CN101984059A - Gossypium barbadense DREB transcription factor gene and application thereof - Google Patents
Gossypium barbadense DREB transcription factor gene and application thereof Download PDFInfo
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Abstract
The invention relates to the field of plant genetic engineering, and provides a gossypium barbadense DREB transcription factor gene which has the nucleotide sequence shown as the SEQ ID NO:1. The gossypium barbadense DREB transcription factor gene is constructed by the steps of constructing a pC-XHDREB plant expression vector, transforming the tobacco by the agrobactrium tumefaciens to obtain the XHDREB gene transformed tobacco, and verifying the XHDREB gene transformed tobacco by the drought simulation experiment. The tobacco has the capacities of drought resistance, saline resistance and cold resistance due to the over-expression of the XHDREB.
Description
Technical field
The present invention relates to plant genetic engineering field, relate in particular to a kind of sea island cotton DREB transcription factor gene with and the plant expression vector that makes up and in the application aspect the drought-enduring transgenic plant development.
Background technology
Transcription factor (transcription factor, TF) be called trans-acting factor again, be meant can with the cis-acting elements generation specificity bonded albumen in the eukaryotic gene promoter region, by between them and and other associated protein between interaction activate or suppress some gene transcription.Difference according to combination can be divided into two classes to transcription factor: a class is the ubiquity transcription factor, another kind of is idiosyncratic transcription factor, the ubiquity transcription factor can with the core sequence TATA frame combination of promotor, can activate all gene transcription, and idiosyncratic transcription factor combines with other regulatory element on the dna sequence dna, can only activate special genes.Since Paz-Ares reported first in 1987 behind the corn transcription factor gene, so far from higher plant isolated regulation and control arid, low temperature, high salt, hormone, grow and the transcription factor of related gene expression such as cause of disease reaction has reached hundreds of.The gene of discovering the encoding transcription factor in Arabidopis thaliana has 1533 at least, accounts for 5.9% of its gene number, and these transcription factors are grown and played an important role aspect the response of environment to external world in coordinate plant growth.
DREB (dehydration responsive element binding protein) transcription factor is to be present in the plant, combine with DRE (dehydration responsive element) cis-acting elements, the protein of related gene expression is replied in regulation and control to adverse circumstances such as arid, high salt, low temperature.
The discovery of DREB class transcription factor is one of breakthrough progress of the tool in plant stress-resistance Journal of Sex Research aspect in recent years.It plays important regulation in plant is replied the process of arid, high salt and low temperature stress signal.
Since 1997, Thotnashow research group and Liu Qiang in 1998 etc. take the lead in cloneing and isolateing CBF1 (DREB1B) from Arabidopis thaliana, CBF2, (DREB1C) and since 3 CBF/DREB1 classes of CBF3 (DREB1A) transcription factor, the research of this class transcription factor in the plant has been subjected to paying close attention to widely.The expression of most of transcription factor genes is subjected to external stimulus and the inducing of process of growing, and the dreb gene mediation is complicated coerces the signal delivery network, participates in multiple different signal transmission.The expression of most of dreb gene all is subjected to inducing of environment stresses such as arid, high salt, low temperature, but different types of DREB transcription factor has different expression mechanisms, even same kind, also have different expression mechanisms, plant to the reaction of environment stress and hormone response in the different regulating and controlling effect of performance.Have not yet to see relevant report about sea island cotton DREB transcription factor gene.
Summary of the invention
The object of the present invention is to provide a kind of sea island cotton DREB transcription factor gene, its nucleotide sequence is shown in SEQ ID NO:1.
Second purpose of the present invention is to provide a kind of sea island cotton DREB transcription factor, and by nucleotide sequence coded shown in the SEQ ID NO:1, its aminoacid sequence is shown in SEQ ID NO:2.
The 3rd purpose of the present invention is to provide the plant expression vector that contains described sea island cotton DREB transcription factor gene.
The 4th purpose of the present invention is to provide with described plant expression vector plant transformed cell, tissue or plant.
The 5th purpose of the present invention is that described sea island cotton DREB transcription factor gene is in the application of cultivating in adversity resistant plant, especially the drought-enduring plant kind.
Technological line of the present invention is:
1) gets new extra large 16 blades of sea island cotton;
2) extract genomic dna and total RNA from blade;
3) carrying out reverse transcription reaction with total RNA, obtain cDNA, is template pcr amplification DREB transcription factor gene with cDNA, called after: XHDREB;
4) make up the pC-XHDREB plant expression vector, use the agrobacterium tumefaciens transformation of tobacco, obtaining changes the XHDREB genetic tobacco, adopts arid simulated experiment checking to change the XHDREB genetic tobacco, because of the expression of crossing of XHDREB has drought-enduring, salt tolerant, anti-cold energy power.
The present invention has cloned a kind of sea island cotton DREB transcription factor gene XHDREB, make up the pC-XHDREB plant expression vector, use the agrobacterium tumefaciens transformation of tobacco, obtain to change the XHDREB genetic tobacco, adopt arid simulated experiment checking to change the XHDREB genetic tobacco, express because of crossing of XHDREB have drought-enduring, salt tolerant, anti-cold energy power.
Description of drawings
Fig. 1 sea island cotton dreb gene pcr amplification electrophorogram, the amplified production of the amplified production of swimming lane: cDNA, swimming lane 2:DNA, swimming lane M:DNA standard molecular weight DL2000;
Fig. 2 recombinant plasmid pC-XHDREB makes up schema
The PCR of Fig. 3 transgene tobacco detects electrophorogram, swimming lane 1: positive control, swimming lane 2-9: transfer-gen plant, swimming lane 10: negative control, swimming lane M:DNA standard molecular weight DL2000;
The drought tolerance analytical results figure of Fig. 4 A-4D transgene tobacco, Fig. 4 A: arid is handled commentaries on classics XHDREB genetic tobacco, Fig. 4 B of 3d: arid is handled non-transgenic tobacco, Fig. 4 C of 3d: rehydration is recovered commentaries on classics XHDREB genetic tobacco, Fig. 4 D of growth 3d: rehydration is recovered the non-transgenic tobacco of growth 3d;
The salt tolerance analytical results figure of Fig. 5 A-5D transgene tobacco, Fig. 5 A:200mmol/L NaCl handle the commentaries on classics XHDREB genetic tobacco of 3d, non-transgenic tobacco, Fig. 5 C that Fig. 5 B:200mmol/LNaCl handles 3d continuously continuously: commentaries on classics XHDREB genetic tobacco, Fig. 5 D behind the recovery growth 3d: the non-transgenic tobacco behind the recovery growth 3d;
The lower temperature resistance analytical results figure of Fig. 6 A-6D transgene tobacco, non-transgenic tobacco, Fig. 6 C of the commentaries on classics XHDREB genetic tobacco of Fig. 6 A:4 ℃ of processing 24h, Fig. 6 B:4 ℃ processing 24h: commentaries on classics XHDREB genetic tobacco, 6D under the room temperature behind the recovery cultivation 3d: the non-transgenic tobacco under the room temperature behind the recovery cultivation 3d.
Embodiment
Below in conjunction with specific embodiment technological line of the present invention is described in further details.
Toolenzyme and reagent
(1) restriction enzyme, modifying enzyme and test kit: the T4 dna ligase, the Taq enzyme, ThermoScript II AMV, DNA reclaim purification kit all available from sky, Beijing root biotech company.Plasmid extracts test kit in a small amount available from vast Imtech, and other chemical reagent is available from Beijing ancient cooking vessel state company.
(2) other medicines: agarose, Tris, diethylpyrocarbonate (DEPC) are Beijing ancient cooking vessel state company product.Peptone, yeast extract, chloroform, primary isoamyl alcohol, ethanol, Virahol, NaCl, CaCl2 etc. are homemade analytical reagent.5-bromo-4-chloro-3-indoles galactoside (X-gal), isopropylthio-β-D galactoside (IPTG), penbritins etc. are available from Beijing ancient cooking vessel state company.Dna molecular amount standard Mark Ladder 2000 is available from sky, Beijing root biotech company.
The configuration of solution
(1) plant genome DNA extracts damping fluid 500mL:0.35mol/L glucose; 0.1mol/L Tris-HCl (pH 8.0); 5mmol/L Na-EDTA (pH 8.0); 2% PVP; 1% β-Me
(2) plant genome DNA extracts lysate 500mL:1.4mmol/L NaCl; 0.1mol/L Tris-HCl (pH 8.0); 20mmol/L Na-EDTA (pH 8.0); 2% CTAB; 2% PVP; 1% β-Me
(3) plant genome DNA lysate TE (pH 8.0) 1L:10mmol/L Tris-HCl; 1mmol/LEDTA (pH 8.0)
(4) plant genome DNA electrophoretic buffer 50 * TBE (500mL): Tris 121.0g; Glacial acetic acid 28.5mL; 0.5mol/L EDTA 50.0mL (pH 8.0); 1mol/L Tris-HCl (pH 8.0) 500.0mL; Tris 60.0g; Water 400.0mL; Dense HCl 21.0mL
Substratum
(1) LB liquid nutrient medium: Tryptones 10.0g, yeast extract 5.0g, NaCl 10.0, and pH 7.0 constant volumes are to 1000mL.
(2) LB solid medium: Tryptones 10.0g, yeast extract 5.0g, NaCl 10.0g, agar powder 15.0g, pH 7.0 constant volumes are to 1000mL.
(3) LB selects substratum: before LB paves plate, treat that the substratum temperature drops to about 50 ℃, adding microbiotic to concentration is 100mg/L, paves plate after shaking up.
Key instrument
Pcr amplification instrument (Tachne T312), high speed freezing centrifuge (Hettich MIKRO 200R), electrophoresis equipment (Bio RAD), gel imaging system (Bio RAD).
The clone of embodiment 1 sea island cotton XHDREB gene
1, the cultivation of vegetable material
The new sea 16 of sea island cotton (Gossypium barbadense L.), the seed distilled water immersion of sterilization, in 37 ℃ of insulation cans, place 48h, clean up with the distilled water of sterilizing again after showing money or valuables one carries unintentionally, squeeze out kind of a skin and be placed in the 1/2 MS substratum, under 25 ℃, intensity of illumination 2500 Lux and 12h photoperiod condition, cultivate.After one week, win blade from aseptic seedling and be stored in liquid nitrogen to extract genomic dna, RNA.
2, the extraction of sea island cotton genomic dna
Extracting method is with reference to the CTAB method, and concrete steps are as follows:
Get blade 3～5g, after in liquid nitrogen, grinding, the 1.5ml centrifuge tube of packing into; In the 1.5ml centrifuge tube, add the DNA extraction damping fluid (65 ℃ of preheatings) of 800 μ L, and add 50 μ L β-thin basic ethanol, 65 ℃ of water-bath 20min behind the mixing, middle mixing is several times; Add 300 μ L 5mmol/L Potassium ethanoate (pH 7.0) mixings, behind ice bath 20～60min, at room temperature thaw; The DNA extraction lysate that adds 300 μ L, mixing, 65 ℃ of water-bath 20min, middle mixing several times, room temperature, the centrifugal 10min of 10000rpm; Draw supernatant and place a new 1.5ml centrifuge tube, add 600 μ L chloroforms: primary isoamyl alcohol (24: 1) mixing, the centrifugal 10min of 10000rpm; Get supernatant and place a new 1.5ml centrifuge tube, add the Virahol of 4 ℃ of precoolings of equal-volume.Mixing gently is more than-20 ℃ of ice bath 30min; The centrifugal 5min of 6000rpm abandons supernatant, precipitates 2 times with 70% washing with alcohol, and dehydrated alcohol is washed 1 time, dries up; Add 50 μ L TE dissolving DNAs.
3, the extraction of the total RNA of sea island cotton
Hot phenol method with improvement extracts sea island cotton (new sea 16) the total RNA of blade, and concrete grammar is as follows:
4mL is extracted damping fluid (with preceding adding PVP and β-thin basic ethanol) add in the centrifuge tube, behind the mixing, place 65 ℃ of water-bath water-bath 20min with the 4mL phenol-chloroform; Get sea island cotton blade 1～2g, change over to after in liquid nitrogen, grinding and be equipped with in the centrifuge tube that extracts damping fluid and phenol-chloroform mixture, vortex vibration 30s; 4 ℃, behind the 12000rpm high speed centrifugation 5min, supernatant is moved to adding 4mL phenol-chloroform mixed solution in another centrifuge tube, mixing, centrifugal, repeat this operation one to twice, get supernatant then; The LiCl that adds isopyknic 4mol/L, mixing is placed more than the 2h the centrifugal 20min of 12000rpm for-20 ℃; Abandon supernatant, with 70% washing with alcohol precipitation, then with resolution of precipitate in the TE of 0.5mL, mixed solution is changed in the centrifuge tube of 1.5mL; Add the extracting one of isopyknic phenol-chloroform mixed solution to twice, supernatant is changed in another centrifuge tube, the dehydrated alcohol that adds 1/10 volume 3mol/L sodium-acetate (pH 5.3) and 2～3 times of volumes, mixing, place 1～2h, 14000rpm, 4 ℃ of centrifugal 5min for-20 ℃, with 70% washing with alcohol precipitation, room temperature is placed 2～3min; The DEPC water that adds 200～500 μ L, dissolution precipitation ,-80 ℃ of preservations are standby.
The total RNA of sea island cotton blade that extracts detects its integrity by agarose gel electrophoresis, and the uv-spectrophotometric instrument detects can see that two obvious RNA bands show that then sample can use.
4, the acquisition and the clone of sea island cotton XHDREB transcription factor gene total length
The first chain cDNA's is synthetic
Undertaken by Promega company reverse transcription test kit specification sheets.Be divided into pre-sex change and reverse transcription two portions, pre-reaction of degeneration system sees Table 1, and reaction conditions is 70 ℃ of incubation 5min, puts 10min on ice immediately.The reverse transcription reaction system sees Table 2, and reaction conditions is 25 ℃, 5min, 42 ℃, 1h, and 70 ℃, 15min deactivation ThermoScript II ,-20 ℃ of refrigerators are preserved reverse transcription product.
The pre-reaction of degeneration system of table 1
Each composition of table 2 reverse transcription reaction
Sea island cotton XHDREB gene PCR clone
Be template with sea island cotton (new sea 16) cDNA and genomic dna respectively, P1 and P2 are primer, carry out the PCR reaction, and reaction system sees Table 3, and amplification obtains cDNA and genomic dna total length and cloning and sequencing.And be XHDREB with the unnamed gene that obtains.
Table 3 PCR reaction system
Reaction conditions: 94 ℃, 3min; 94 ℃, 1min, 50 ℃, 45s, 72 ℃, 1min (35 circulations); 72 ℃, 10min, 4 ℃ of coolings ,-20 ℃ of preservations.
The amplimer sequence is as follows:
P1：5′-ATGGATTTTTTAGTTCAAGATTA-3′
P2：5′-TTAAATAGAATAACTCCATAAAGGT-3′
Be template with its cDNA and genomic dna respectively, carry out pcr amplification, obtain the specific band of identical size,, show that sea island cotton XHDREB gene inside does not have intron referring to Fig. 1 with Auele Specific Primer P1, P2.
The XHDREB gene nucleotide series is shown in SEQ ID NO:1.
The proteic aminoacid sequence of XHDREB genes encoding is shown in SEQ ID NO:2.
