw stem rot and encoding gene and application thereof
CN113621625B (en) Application of sesame SiERF103 gene in enhancing plant resistance
CN109355295B (en) Peanut AhWRKY75 gene and application thereof in improving peanut salt tolerance
CN113444734B (en) Preparation method and application of salt-tolerant transgenic poplar
CN106834315A (en) One kind is than white thorn NbCIPK25 genes and its expressing protein and the application of undercuting
CN110229818A (en) Wax plum CpSNAC1 gene promoter and its application
CN113754748A (en) Polypeptide immune activator for improving insect resistance and disease resistance of rice
CN110819634B (en) Clone of lilium tenuifolium gene LpNAC6 and application thereof
CN103266118A (en) Cabbage type rape anti-sulfonylurea herbicide gene and application thereof
CN104404043A (en) Promoter of gene Me094 related to bacterial-blight resistance of Oryza meyeriana
CN108864264B (en) Corn OXS2a gene, and encoding protein and application thereof
CN112794887B (en) Duli pear transcription factor PbrWRKY40 and application thereof in improving total acid content of plants and salt resistance genetic improvement
CN111154770B (en) Application of rice gene OsABCC2 in regulation of absorption and transportation of pesticides
CN101712718A (en) Protein relevant to plant drought resistance, coding gene and application thereof
CN100432101C (en) Tomato RNA virus host factor and its coding gene and use thereof
CN114032245A (en) Gene VLNHX3D in regulating plant cell Na+And/or K+Application in concentration
CN114717245B (en) MsbHLH35 gene and application of encoding protein thereof in regulation and control of alfalfa yield and stain resistance
CN109053870A (en) Application of AtERF49 gene during plant responding high temperature stress
CN112592392B (en) Application of pleiotropic gene SbSnf4 in improvement of cane sugar yield, plant height, fresh weight of stems and/or juice amount

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant