CN102373217A - Paddy DREBs (dehydration-responsive element binding) transcription factor and application thereof - Google Patents

Abstract

The invention belongs to the technical field of gene engineering, in particular relates to a paddy DREBs (dehydration-responsive element binding) transcription factor and application thereof, and discloses application of the paddy DREBs transcription factor in preparing transgenic drought-resistant plant varieties. In the transcription factor, the nucleotide sequence is shown as SEQ ID NO.1; or the amino acid sequence is an amino acid sequence which has at least 90 percent homology with the amino acid sequence shown as SEQ ID NO.2 and can improve plant stress resistance. The DREBs transcription factor and a gene thereof have remarkable effect in osmotic stress resistance, so that after the gene provided by the invention is combined with any stress-inducible promoter, introduced to a proper expression carrier and then transformed to a plant host, loss on biological production of various crops planted on a drought land can be remarkably reduced.

Description

A kind of paddy rice DREBs class transcription factor and application thereof

Technical field

The invention belongs to gene engineering technology field, be specifically related to a kind of paddy rice DREBs class transcription factor and application thereof.

Background technology

Plant can suffer from various physical environments in process of growth, wherein some natural disaster often causes a large amount of underproduction of farm crop, like arid, stagnant waterlogging and disease and pest etc.Cultivating the resistance crop varieties is one of important goal of agricultural cience and farming techniques research.When previous important breeding technique be various adversity genes are carried out farm crop genetic engineering modified, to obtain degeneration-resistant new crop varieties.

For resisting coercing of bad external environment, plant soma is experienced the variation of external environment and is passed the signal along in the cell, and the meeting inducing cell is expressed various response genes and resisted the injury of poor environment to plant materials jointly.DREBs (Dehydration-responsive element binding) type of transcription factor is special as AP2/EREBP (APETALA2/ethlene responsive element binding protein; The APETALA2/ element responsive to ethylene is conjugated protein) class members of family, involved in plant being replied and responding abiotic stress such as arid, high salt and low temperature.DRE (Dehydration-responsive element can discerned and combine to DREBs class transcription factor specifically; The arid response element); And then participate in the transmission of adverse circumstance signal, and the expression of regulation and control downstream adverse circumstance response gene, thus plant resistance to environment stress is carried out comprehensive improvement.Yamaguchi-Shinozakiaib equal in the nuclear extract of Arabidopis thaliana, to detect first in 1994 can with rho factor DRBF1 (the Yamaguchi-Shinozakiaib K of DRE effect; Shinozaki K.Plant Cell.1994,6 (2): 251-264).Subsequently the clone of a plurality of these genoids with discover; The DRE/CRT cis-acting elements is prevalent in the promotor of stress response genes involved; This cis-acting elements can discerned and combine to the DREBs transcription factor specifically, and then participate in regulation and control Expression of Related Genes and function enforcement.

But it is found that this gene family is extended familys, in paddy rice, just have 196 members, not only relevant with the abiotic stress response of plant, and regulation and control growth and development of plant and fruit maturation.Find that member new in this gene family coerces the effect in the reaction in growth and development of plant with biological, the resistance that searching can improve plant can be improved the gene of plant-growth again, will the new variety of plant of cultivating stable high yield be had great importance.

Summary of the invention

Technical problem to be solved by this invention is to find new DREBs class transcription factor and nucleotide sequence thereof, to expand the gene that can be applicable in the current Plant Biotechnology improved the plant drought ability, obtains novel transgenic drought-resistant plant variety.

For this reason, the invention discloses the application of a kind of paddy rice DREBs class transcription factor in preparation transgenic drought-resistant plant variety, it is characterized in that:

(a) its aminoacid sequence is shown in SEQ ID NO.2; Or

(b) its aminoacid sequence is at least with aminoacid sequence 90% homology shown in the SEQ ID NO.2 and the aminoacid sequence with the resistance that improves plant.

In specific embodiment, above-mentioned (a) DREBs class transcription factor is generated by nucleic acid sequence encoding shown in SEQ ID NO.1.

On the other hand, the invention discloses a kind of recombinant vectors that contains above-mentioned nucleotide sequence.

In one embodiment, said nucleotide sequence is shown in SEQ ID NO.1.

On the other hand, the invention also discloses a kind of transformant that contains above-mentioned recombinant vectors.

In certain embodiments; Said transformant is vegetable cell or organism; Said organism is the transgenic drought-resistant plant; For one of in paddy rice, corn, wheat, barley, tobacco, soybean, Chinese sorghum, cotton, crudefiber crop, peanut, rape, sesame, sugarcane or the beet, wherein be preferably paddy rice.

DREBs class transcription factor according to the invention and gene thereof; Have to coerce and have tangible effect at impermeabilisation; Therefore can gene according to the invention be combined back importing suitable expression vector and transform plant host with any adverse circumstance evoked promoter, can obviously be reduced in the loss of various farm crop on biological yield of planting on the Arid lands.

Description of drawings

Fig. 1. adopt ClustalW software with the protein sequence of OsDREB1 predictive genes and the DREB homologous protein sequence figure that compares;

Fig. 2. the structure synoptic diagram of expression vector pCAMBIA1300-OsDREB1;

Fig. 3 .real-time PCR method detects the OsDREB1 gene at the expression level of four leaf phase of paddy rice blade under NaCl and ABA processing.

Fig. 4. the comparison of the upgrowth situation of non-transgenic plant and two overexpression transfer-gen plants (T1) under drought stress.

Embodiment

At this paper; Term " isolating ", " purifying " DNA are meant; This DNA or fragment have been arranged in the sequence of its both sides under native state separates, and refers to that also this DNA or fragment with under the native state follow the component of nucleic acid to separate, and separates with in cell, following its protein.

The present invention also comprises the variant form of open reading frame sequence among the proteic SEQ ID NO.1 with OsDREB1 identical function that can encode.These variant forms comprise (but being not limited to): several (are generally 1-90; 1-60 preferably, more preferably 1-20,1-10 best) disappearance, insertion and/or the replacement of Nucleotide; And several (are generally in 60 to hold interpolation 5 ' and/or 3 '; Preferably being in 30, more preferably is in 10, is in 5 best) Nucleotide.

In the present invention, also comprise having and variant form OsDREB 1 identical function, SEQ ID NO.2 sequence.These variant forms comprise (but being not limited to): several (are generally 1-50; Preferably 1-30; 1-20 more preferably, 1-10 best) amino acid whose disappearance, insertion and/or replacement, and add one or several at C-terminal and/or N-terminal and (be generally in 20; Preferably being in 10, more preferably is in 5) amino acid.For example, in the art, when replacing, can not change proteinic function usually with the close or similar amino acid of performance.Again such as, add one or several amino acid at C-terminal and/or N-terminal and also can not change proteinic function usually.

Proteic percent homology is analyzed (GCG program) by GAP (Needleman and Wunsh, 1970) and is confirmed, parameter gap creation penalty=5 wherein, gap extension penalty=0.3.

When the sequence length of being analyzed was at least 15 amino acid, GAP just analyzed and tests in 15 the amino acid whose zones that are at least of two sequences of participating in test.More preferably, when the sequence length of being analyzed was at least 50 amino acid, GAP just analyzed and tests in 50 the amino acid whose zones that are at least of two sequences of participating in test.More preferably, when the sequence length of being analyzed was at least 100 amino acid, GAP just analyzed and tests in 100 the amino acid whose zones that are at least of two sequences of participating in test.More preferably, when the sequence length of being analyzed was at least 250 amino acid, GAP just analyzed and tests in 250 the amino acid whose zones that are at least of two sequences of participating in test.Even more preferably, when the sequence length of being analyzed was at least 500 amino acid, GAP just analyzed and tests in 500 the amino acid whose zones that are at least of two sequences of participating in test.

Can separate and purifying polynucleotide (DNA or RNA), carrier, transformant and organism through methods known in the art.

The isolated polynucleotide of the present invention include but not limited to: the nucleotide sequence of SEQ ID NO.1 coding OsDREB 1 gene; Perhaps this nucleotide sequence can with SEQ ID NO.1 in from the nucleotide sequence hybridization of Nucleotide 532-1404 position; Perhaps its function is equivalent to the subfragment of sequence shown in the SEQ ID NO.1.

Can adopt the OsDREB1 gene of having cloned as probe, screening obtains gene of the present invention or homologous gene from cDNA and genomic library.Equally also can adopt PCR (polymerase chainreaction) technology, amplification obtains OsDREB1 gene and any section of DNA or its homologous section of DNA of this aspect from genome, cDNA.

Being used for carrier of the present invention can be like phage, plasmid, clay, minichromosome, virus or retroviral vector.The carrier that can be used for cloning and/or express polynucleotide of the present invention is to duplicate and/or to express the carrier that duplicates and/or express polynucleotide in the host cell of polynucleotide at need.Generally speaking; The recombinant expression vector that carries nucleotide sequence of the present invention can use Ti-plasmids, plant viral vector, and conventional biotechnological means such as directly DNA conversion, microinjection, electroporation imports vegetable cell (Weissbach, 1998; Method for Plant Molecular Biology VIII; Academy Press, New York, pp.411-463; Geiserson and Corey, 1998, Plant Molecular Biology (2nd Edition).

Having developed several different methods is used for via the complementary sticky end polynucleotide being operated with carrier and links to each other.For example, can add complementary with the aggressiveness sequence fragment by the DNA section in desire is inserted carrier DNA.Then through the hydrogen bond connection carrier between the complementary homopolymeric tail and DNA section to form recombinant DNA molecules.

The synthetic linker that contains one or more restriction site provides the method for another kind of connection DNA section and carrier.Handle the DNA section that produces through the endonuclease restrictive diges-tion with phage T4DNA polysaccharase or e. coli dna polymerase I; Described two kinds of polysaccharases with its 3 '; It is terminal that 5 '-exonucleolytic activity is removed outstanding γ-strand, and mend flat 3 '-recessed end with its polymerization activity.Therefore, these active associatings have produced flush end DNA section, and the enzyme that connects at ability catalysis flush end dna molecular then is as under the existence of phage T4DNA ligase enzyme being incubated the linkers of flush end section with molar excess.Therefore, reaction product is the DNA section that end carries the polylinker sequence, uses these DNA sections of suitable Restriction Enzyme cracking then, and is connected in the expression vector of using enzymatic lysis, and said enzyme can produce and the compatible end of said DNA section.Can buy the synthetic linker that contains a plurality of restriction endonucleases site from a plurality of businessmans.

The polynucleotide inset should be operably connected to on the compatible suitable promotor of the host cell of expressing polynucleotide, and promotor can be strong promoter and/or inducible promoter.The example of some promotors of enumerating comprises phage PL promotor, intestinal bacteria lac, trP, phoA, tac promotor, SV40 is early stage and late promoter and retrovirus LTR promotor; Other suitable promotor is well known by persons skilled in the art.The express recombinant carrier further contains transcription initiation, termination site, and contains the ribosome bind site that is useful on translation at transcriptional domain.The encoding part of the transcript that recombinant vectors is expressed can comprise the translation initiation codon that is positioned at the starting point place and suitably be positioned at by the terminator codon (UAA, UGA or UAG) of the end of translation polypeptide.

As stated, expression vector can comprise at least one selective marker.Said mark comprises the antibiotic resistant gene of coding, for example: neomycin phosphotransferase (Neomycin phosphotransferase) gene npt II, hygromix phosphotransferase (Hygromycin phosphotransferase) gene hpt and Tetrahydrofolate dehydrogenase (Dihydrofolate reductase) gene dhfr; Another kind of is the coding herbicide resistance gene; For example, careless fourth phosphinothricin acetyl transferring enzyme (Phosphinothricin acetyltransferase) gene bar, 5-enol pyruvoyl oxalic acid-3-phosphate synthase (5-Enoylpyruvate shikimatr-3-phosphate) gene epsps.Suitably host's representative example includes but not limited to: protoplasm somatocyte and vegetable cell.The appropriate culture medium of above-mentioned host cell and culture condition are known in the art.

The method for transformation of goal gene or polynucleotide of interest: one type is carrier mediated method for transformation; Be about to along with the transfer of carrier DNA goal gene to be imported in the Plant Genome on the carrier molecules such as DNA of plasmid or virus that goal gene is inserted into Agrobacterium; Agriculture bacillus mediated and virus-mediated method just belongs to this method.Second type is the gene direct guiding method, is meant that the method through physics or chemistry directly imports the external source goal gene in the genome of plant.Physical method comprises that particle gun conversion method, electricity swash conversion method, supersonic method, microinjection and laser microbeam method etc.; Chemical process has PEG mediated transformation method and liposome method etc.The 3rd type is the germplasm systems approach, and this comprises pollen tube passage method, sexual cell dip method, blastular and ovary injection etc.

Among the present invention, the term of use " transformant " (transformant) promptly has the host cell or the organism of allogeneic dna sequence DNA molecule.

The present invention also comprises the host cell that contains nucleotide sequence of the present invention, and said nucleotide sequence can be operated with one or more allos control region (like promotor and/or enhanser) through technology known in the art and link to each other.Can select to regulate the expression of the gene order of inserting, or can be according to the required particular form modification and the host strain of processed gene product.In the presence of some inductor, the expression that some promotor starts can raise.

Can identify by successful cell transformed through well-known technology, promptly contain the cell or the organism of the recombinant vectors of nucleotide sequence according to the invention.

Below in conjunction with specific embodiment, further illustrate the present invention.Should be understood that these embodiment only to be used to the present invention is described and be not used in the restriction scope of the present invention.The experimental technique of unreceipted actual conditions in the following example, usually according to normal condition, or the condition of advising according to manufacturer.

Embodiment 1: separating clone OsDREB1 gene

Through Nese Sreenivasulu et al.2006 at The Plant Journal (2006; 47; The barley grain of 310-327) delivering is grown macroarray result, finds to express sequential label B Q469825/BU986453 and in the grouting ripening stage dormin (ABA) and drought stress is responded.Through homology search paddy rice homologous gene; Adopt ClustalW software (public use software) that the protein sequence and the DREB homologous protein sequence of OsDREB1 predictive genes are compared (among Fig. 1; Ta:GI:84795234, sequence source: Triticum aestivum wheat; Os1:GI:115476366, sequence source: Oryza sativa paddy rice; Os2:GI:31745669, sequence source: Oryza sativa paddy rice; Sb:GI:242044532, sequence source: Sorghum bicolor Chinese sorghum; Zm:GI:195626986, sequence source: Zea mays corn; Pt:GI:224063209, sequence source: Populus trichocarpa black poplar; At:GI:13877779, sequence source: Arabidopsis thaliana Arabidopis thaliana), find that this gene is positioned at paddy rice the 9th karyomit(e) normal chain 12,215,431-12,217,345, called after OsDREB1.Adopt TRIzol reagent (GIBCO BRL, USA) the total RNA of extracting rice leaf.(Tiangen China) becomes cDNA with its reverse transcription to utilize ThermoScript II MLV.With primer DBF (5 '-CCCGCCCACCATTCCCACTTTCGC-3 ') and primer DBR (5 ' CACAACAATTTTAACAACGACGCTAA-3 '), amplify the full-length cDNA (1914bp) of gene.The PCR reaction conditions is: 94 ℃ of preparatory sex change 3min; 94 ℃ of 30sec, 55 ℃ of 30sec, 72 ℃ of 90sec totally 35 circulations; 72 ℃ are extended 5min.The PCR product that obtains increasing is connected into the pGEM-T carrier, and (Promega, USA), screening positive clone also checks order, and obtains the full length sequence (SEQ ID NO.1) of OsDREB1 gene.

The structure and the genetic transformation of embodiment 2:OsDREB1 gene overexpression vector

2.1 contain the structure of destination gene expression carrier:

According to the full length sequence (SEQ ID NO.1) of OsDREB1 gene, design amplifies the primer that complete coding is read frame, and on upstream primer and downstream primer, introduces restriction endonuclease sites (this is decided by the carrier of selecting for use) respectively, so that construction of expression vector.Amplified production to obtain among the embodiment 1 is a template, through high-fidelity Taq enzyme pfu enzyme (Tiangen, China) carry out pcr amplification after, with the OsDREB1 base

Because of cDNA is cloned into intermediate carrier (like pBI23), further transformed into escherichia coli DH5 α is guaranteeing to identify intermediate carrier under the correct prerequisite of reading frame; The extracting plasmid uses EcoRI and HindIII digested plasmid then, so just gene two has been added transcripting promoter and terminator respectively; Constituted a complete expression unit; Reinstall among the plant expression vector pCAMBIA1300, transform Agrobacterium EHA105, carry out the experiment of rice callus metaplasia at last.

2.2 rice genetic transforms

2.2.1 seed disinfection

After shelling, the fine rice paddy seed of sophisticated Japan puts into aseptic triangular flask, with 75% alcohol-pickled 1-2min, and aseptic water washing 2 times; With the 30%NaClO 30min that sterilizes, need often shake therebetween, with aseptic washing 3-4 time, blot redundant moisture with aseptic filter paper again, seed is inoculated on the callus inducing medium (MS+2,4-D 2.0mg/L), about 30 of every ware is in 28 ℃ of dark cultivations.

2.2.2 succeeding transfer culture

Through inducing of nearly January, paddy rice grows the callus that yellow is expanded, and removes its scultellum, callus is gone on the fresh callus inducing medium (MS+2,4-D 2.0mg/L) carry out subculture.Per 2 all subcultures once, general subculture can obtain suitable genetically modified bright yellow, granular embryo callus for 2-4 time., select embryo property particle and be used for genetic transformation after 2 weeks at succeeding transfer culture.

2.2.3 the cultivation of Agrobacterium

Picking list bacterium colony is cultivated in 1ml Agrobacterium substratum on the conversion flat board.In 50ml Agrobacterium substratum (containing corresponding microbiotic), add the above-mentioned culture of 1ml, 200rpm, 28 ℃ are cultivated 5-6hr to OD600 is 0.6-1.0, cultivates the preceding 2hr of end and adds Syringylethanone (AS, final concentration 100uM).Get above-mentioned bacterium liquid at room temperature, 4000rpm, 10min abandons supernatant, adds the resuspended thalline of MS liquid nutrient medium (containing AS 100uM), with on cultivate 2hr under the identical condition, make the OD600=0.5-1 of bacterium liquid, can be used to transformed calli this moment.AS＝acetosringone

2.2.4 cultivate altogether

The EMBRYO IN RICE callus is immersed Agrobacterium bacterium liquid 20-30min, use aseptic thieving paper suck dry moisture again, the callus that will infect places on the common culture medium (MS+2,4-D 2.0mg/L+AS 100uM), 28 ℃ of dark cultivations three days.

2.2.5 wash bacterium

The callus of cultivating altogether is earlier with aseptic water washing 3 times, is immersed in the MS liquid nutrient medium that contains Cef/CN 400mg/L behind the 20-30min again, callus changed on the aseptic filter paper blot.

2.2.6 select to cultivate

The callus of suck dry moisture is inoculated on the selection substratum (MS+2,4-D 2.0mg/L+Hyg 30mg/L+Cef 400mg/L).After 3 weeks, select the callus that newly grows and be inoculated on the selection substratum (MS+2,4-D 2.0mg/L+Hyg 50mg/L+Cef 250mg/L), selected for 2 weeks again.