The structure of embodiment 2 pC-DREB plant expression vectors
See also Fig. 2, with KI and Pst I restriction endonuclease XHDREB gene fragment and expression vector PCAMBIA2300-35SOCS being carried out enzyme cuts, the XHDREB gene that downcuts is inserted between the CaMV35S promotor and OCS terminator of pCAMBIA2300-35SOCS expression vector, has made up plant expression vector pC-DREB.Plant expression vector pCAMBIA2300-35SOCS is a fundamental construction with pCAMBIA2300, has added 35S promoter and multienzyme is cut the site.
Double digestion sea island cotton dreb gene fragment
Primer DR-1, DR-2 with design band KI and Pst I restriction enzyme site carry out pcr amplification to sea island cotton cDNA library, reclaim the XHDREB gene fragment that amplified production obtains having KI and Pst I restriction enzyme site.Use KI then, Pst I carries out double digestion to the gene fragment that reclaims, and the enzyme system of cutting sees Table 4.
Primer sequence is as follows:
Upstream primer DR-1:5 '-
GGGGTACCATGGATTTTTTAGTTCAAGATTA-3 ';
Downstream primer DR-2:5 '-
AACTGCAGTTAAATAGAATAACTCCATAAAGGT-3 ';
The KI of line part for adding, Pst I restriction enzyme site, it is synthetic that primer is given birth to the worker by Shanghai.
Table 4 XHDREB gene enzyme is cut reaction system
Agarose gel electrophoresis with 1% detects, then with the XHDREB gene fragment behind sky, Beijing DNA of the root biotech company recovery purification kit recovery double digestion.
Embodiment 3 utilizes agriculture bacillus mediated conversion method to obtain to change the XHDREB genetic tobacco
1, the preparation of Agrobacterium competent cell
The single colony inoculation of picking EHA105 in 5mLYEB liquid nutrient medium (containing Streptomycin sulphate 100mg/mL, Rifampin 100mg/mL), 28 ℃, the 250rpm overnight incubation; Draw the 2mL culture and change in the 50mLYEB liquid nutrient medium, continuing to be cultured to the OD600 value is 0.6; Bacterium liquid is gone in the aseptic centrifuge tube ice bath 30min, the centrifugal 5min of 5000rpm; With the resuspended thalline of the aseptic CaCl2 of 1.7mL20mmol/L, add the aseptic glycerine of 300 μ L, behind the mixing, be sub-packed in the aseptic 1.5mL Eppendorf tube by every pipe 200 μ L ,-70 ℃ of maintenances are standby.
2, freeze-thaw method changes recombinant plasmid PC-XHDREB over to Agrobacterium
The EHA105 competent cell and the positive recombinant plasmid of 10 μ L that in the 1.5mL centrifuge tube of sterilization, add 200 μ L prepared fresh, ice bath 1h behind the mixing; Centrifuge tube is moved in 42 ℃ of water-baths water-bath 60s (must guard against vibration); Immediately centrifuge tube is placed ice, move into water-bath 5min in 37 ℃ of water-baths behind 2～3min again; In centrifuge tube, add 37 ℃ of 800 μ L YEB liquid nutrient medium, on 37 ℃ of constant temperature oscillators, cultivate 1h with 260r/min; The centrifugal 5min of 4000rpm abandons supernatant, and the residue thalline is used to be coated with the YEB agar plate; Getting the residue thalline evenly coats YEB and selects substratum (to contain Streptomycin sulphate 100mg/mL, kantlex 100mg/mL, Rifampin 100mg/mL) surface, after treating that thalline is absorbed fully, flat board is inverted in 28 ℃ of incubators cultivates 48h, after treating to grow single bacterium colony on the substratum, carry out bacterium colony PCR, the screening positive recombinant, PCR application of sample system and reaction system are with reference to the 2nd chapter; The positive bacterium colony that picking filters out in the YEB liquid nutrient medium, is cultivated 48h for 28 ℃, then bacterium liquid is prepared into glycerol stock, and-20 ℃ of preservations are used for subsequent experimental.
3, genetic transformation of tobacco and plant regeneration
Adopt the agrobacterium-mediated transformation transformation of tobacco, obtain to change the XHDREB genetic tobacco.
1) get the Agrobacterium bacterium liquid 100 μ L that preserve in-20 ℃ of refrigerators and be inoculated in the 3mLYEB liquid nutrient medium, 28 ℃, the 180rpm incubated overnight.Get 50% glycerine, the 200 μ L that activation bacterium liquid 200 μ L add sterilization and be stored in the centrifuge tube, be put in-20 ℃ of refrigerators standby.Get in the YEB liquid nutrient medium of activation bacterium liquid or above-mentioned standby bacterium liquid 100 μ L adding 10mL, 28 ℃, 180rpm shaking culture to OD600 value is 0.6～0.8.Bacterium liquid branch is installed in the 50mL triangular flask, be used to infect;
2) get tobacco aseptic seedling blade and cut leaf disc, cultivate 2～3d in advance, will soak 10～15min through the pre-Agrobacterium bacterium liquid of cultivating after the leaf dish of handling is put into activation;
3) after the leaf panel surface bacterium liquid that will infect with the thieving paper of sterilizing blots, the leaf dish is put on the division culture medium, 25 ℃, secretly cultivates 48h;
4) dark cultivate finish after, the leaf dish is put in selects to cultivate on the division culture medium, later per 10～15d subculture once;
5) after selection cultivated for 2～3 weeks, differentiate resistant buds around the leaf dish, treat that resistant buds grows to 2～3cm, downcut to be inserted into and select to carry out root culture in the root media, obtain changeing the XHDREB genetic tobacco.
4, the evaluation of transgenic tobacco plant and detection
The DNA detection of transgene tobacco
Extract the total DNA of transgene tobacco blade, with the negative contrast of the total DNA of non-transgenic tobacco, carry out pcr amplification reaction, amplification system and reaction system are referring to embodiment 1.
Get amplified production respectively and carry out electrophoresis detection on 1% sepharose, can obtain the band of about 650bp, illustrate that the XHDREB gene has been integrated into the tobacco gene group, electrophoresis result is referring to Fig. 3.
5, express the degeneration-resistant simulated experiment and the Function Identification of XHDREB transgene tobacco excessively
1) the drought resistance analysis of transgene tobacco
Compare with the non-transgenic tobacco, will change XHDREB gene masculine tobacco and at room temperature carry out natural drought 3d with contrast, rehydration 3d observes its profile then.
Shown in Fig. 4 A and 4B, after arid is handled 3d, the wilting phenomenon has all appearred in transgenosis and non-transgenic tobacco, the blade edge of transgene tobacco begins to curl but the wilting degree of blade is not very serious, but not transgene tobacco is followed down several second leaf except lobus cardiacus and also kept certain turgescence rest blade all to wilt.Shown in Fig. 4 C and 4D, after rehydration is recovered growth 3d, the recovery extent of transgene tobacco blade is significantly better than wild-type as can be seen, the blade of transgene tobacco has all recovered turgescence, but not transgene tobacco has only lobus cardiacus and second and third layer blade to recover turgescence, and rest blade all is in wilted condition.This shows that the overexpression of XHDREB gene in tobacco can improve the resistance of transgenosis grass planting to drought stress really.
2) the salt tolerance analysis of transgene tobacco
Compare with not genetically modified tobacco seedling, will change XHDREB gene masculine tobacco and handle 3d continuously with 200mmol/LNaCl, recover under normal operation then to grow behind the 3d, observe its profile with contrast.
Shown in Fig. 5 A and 5B, after handling 3d continuously with 200mmol/L NaCl, the blade of non-transgenic tobacco has only undermost two to wilt, rest blade all keeps certain turgescence, still keep certain turgescence and compare the non-transgenic tobacco with transgene tobacco except lobus cardiacus and second layer blade, rest blade is all wilted.Shown in Fig. 5 C and 5D, to recover under normal operation to grow behind the 3d, the recovery extent that can see the transgene tobacco blade is significantly better than the non-transgenic tobacco.The result shows: the salt tolerance of changeing the XHDREB genetic tobacco is improved.
