CN114644694A - ZmOST1 protein, coding gene thereof and application thereof in cultivating drought-resistant plants - Google Patents

Abstract

The invention discloses ZmOST1 protein, a coding gene thereof and application thereof in cultivating drought-resistant plants. The ZmOST1 protein provided by the invention has an amino acid sequence of SEQ ID No.1 or is a fusion protein which is obtained by substituting and/or deleting and/or adding one or more amino acid residues of the ZmOST1 protein, has the same function with corn or more than 80% of identity with the corn and has the same function with the corn or is obtained by connecting a protein tag at the N end and/or the C end of the protein. Compared with the wild drought resistance, the drought resistance of the positive strain obtained by over-expressing ZmOST1 in the corn B73 is enhanced, compared with the traditional breeding mode, the time of the positive strain is short, the purpose is strong, and the positive strain has important significance for cultivating and improving new varieties of drought-resistant plants.

Description

ZmOST1 protein, coding gene thereof and application thereof in cultivating drought-resistant plants

Technical Field

The invention relates to the technical field of biology, in particular to ZmOST1 protein, a coding gene thereof and application thereof in cultivating drought-resistant plants.

Background

More than half of the corns in China are planted on dry land which depends on natural rainfall in the northwest, southwest, northwest and northeast areas, the water loss is fast, the rainfall rate is high, and the growth of the corns is seriously influenced. Corn is an important food crop in China, and the research on the drought resistance of the corn is of great significance for solving the problem of corn yield in China.

Drought can cause osmotic stress in plants, the osmotic pressure in the plant is lower than the osmotic pressure of the environment (such as soil solution), and the plant cannot absorb water or even lose water. Osmotic stress has two pathways: ABA-dependent and ABA-independent pathways. When subjected to environmental stress, ABA, PYL, PYR and RCAR are ABA receptors generated in an ABA pathway and can be combined with ABA. The activity of protein phosphatase of PP2C is inhibited, so that SnRK2 has kinase activity and can phosphorylate downstream transcription factors and the like, thereby regulating and controlling the expression of downstream stress response genes, inhibiting stomatal opening, regulating and controlling ABA sensitivity and generating other responses. DREB2A is phosphorylated, thereby regulating downstream stress response gene expression, independent of the ABA pathway. The drought stress can induce the abscisic acid (ABA) in the plant body, the ABA can guide stomata to close in time so as to reduce the water loss in the plant body, and simultaneously the ABA can activate the expression of a large number of drought-related genes so as to cause various drought stress responses of the plant. ABA is an extremely important plant hormone, and plays a key role in stomata closure, expression of stress response genes, and the like.

Sucrose non-glycolysis related protein kinase (SnRK) is a serine/threonine (Ser/Thr) protein kinase specific to plants and plays an important role in plant stress resistance research. Under drought conditions, ABA hormone level rises, then the ABA hormone level is combined with an ABA receptor PYR/PYL/RCARs, then a receptor protein can be combined with downstream phosphokinase PP2C to inhibit the activity of protein phosphatase, and further SnRK protein kinase in an inactivated state is released. OST1(Open Stomata 1) is a typical SnRK protein kinase, originally found in fava beans and named AAPK, which is activated by ABA signals and is involved in regulating stomatal movement. There are ten members of the SnRK2 family in Arabidopsis thaliana, named SnRK2.1-SnRK2.10, where OST1 is SnRK2.6. There are twelve members of the SnRK2 family in maize, designated SnRK2.1-SnRK2.12, where OST1 is SnRK2.8.

Some related reports on the function study of the SnRK family proteins in plants exist at present, but the role of the SnRK family proteins in monocotyledonous crop corn is not clear.

Disclosure of Invention

The invention aims to provide ZmOST1 protein, a coding gene thereof and application thereof in cultivating drought-resistant plants.

In a first aspect, the invention claims a protein.

The protein claimed by the invention is derived from corn (Zea mays L.) and is named ZmOST1 protein. Specifically, any one of the following may be used:

(A1) protein with an amino acid sequence of SEQ ID No. 1;

(A2) the amino acid sequence shown in SEQ ID No.1 is substituted and/or deleted and/or added by one or more amino acid residues and is derived from the protein with the same function of corn;

(A3) a protein having 99% or more, 95% or more, 90% or more, 85% or more, or 80% or more identity to the amino acid sequence defined in any one of (A1) - (A2) and derived from maize having the same function;

(A4) a fusion protein obtained by attaching a protein tag to the N-terminus and/or C-terminus of the protein defined in any one of (A1) to (A3).