2.2.7 differentiation culture

To be transferred to presorting substratum (N6+KT 2.0mg/L+NAA 0.2mg/L+6-BA 2.0mg/L+Hyg 30mg/L+Cef 200mg/L+ agar 9g/L+ sucrose 45g/L) through the resistant calli that 2 selections obtain and go up dark the cultivation about 10 days, and forward division culture medium (N6+KT 2.0mg/L+NAA 0.2mg/L+6-BA 2.0mg/L+Hyg 30mg/L+ agar 4.5g/L+ sucrose 30g/L) again to and go up the illumination cultivation.

2.2.8 root culture

About 1-2 month, forward seedling high about 2cm to generation that root media (1/2MS+Hyg 15mg/L+ agar 4.5g/L+ sucrose 20g/L) is gone up inducing adventitious root.

2.2.9 the transplanting of transgenic seedling

When seedling grows to 10cm when high, seedling is taken out, clean the solid medium that adheres to sterilized water, move in the earth, just begun to treat to take off lens again behind the robust plant with lens cover several days, cultivate in the greenhouse.

Embodiment 3: the expression analysis of endogenous OsDREB1 gene in paddy rice

3.1 material is prepared

After the fine germination of rice varieties Japan, transplant in liquid nutrient medium (tap water is mixed with the 1/5MS macroelement).Behind the growth of seedling 15d, use the sodium-chlor of 50mM concentration and 100 μ M dormins (ABA) to handle respectively respectively 2 minutes, 5 minutes, 30 minutes, 2 hours and 6 hours, the clip blade drops into the liquid nitrogen preservation fast then, is used for the extracting of RNA.

3.2 there is not total RNA preparation of DNA

The leaf RNA extraction agent box working instructions extracting in a small amount that provides by Shanghai China Shun Bioisystech Co., Ltd.Use Beckman Coulter ^TMDU

640 ultraviolet spectrophotometers are measured RNA concentration.For removing the DNA that remains among the RNA; Each total RNA sample is got 5 μ g; Add 1 μ L DNAaseI (American I nvitrogen company) and 1 μ L10 * reaction buffer, supply volume to 10 μ L, normal-temperature reaction 30min; Every then pipe adds 1 μ L 2mmol L-1EDTA termination reaction, makes DNAase I inactivation at 70 ℃ of heating 10min at last.

3.3 the first chain cDNA's is synthetic

Above-mentioned RNA sample is respectively got 2 μ L; The reagent that provides by U.S. Promega company reverse transcription test kit adds 4 μ L 25mmol L-1MgCl2 successively; 2 μ L10 * RT damping fluid, mixed liquid of 2 μ L dNTP and 1 μ L oligo (dT) 15 add water and supply volume to 18.5 μ L; At 70 ℃ of heat denatured 10min, fast in cooled on ice.Add 0.5 μ L RNase inhibitor and 1 μ L AMVRTase then, at 42 ℃ of water-bath 60min, 70 ℃ are heated the 10min termination reaction down.

3.4 quantitative PCR

Sequences Design Auele Specific Primer RF:5 '-CCGTGCAGATGGGATTTTAGAC-3 ' according to gene OsDREB1; RR:5 '-CTGCCGGTAGCATCGGCGTC-3 ' is used for quantitative fluorescent PCR; According to cDNA sequences Design Auele Specific Primer the AF:5 '-CTTCCTCATGCCATCCTGC-3 ' of Actin (GenBank accession No.AY212324) gene, AR:5 '-GCAAGCTTCTCCTTGATGTCC-3 ' is used for the quantitative fluorescent PCR of reference gene.PCR uses American AB I PRISM

7000 quantitative PCR appearance, and each PCR is provided with once and repeats.Reaction system comprises SYBR Premix ExTaq ^TM(2 *) 10 μ L, each 0.5 μ L of forward and reverse primer, the cDNA template 1 μ L of various processing adds water and supplies volume to 25 μ L.Response procedures is: 95 ℃ of 30s, and then at 95 ℃ of 10s, 60 ℃ of 35s circulate 40 times down; Be set in each circulation and read fluorescent value during 60 ℃ of 35s; Carry out the ROX value simultaneously and proofread and correct, add the analysis of fluorescence PCR products melt curve analysis at last, other operations see the instrument working instructions for details.In order to detect the pollution that whether has DNA in the RNA sample, 3 samples of picked at random are respectively got 1 μ L

RNA carries out PCR as template, and method is the same.