3) the cold resistance analysis of transgene tobacco
Compare with non-transgenic tobacco seedling, will change XHDREB gene masculine tobacco and, at room temperature recover cultivation 3d then, observe its profile impinging upon 24h processing continuously in 4 ℃ of incubators.
Shown in Fig. 6 A-6D, 4 ℃ of wilting degree of handling 24h non-transgenic tobacco are obviously greater than transgene tobacco, and after at room temperature recovering to cultivate 3d, the non-transgenic tobacco is wilted fully, the blade blackout is near dead, and the blade major part of transgene tobacco all recovers normal.This shows that the overexpression of XHDREB gene in tobacco improves to the cold resistance tool of transgene tobacco.
Claims (6)
1. sea island cotton DREB transcription factor gene, its nucleotide sequence is shown in SEQ ID NO:1.
2. the described sea island cotton DREB of claim 1 transcription factor gene encoded protein matter has the aminoacid sequence shown in the SEQ ID NO:2.
3. the plant expression vector that contains the described sea island cotton DREB of claim 1 transcription factor gene.
4. with the described plant expression vector plant transformed of claim 3 cell, tissue or plant.
5. the application of the described sea island cotton DREB of claim 1 transcription factor gene in cultivating the adversity resistant plant kind.
6. application according to claim 5 is characterized in that: the application of described sea island cotton DREB transcription factor gene in cultivating drought resistant plant variety.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2013181834A1 (en) * | 2012-06-08 | 2013-12-12 | 创世纪转基因技术有限公司 | Dreb1 transcription factor of cotton and coding gene and application thereof |
| WO2013185256A1 (en) * | 2012-06-11 | 2013-12-19 | 创世纪转基因技术有限公司 | Dreb1-like transcription factor of cotton and coding gene and application thereof |
| WO2013185257A1 (en) * | 2012-06-11 | 2013-12-19 | 创世纪转基因技术有限公司 | Dreb1-like transcription factor of cotton and coding gene and application thereof |
| CN112111508A (en) * | 2019-06-20 | 2020-12-22 | 新疆农业科学院核技术生物技术研究所（新疆维吾尔自治区生物技术研究中心） | Cotton stress-tolerant gene GhCBF and coding protein and application thereof |
| CN113584018A (en) * | 2021-08-24 | 2021-11-02 | 海南师范大学 | Improved CTAB method for extracting nucleic acid from mallotus japonicus |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2013181834A1 (en) * | 2012-06-08 | 2013-12-12 | 创世纪转基因技术有限公司 | Dreb1 transcription factor of cotton and coding gene and application thereof |
| WO2013185256A1 (en) * | 2012-06-11 | 2013-12-19 | 创世纪转基因技术有限公司 | Dreb1-like transcription factor of cotton and coding gene and application thereof |
| WO2013185257A1 (en) * | 2012-06-11 | 2013-12-19 | 创世纪转基因技术有限公司 | Dreb1-like transcription factor of cotton and coding gene and application thereof |
| CN103703020A (en) * | 2012-06-11 | 2014-04-02 | 创世纪转基因技术有限公司 | Dreb1-like transcription factor of cotton and coding gene and application thereof |
| CN103703020B (en) * | 2012-06-11 | 2017-12-12 | 创世纪种业有限公司 | A DREB1 classes transcription factor for cotton and its encoding gene and application |
| CN112111508A (en) * | 2019-06-20 | 2020-12-22 | 新疆农业科学院核技术生物技术研究所（新疆维吾尔自治区生物技术研究中心） | Cotton stress-tolerant gene GhCBF and coding protein and application thereof |
| CN113584018A (en) * | 2021-08-24 | 2021-11-02 | 海南师范大学 | Improved CTAB method for extracting nucleic acid from mallotus japonicus |
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