In the above proteins, the protein tag (protein-tag) refers to a polypeptide or protein that is expressed by fusion with a target protein using in vitro recombinant DNA technology, so as to facilitate the expression, detection, tracking and/or purification of the target protein. The protein tag may be a Flag tag, a His tag, an MBP tag, an HA tag, a myc tag, a GST tag, and/or a SUMO tag, among others.

In the above proteins, identity refers to the identity of amino acid sequences. The identity of the amino acid sequences can be determined using homology search sites on the Internet, such as the BLAST web pages of the NCBI home website. For example, in the advanced BLAST2.1, by using blastp as a program, setting the value of Expect to 10, setting all filters to OFF, using BLOSUM62 as a Matrix, setting Gap existence cost, Per residual Gap cost, and Lambda ratio to 11, 1, and 0.85 (default values), respectively, and performing a calculation by searching for the identity of a pair of amino acid sequences, a value (%) of identity can be obtained.

In the above protein, the homology of 95% or more may be at least 96%, 97%, 98% identity. The homology of 90% or more may be at least 91%, 92%, 93%, 94% identity. The homology of 85% or more may be at least 86%, 87%, 88%, 89% identity. The homology of 80% or more may be at least 81%, 82%, 83%, 84% identity.

In a second aspect, the invention claims nucleic acid molecules encoding the proteins described in the first aspect hereinbefore.

Further, the nucleic acid molecule may be DNA, such as cDNA, genomic DNA, or recombinant DNA; the nucleic acid molecule may also be RNA, such as mRNA, and the like.

Furthermore, the nucleic acid molecule (ZmOST1 gene) is specifically any one of the following:

(B1) a DNA molecule (complete genome sequence) shown as SEQ ID No. 2;

(B2) a DNA molecule (T01 transcript) as shown in positions 843-4266 of SEQ ID No. 2;

(B3) a DNA molecule (cDNA) shown as SEQ ID No. 3;

(B4) a DNA molecule which hybridizes under stringent conditions to a DNA molecule as defined in any one of (B1) to (B3) and which encodes a protein as defined in the first aspect hereinbefore;

(B5) a DNA molecule having 99% or more, 95% or more, 90% or more, 85% or more or 80% or more identity to a DNA sequence defined in any one of (B1) to (B4) and encoding a protein as defined in the first aspect hereinbefore.

In the above nucleic acid molecule, the stringent conditions may be as follows: 50 ℃ in 7% Sodium Dodecyl Sulfate (SDS), 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing in2 XSSC, 0.1% SDS at 50 ℃; also can be: 50 ℃ in 7% SDS, 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing at 50 ℃ in 1 XSSC, 0.1% SDS; it can also be: 50 ℃ in 7% SDS, 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing in 0.5 XSSC, 0.1% SDS at 50 ℃; also can be: 50In 7% SDS and 0.5M Na at DEG C₃PO₄Hybridization with 1mM EDTA, rinsing in 0.1 XSSC, 0.1% SDS at 50 ℃; also can be: 50 ℃ in 7% SDS, 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing in 0.1 XSSC, 0.1% SDS at 65 ℃; can also be: in a solution of 6 XSSC, 0.5% SDS at 65 ℃ and then washed once with each of 2 XSSC, 0.1% SDS and 1 XSSC, 0.1% SDS.

In the above nucleic acid molecules, homology refers to the identity of nucleotide sequences. The identity of the nucleotide sequences can be determined using homology search sites on the Internet, such as the BLAST web page of the NCBI home website. For example, in the advanced BLAST2.1, by using blastp as a program, setting the value of Expect to 10, setting all filters to OFF, using BLOSUM62 as a Matrix, setting Gap existence cost, Per residual Gap cost, and Lambda ratio to 11, 1, and 0.85 (default values), respectively, and performing a calculation by searching for the identity of a pair of nucleotide sequences, a value (%) of identity can be obtained.

In the above nucleic acid molecule, the 95% or more homology may be at least 96%, 97%, 98% identity. The homology of 90% or more may be at least 91%, 92%, 93%, 94% identity. The homology of 85% or more may be at least 86%, 87%, 88%, 89% identity. The homology of 80% or more may be at least 81%, 82%, 83%, 84% identity.

In a third aspect, the invention claims an expression cassette, a recombinant vector, a recombinant bacterium or a transgenic cell line comprising a nucleic acid molecule as described in the second aspect above.

The expression cassette refers to a DNA capable of expressing ZmOST1 in a host cell, and the DNA may include not only a promoter for initiating transcription of ZmOST1 gene, but also a terminator for terminating transcription of ZmOST 1. Further, the expression cassette may also include an enhancer sequence. Promoters useful in the present invention include, but are not limited to: constitutive promoters, tissue, organ and development specific promoters, and inducible promoters. Examples of promoters include, but are not limited to: ubiquitin gene Ubiqutin promoter (pU)bi); the constitutive promoter of cauliflower mosaic virus 35S; the wound-inducible promoter from tomato, leucine aminopeptidase ("LAP", Chao et al (1999) Plant Physiol120: 979-992); a chemically inducible promoter from tobacco, pathogenesis-related 1(PR1) (induced by salicylic acid and BTH (benzothiadiazole-7-carbothioic acid S-methyl ester)); tomato proteinase inhibitor II promoter (PIN2) or LAP promoter (both inducible with jasmonic acid ester); heat shock promoters (U.S. patent 5,187,267); tetracycline-inducible promoters (U.S. Pat. No. 5,057,422); seed-specific promoters, such as the millet seed-specific promoter pF128(CN101063139B (Chinese patent 200710099169.7)), seed storage protein-specific promoters (e.g., the promoters of phaseolin, napin, oleosin and soybean beta conglycin (Beachy et al (1985) EMBO J.4: 3047-3053)). They can be used alone or in combination with other plant promoters. All references cited herein are incorporated by reference in their entirety. Suitable transcription terminators include, but are not limited to: agrobacterium nopaline synthase terminator (NOS terminator), cauliflower mosaic virus CaMV 35S terminator, tml terminator, pea rbcS E9 terminator and nopaline and octopine synthase terminators (see, e.g., Odell et al (I)⁹⁸⁵) Nature 313: 810; rosenberg et al (1987) Gene,56: 125; guerineau et al (1991) mol.gen.genet,262: 141; proudfoot (1991) Cell,64: 671; sanfacon et al Genes Dev.,5: 141; mogen et al (1990) Plant Cell,2: 1261; munroe et al (1990) Gene,91: 151; ballad et al (1989) Nucleic Acids Res.17: 7891; joshi et al (1987) Nucleic Acid Res, 15: 9627).

Constructing a recombinant expression vector containing the ZmOST1 gene expression cassette. The plant expression vector used may be a binary Agrobacterium vector or a Gateway system vector, etc., such as pCXUN, pBin438, pCAMBIA1302, pCAMBIA2301, pCAMBIA1301, pCAMBIA1300, pBI121, pGWB411, pGWB412, pGWB405, pCAMBIA1391-Xa or pCAMBIA 1391-Xb. When ZmEREB167 is used to construct a recombinant expression vector, any one of enhanced, constitutive, tissue-specific or inducible promoters, such as cauliflower mosaic virus (CAMV)35S promoter, ubiquitin gene Ubiqutin promoter (pUbi), etc., can be added before the transcription initiation nucleotide, and can be used alone or in combination with other plant promoters; in addition, when the gene of the present invention is used to construct plant expression vectors, enhancers, including translational or transcriptional enhancers, may be used, and these enhancer regions may be ATG initiation codon or initiation codon of adjacent regions, etc., but must be in the same reading frame as the coding sequence to ensure proper translation of the entire sequence. The translational control signals and initiation codons are widely derived, either naturally or synthetically. The translation initiation region may be derived from a transcription initiation region or a structural gene.

In order to facilitate the identification and screening of transgenic plant cells or plants, plant expression vectors to be used may be processed, for example, by adding a gene encoding an enzyme or a luminescent compound which can produce a color change (GUS gene, luciferase gene, etc.), an antibiotic marker having resistance (gentamicin marker, kanamycin marker, etc.), or a chemical-resistant marker gene (e.g., herbicide-resistant gene), etc., which can be expressed in plants.

In a fourth aspect, the invention claims the use of a protein as described in the first aspect above or a nucleic acid molecule as described in the second aspect above or an expression cassette, recombinant vector, recombinant bacterium or transgenic cell line as described in the third aspect above for modulating drought resistance in a plant.

Further, the method is carried out. In the plant, the expression level and/or activity of the protein is increased, and the drought resistance of the plant is increased; the expression level and/or activity of the aforementioned protein is decreased, and the drought resistance of the plant is decreased.

In a fifth aspect, the invention claims a method of cultivating plants with improved drought resistance.

The method of growing plants with increased drought resistance as claimed in the present invention may comprise the step of increasing the expression level and/or activity of a protein as described in the first aspect hereinbefore in the recipient plant.

The method can be realized by means of hybridization or by means of transgenosis.

In a sixth aspect, the invention claims a method of breeding transgenic plants with improved drought resistance.

The method for cultivating the transgenic plant with improved drought resistance, which is claimed by the invention, can comprise the following steps: introducing into a recipient plant a nucleic acid molecule capable of expressing a protein according to the first aspect of the invention, to obtain a transgenic plant; the transgenic plant has increased drought resistance compared to the recipient plant.

Further, a nucleic acid molecule capable of expressing a protein as described in the first aspect above may be introduced into the recipient plant by means of a recombinant expression vector.

Wherein, the nucleic acid molecule (ZmOST1 gene) can be modified as follows and then introduced into the receptor plant to achieve better expression effect:

1) modifying the sequence of the gene adjacent to the initiating methionine to allow efficient initiation of translation; for example, modifications are made using sequences known to be effective in plants;

2) linking with promoters expressed by various plants to facilitate the expression of the promoters in the plants; such promoters may include constitutive, inducible, time-regulated, developmentally regulated, chemically regulated, tissue-preferred, and tissue-specific promoters; the choice of promoter will vary with the time and space requirements of expression, and will also depend on the target species; for example, tissue or organ specific expression promoters, depending on the stage of development of the desired receptor; although many promoters derived from dicots have been demonstrated to be functional in monocots and vice versa, desirably, dicot promoters are selected for expression in dicots and monocot promoters for expression in monocots;

3) the expression efficiency of the gene of the present invention can also be improved by linking to a suitable transcription terminator; for example tml from CaMV, E9 from rbcS; any available terminator which is known to function in plants may be linked to the gene of the invention;

4) enhancer sequences, such as intron sequences (e.g., from Adhl and bronzel) and viral leader sequences (e.g., from TMV, MCMV, and AMV) were introduced.

In the above aspects, the plant is a monocot or a dicot.

Further, the plant is a gramineous plant.

Further, the plant is a plant of the genus zea, such as corn.

In a specific embodiment of the invention, the plant is maize B73.

Because different transcripts can be generated by the same DNA segment sequence of the corn and different proteins can be translated, the different transcripts generated by the sequence shown in SEQ ID No.2 and the translated different proteins are all in the protection scope of the invention.

More than one transcript of the ZmOST1 gene can resist drought sensitivity after the cDNA corresponding to other forms of transcripts is over-expressed, which belongs to the protection scope of the invention.

Compared with the wild drought resistance, the drought resistance of the positive strain obtained by over-expressing ZmOST1 in the corn B73 is enhanced, compared with the traditional breeding mode, the time of the positive strain is short, the purpose is strong, and the positive strain has important significance for cultivating and improving new varieties of drought-resistant plants.

Drawings

FIG. 1 shows the detection of the expression level of ZmOST1 gene in control (WT) and maize over-expression strain. The OST1 OE is a randomly selected strain from ZmOST1 transgenic line.

Detailed Description

The present invention is described in further detail below with reference to specific embodiments, which are given for the purpose of illustration only and are not intended to limit the scope of the invention. The examples provided below serve as a guide for further modifications by a person skilled in the art and do not constitute a limitation of the invention in any way.

The experimental procedures in the following examples, unless otherwise indicated, are conventional and are carried out according to the techniques or conditions described in the literature in the field or according to the instructions of the products. Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.

The corn ecotype is B73; the agrobacterium strain is EHA 105. The main reagents comprise: restriction enzymes, DNA polymerases, T4 ligases, etc. from biological companies such as NEB and Toyobo; reverse transcription kit from Thermo corporation; RNA extraction kit from magenta; quantitative PCR reagents of Taraka corporation; the plasmid extraction kit and the DNA recovery kit are products of Tiangen company; MS culture medium, agar powder, agarose, ampicillin, kanamycin, gentamicin sulfate, rifampicin and other antibiotics are sigma products; the various other chemical reagents used in the examples were all imported or domestic analytical reagents; primer synthesis and sequencing was done by Yingjun corporation.

Example 1, ZmOST1 protein and application of encoding gene thereof in regulating and controlling plant drought resistance

Discovery of ZmOST1 protein and coding gene thereof

The invention discovers a novel protein from corn (Zea mays L.), named ZmOST1 protein, and the amino acid sequence of the protein is SEQ ID No. 1. The sequence of the ZmOST1 gene in the maize B73 genome is SEQ ID No.2, which consists of 9292 bases, and the reading frame of the T01 transcript is from 843 rd to 4266 th bases of the 5' end of the SEQ ID No. 2. The gene consists of 9 exons, wherein the T01 transcript encodes 5 exons, and the reading frame (i.e. SEQ ID No.2 from 843 th to 4266 th at the 5' end) has 1 st to 192 th bases, 2810 th to 2884 th bases, 2962 th to 3063 th bases, 3147 th to 3200 th bases, 3299 th to 3424 th bases and the rest is the intron sequence thereof. The CDS sequence of ZmOST1 gene is shown in SEQ ID No. 3.

Construction and detection of over-expression vector of ZmOST1 gene

Extracting total RNA from corn (Zea mays L.) B73, reverse transcribing to obtain cDNA, amplifying ZmOST1 gene with cDNA as template and F and R as primer, and enzyme cutting and connecting to over-expression vector.

The vector construction method comprises the following steps:

(1) total RNA of corn B73 was extracted using the RNA extraction kit from magenta, according to the kit instructions.

(2) The RNA was reverse transcribed to give cDNA using a reverse transcription kit from thermo, and the detailed procedures were as described in the kit's instructions.

(3) Using cDNA as a template and F and R as primers, amplifying ZmOST1 gene cDNA, running electrophoresis on the amplification product, cutting gel and recovering, wherein the recovery method refers to a Tiangen company kit.

The primers used were:

an upstream primer F: 5'-ATGGCAGGGCCGGCGCCG-3', respectively;

a downstream primer R: 5'-CTAGAACAAGATGAAGTTGACATGGG-3' are provided.

(4) The recovered ZmOST1 gene cDNA (SEQ ID No.3) was cloned into pBCXUN vector, 5. mu.L of ligation product was taken, and E.coli competence was transformed. Screening was performed on LB plates containing 50. mu.g/mL kanamycin. And (5) identifying the single clone by colony PCR, and selecting a positive clone for sequencing. The obtained recombinant expression vector with correct sequencing is named pBCXUN-ZmOST 1. Colony PCR and sequencing universal primers were as follows:

UbiP-seq：5’-TTTTAGCCCTGCCTTCATACGC-3’；

NosR-seq：5’-AGACCGGCAACAGGATTCAATC-3’。

in the recombinant plasmid pBCXUN-ZmOST1, the ZmOST1 gene shown in SEQ ID No.3 is transcribed from the Ubi promoter and terminated by the Nos terminator, thereby expressing the objective protein ZmOST1(SEQ ID No. 1).

The pBCXUN vector is an expression vector obtained by replacing the HYG gene (hptII, hygromycin resistance gene) of the pCXUN vector (GenBank: FJ905215.1, 06-JUL-2009) with the Bar gene (encoding phosphinothricin acetyltransferase) (GenBank: 284-835 nucleotides in MG719235.1, 02-OCT-2018) and keeping the other nucleotides of pCXUN unchanged.

Construction and detection of ZmOST1 gene over-expression plant

And (3) transforming the pBCXUN-ZmOST1 overexpression plasmid constructed in the second step into a competent Agrobacterium EHA105 strain by a heat shock method, and identifying positive clones by colony PCR. Inoculating single colony of correctly identified Agrobacterium into 2-3mL liquid culture medium containing 100 μ g/mL kanamycin and 50 μ g/mL rifampicin, shake culturing at 28 deg.C overnight, inoculating to liquid culture medium containing large amount of antibiotics in the next day, shake culturing, collecting thallus after several times of inoculation, and resuspending to OD₆₀₀Between 0.8 and 1.0. By usingAnd infecting the young B73 corn embryo picked out under aseptic condition with the obtained recombinant agrobacterium suspension, and inducing the young corn embryo to callus and grow seedlings. Transgenic plants are obtained by self-crossing and seed-breeding T3 generation for subsequent experiments.

RNA of different transgenic inbred lines is extracted, cDNA is reversely transcribed, the relative expression level of the ZmOST1 gene is detected by quantitative PCR, and the result is shown in figure 1, which shows that the expression level of the ZmOST1 gene in the ZmOST1 transgenic plant is about 19 times of that of the B73 of a non-transgenic control plant and is far higher than that of the B73 of the non-transgenic control plant.

Fourth, statistics of drought treatment survival rate of over-expressed corn with ZmOST1 gene

1. Homozygous T identified as positive by step three₃Transgenic corn seeds (OE1 and OE2) and B73 wild-type (WT) corn seeds of the generation ZmOST1 were sown in small pots with nutrient soil, 6 pots per line, 10 pots per pot, respectively.

2. After growing for 7 days in a culture room at 25 ℃, selecting seedlings with consistent growth vigor, transplanting the seedlings into large rectangular pots filled with 2500g of nutrient soil, planting Wild Type (WT) in one half area of each pot, and planting ZmOST1 transgenic lines in the other half area of each pot, wherein the number of the seedlings is three, and each row comprises 5 seedlings, namely 15 seedlings of the wild type and the transgenic lines in each large rectangular pot. Three replicates were set, 5 pots per replicate.

3. After 7 days of growth under normal watering conditions, the watering was stopped for drought treatment.

4. After 20 days, significant differences in the phenotype of the Wild Type (WT) and ZmOST1 transgenic plants were observed.

5. After being rehydrated for 7 days, the survival of each strain is observed and the survival rate is counted. Survival rate is the percentage of the number of surviving plants in each line to the total number of plants. And taking the average value of multiple repetitions for statistical analysis.

The results show that: the survival rate of maize B73 plants (WT) was 40% ± 4%. The survival rate of plants of OE1 strain is 80% + -5%, and the survival rate of plants of OE2 strain is 83 + -4%.

The present invention has been described in detail above. It will be apparent to those skilled in the art that the invention can be practiced in a wide range of equivalent parameters, concentrations, and conditions without departing from the spirit and scope of the invention and without undue experimentation. While the invention has been described with reference to specific embodiments, it will be appreciated that the invention can be further modified. In general, this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains. The use of some of the essential features is possible within the scope of the claims attached below.

<110> university of agriculture in China

<120> ZmOST1 protein, coding gene thereof and application thereof in cultivating drought-resistant plants

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ttcttttatc aactatttgc aaatttgcct ccactttaat attaactttg gaatataatg 2280

atggcttaaa tctgaccata aattttggaa atcaatactt caactttaaa tcaaaaaaat 2340

gctgaaaata aattgaattt gacactacat cttgacaaaa ttgacaacat tttaattgta 2400

attttgaaat atttcaatta taatatattt tcaagatttt aatttaaata ttgttattct 2460

gtcaacattt aatgttaact tcaatttatt ttcatcaatt tagaaatgtt attaaattca 2520

aatttgaaat attgattttc aaagtttatt gttagattta tatcacaatt atattctaaa 2580

ctttcatatc aaagtggcgg cgaatttgca aacaattgat aaaaggaggg acaatggtgc 2640

aagggtatta acgttttttt tgtgggctga gttaacactg ttaagtgatg ataacggaat 2700

agggaaaact acctatctct ttcaaaaaag ggcatacatg taaactttaa aaaacagggg 2760

taaaacgaag ttgtatctcc gagtaggggc aaccattcaa cgacctcaat tggtaatgat 2820

gctccataac tctatgccac aactaacttc tactacttga tgcatgggag tgtatgtctt 2880

ataaaaagaa attgaaacag tatgtgtcta ggacccaaca aaatattgca tgaatttcac 2940

aagaccacat tatactggga aaagtttttg caacttgcaa gcctttcctg ttcctttaac 3000

taatattgag gtagttgcat acttgttata ctgtactgta caagagccga cacgttttga 3060

ttgcatagcg aggtggggga aagtggaatc cttttgatac actatgaagt agaatacttg 3120

tactagtaat tccatctcat tcagctccat ccagccaagc aattgcattg atgtgtaagc 3180

tctttactga atgaacatgt tttatccatg tgtgtgtatt tctaattgga ggtacctgtg 3240

cattgttttg taataccatt gctttactgt tgtggccata tggtctagta gtttttgata 3300

gaacaaataa tcaataatat cttgaacttc tgaaaccaag gatagttgag cagattcttg 3360

ttacattcct ttgattttgg cttcaacccc tgagagacca acatcaattc atactaaaag 3420

cccaatattg tttatgatct ctatcaatag caagacattg ggtctagttt atggaatttc 3480

ctctctcatt catttggtga tttgatttaa ataattccaa tgtccatagt gccttaaatc 3540

tgttttttgt gcattttctt cttctatcat aagtgctagg ttgtcatttt ttactgcact 3600

cagttgtttt atcttggttt tctttaatat gtattccgtt gtcatgtgca gatagatgag 3660