3.5 analytical procedure

Ct produces through manual 0.2 back of confirming as through the fluorescence thresholding of 7000system SDS Version1.2.3 software at PCR, enters data into EXCEL and carries out computational analysis.Data analysis employing method is 2 ^{-Δ Δ CT}, utilize the EXCEL table to make to express the difference histogram then.

3.6 analytical results

50mM sodium-chlor is handled after 5 minutes this expression of gene amount and is obviously brought up to 2 times, and expression amount progressively raises subsequently; 100 μ MABA handled after 5 minutes, and the rising of its expression amount reaches rapidly about 6 times, expresses after 30 minutes and tends towards stability, and meets the characteristic of transcription factor at the early expression of gene.Explain that this gene is to rely on ABA, progressively strengthens and expresses (see figure 3) under infiltration and ion stress conditions.

T is for the upgrowth situation of family plant under drought condition for embodiment 4:OsDREB1 gene overexpression transgenic

Chosen among 2 embodiment 2 OsDREB1 gene overexpression transgenic T1 and carried out the drought stress experiment of fringe phase for the family plant.Concrete steps are following: every part of material plantation 3 row; Every row 7 strains, most materials gets into ear differentiation period (paddy rice to moisture tricky time) in colony, beginning emptying field moisture; 2 of drought stresses are observed the upgrowth situation of transfer-gen plant more than week then.The result shows that the growth of 2 transfer-gen plants of the present invention clone's OsDREB1 gene overexpression does not have tangible difference with contrast under normal operation, will be superior to contrast significantly but handle its growth of back at environment stress.After arid handled for 3 weeks, observe, the surviving rate (average 82%) of finding transfer-gen plant is apparently higher than transfer-gen plant (36%) not, and setting percentage (average 73%) is also apparently higher than transfer-gen plant (32%) not.These 3 parts of materials are copied kind of a test; The raising of finding the real grain of every fringe number, setting percentage and harvest index reaches utmost point level of signification; Its raising the output has also reached level of signification, and than non-transgenic plant raising the output 200%-300%, plant height becomes short (Fig. 4 although arid is handled down; Contrast expression non-transgenic plant, OX1 and OX2 represent overexpression transfer-gen plant 1 and 2.Effectively fringe, every fringe real grain number and every fringe grain husk spend several units for, the unit of setting percentage and harvest index is a per-cent, plant height unit be centimetre that the unit of economics output and overground part biological yield restrains. ^*Expression P≤0.001, ^*Expression P≤0.05).The OsDREB1 base is described

The expression of cause can be alleviated the paddy rice plant strain growth that drought stress causes after getting into reproductive growth and is obstructed, and has improved the setting percentage and seed grouting of plant, and the enhancing transgenic paddy rice improves rice yield to the resistance of abiotic stress.

Scope of the present invention does not receive the restriction of said specific embodiments, and said embodiment also comprises the method and the component of functional equivalent only as the single example of illustrating all respects of the present invention in the scope of the invention.In fact, except content as herein described, those skilled in the art can easily grasp multiple improvement of the present invention with reference to the description and the accompanying drawing of preceding text.Said improvement also falls within the scope of appended claims.Every piece of reference that preceding text are mentioned is listed this paper in as a reference all in full.

Claims (2)

1. the application of paddy rice DREBs class transcription factor in preparation transgenic drought-resistant plant variety is characterized in that:

(a) its nucleotide sequence is shown in SEQ ID NO.1; Or

(b) its aminoacid sequence is at least with aminoacid sequence 90% homology shown in the SEQ ID NO.2 and the aminoacid sequence with the resistance that improves plant.

2. application as claimed in claim 1 is characterized in that said transgenic drought-resistant plant is one of in paddy rice, corn, wheat, barley, tobacco, soybean, Chinese sorghum, cotton, crudefiber crop, peanut, rape, sesame, sugarcane or the beet.

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