aatgtccagc gtgagataat taaccataga tcattgaagc accctaacat tattaggttt 3720

aaagaggtga gttgggattg tgatgctgtt tctccccaga attccattgt taaacatttt 3780

tttattcttc tttgtatgtc taggttattt taacaccgac ccatcttgct attgtcatgg 3840

aatatgcctc tggcggtgag ctttttgaga ggatatgtaa gaacgtgcga ttcagtgaag 3900

atgaggtttg tgcttgccac tttgttcatc tttcttcttg acacatctct ttcttgatcc 3960

taccaaatgc acatatatat ctttccaggc tcgctacttc ttccagcagc ttatttcagg 4020

agtaagctac tgccattcaa tggtattagt tattatatag tggcatatct tccatcctat 4080

tgttactgtt tactcatacc atgcattgag ttgagctgcc tttttattct tattttcaag 4140

caagtatgcc accgtgattt gaagttggag aacacacttc tagatgggag tgacgctcct 4200

cgcttgaaga tttgcgattt cggttattcc aaggttctta cccatgtcaa cttcatcttg 4260

ttctagctta tatttatgtg aatatgtcag gccaccccgt atgtatcaga tatgttgctg 4320

agatggatat gcgtgggaca caacatggat acatctccat ggagtttcgg ggaaaaacat 4380

aaatgagagc aattctgata catgagtgaa gatgtattga ggggctttgt ggatacggtt 4440

cagtccagta ggcctactat aacccttgac tcacatgcgc tgatgtgcac cggcagtcag 4500

tgacacgaca tgcctgccgc tgccgctgcc cacctgactt cctgctcgcc cagctgcctg 4560

cctgccccca ccatgagaaa tccagcgttg tttgcacact tgagcaggct gtggctgtgg 4620

acgcacacca cccacaagat gcacgccgga aattggtgag cacttcacgg cactgcaacg 4680

agcttcacca cacaccggtg ataaatcagc accaagctgg tgatgagtta gcagtgattc 4740

cttccatgtt tcgtcttctc tcctgtagcc tcttcctgcc ccattcttcg tatcaaattt 4800

gaactagaac tgaaagggga agggattgga tcagtgattt actcatctcc tccaagcctg 4860

aagcctcacc ctcaagtaaa aagtgatgta cttagttgag tcctccctgt ttaactagtt 4920

ttactactgc actaagtgct tatatgttct cactaggtta gtacccgtgc gttgcaacgg 4980

gaacatataa taccatgata acttatatac aaaatgtgtc ttatattgtt ataagaaaat 5040

atttcataat ccatttgtga tcctagcaat acataaattt tgttatttta atttagttgt 5100

ttcactacta cattgcaacc atcagtatca tgcagacttc gatatatgtc atgatttgca 5160

tggtctcatt attggagagc acgtgccaca cctgccggta gaagttccgt cgtacatcgt 5220

tagtcatcag gcacgcacca ccatacacgc ttgcttaaac aaaaaatgca agtgtgtgtt 5280

tgcgaagaga attaaaggca ggccggcaca aaaggtaccc cgacgatggc gagtggtcat 5340

tgttgtcggt ccacctctgc tcacctccgg tgtcgagatg acgccacaat ccttgatata 5400

atagtcgtcg aacgcgcgtg atatggtgag taccgatcac tcttggctgg gctgccaaat 5460

gaagtgcacc ccgggctcat cagcgaggta gtacacctgg tcgttgcacc accggatgtg 5520

ctactcctct acatacatct tgttcgagga cactcacaca acaacaacaa tggtcatcat 5580

tccagcgcac aagaattcat ggtcggtcag tagcgactta catagcaggt taggcttcag 5640

gtggatgatg agctagacag cgtgatggcg tcgtcgtagg ttgcgatgcc cagaacaacc 5700

cgagagtcgt cgacattggc gacgaccatg aggtccccat gtttgacgat ggacagcgcg 5760

gtgcagccgc tctggaccac gtccaagcgg cggctgcgcc ggagcttgcc gtacacagcg 5820

gcgcatgggc catgtaggac tgcttccaga gttcgaactg gcagtcgcca agtttcttct 5880

cgtcgtcggt gagcgacgcc aacgcgagcg cctcgtgcta gtggtgtttg ttcggggttt 5940

ccaagtaaga aacatggatt catctttggc attggtttat gaaaatgatt tataagttag 6000

atccatggaa aaatattatg gagaataaat gttacacatg caaaaaaagt atttaagttg 6060

aaaacattat tcaaacaaaa gaaattgcat gcaaggctct tctttaaata ctactccctc 6120

aatccaaaaa tataatttaa taatctcagt gatacttatc tactactaca cattgtgcaa 6180

gggtagcagg tgggcttcgg gggagatgta ttatatgttt ttactataat agatgtagac 6240

ataaacacac atgtggtgta gtggtagcta caaacacatt tatttgagag gtcgcgggtt 6300

cgaatcccct tggagcctgt ttttttaatt taattttagc taggcgtggg atgcacgtgg 6360

gaatgggaat agactttgtg ggagggggaa tgagaaagac acgtgatagc cgggaatgga 6420

aatgtgaatg ggctttgcag ggagggggaa tgagaaagac aggcagctgg gaatgagaat 6480

gggctttgca gtgaggacgg aatggcaatg gcagaacacg gaatggggca gcctacccct 6540

taccgtctta ataggtagta gagattttga ttctgcaagt ttgactttga ggtttggatg 6600

gagttgctcg gcaagcttgg ctattggagt gttggctaaa tggaaatatg aaataatcag 6660

cagaacacaa ttgcaagaat gcaaggaaat ttcaagttcc tgctagtgtg ttacgggctt 6720

ttgaatttta atatgcctta ttatttattt attaaaaata ctaaaaacat atctgcatat 6780

tgttctttaa gaaaaatgga catttgtgta tctgcctgat actgatatgc tgaattttaa 6840

tatgccttat tatttattta ttaaaaatac taaaaacata tctgcatatt gttctttaag 6900

aaaaatggat atttgtgtat ctgcccgata ctgatatgca tatctatacc tgtgctgcat 6960

agtgttcgag tacatgtttc tgtgggtttg tggctagcac tcattgatgt tctattgttc 7020

tatggcttaa tgtttacacg atggtagtag aataatgaat atgatcatag cgttgtcagt 7080

tttcttttaa ctaacaatac catgcatttt tcattctctg cagtcatctg ttcttcattc 7140

ccagccgaag tccactgttg gaacacctgc ttatattgca cctgaagttt tgttgaaaaa 7200

agaatatgat ggcaaggtac ttgtgctatc acttttttta tcttgagaaa tgtgtcttga 7260

gagttgtgca tacctctgca tcagtgcact gttgttccat atacatgtgc cacatgttct 7320

tagtggtcag aaaacttcca cttatgattt ctcttttagt gttcactgtc tggttacctc 7380

ctatatgagg aaagcttctc ctcaccgatg gatttgttta cgacaaaaca gattgctgac 7440

gtttggtcat gtggtgtaac cctctatgta atggttgttg gcgcatatcc ttttgaagat 7500

ccagaagagc ctaagaactt ccgcaagaca attcaggtta tctttttttt tcagcatggg 7560

tacaagttga atgcacaaga gtgcacacgt acttctgtca gttatagcac atatgtgtgt 7620

gtcctagcac gcacccacct aaactctaga tcggctactt tctgagtctg catgtttaac 7680

ttaatgcatt atcctgtttt agctaacaac taaactaatg gtagtgtcat cttgttatag 7740

cgtatcttga atgttcagta cgcaattcca gacaacgtga acatatctcc agagtgcagg 7800

catctaattt cgaggatttt tgttggcgat cctgctacgg taagtccttg tatgttgtag 7860

tcttgctttg accttgatat agaaggttca gaagatatgc tgttttgagt tttgtcatat 7920

ttatctgatt ttgttgtatt catttgtcat aaattatgca gcgaataaca atccctgaaa 7980

tccggaatca tagttggttc ctgaagaacc ttcctgctga tctgatggat gatgatagca 8040

tgagcaacca atatgaggag cctgaccagc caatgcagac catggatcag atcatgcaga 8100

ttttaacaga ggccaccata ccacctgcct gttctcgcag tataaatgtc ctagctgatg 8160

gactggacat ggatgacgac atggatgatc ttgattccga ctcagatctt gatgttgaca 8220

gcagcggtga gattgtgtac gcaatgtgag taacatctca gcagccatct cagcagtcaa 8280

ccaaggaacc aatcaggcat gggatgccct gttgtatact ctggcaaagg gcatgcttcc 8340

ttttgtgaaa aacatccatg tagttttctc aaggaatcgc aatgtgttct gtggaacaca 8400

tatgttgtag tcaaactcgg gttactcagc ttgtaaaact agcctcctct tgcttcctgg 8460

ttactgggct ttggatactc tcaggcttcg acttgtgtgt tgctgttaaa tatgtataag 8520

aaacttctac tcaggttgat ctgtgaatct ggattccggt agtggttttg ctcagtgttg 8580

ggtgtattcc cgatttctgc atttcaaact ctgatgatta ggtcttgtat ttaggcggtt 8640

tctggatttc aaaatttgtg tttttttgtg caatttacat tttcggggtg ttaggcactt 8700

tcatgtttaa tttaaggggt gtttggttcc cacggagtta atgggctcta aattttacac 8760

tataaaatct atcgtgttca ttttgaacta aagttaatta agggttctaa aaatttgtga 8820

ataaatattt gaatcgtgat ctattaccac ccttattccc accttgttat tcccaccttg 8880

gcagtgagaa ggttgtcatg tctagaatag ggatggtagt ggggagccga tttctgtggg 8940

aaattcctcc attagagctt gtttgatatc cacaaaatag aggaggatca gagagctaaa 9000

atcctctctt tatttaattt taaatgagaa gtggatttta gcctctctaa cccctccggt 9060

ttttggactt tcaaactagc ccttagaaga cggggatggg gccaatttaa cccctccggt 9120

ttttggactt ctaaactatc ttccgcactc cgaattattt ctaacactaa ccccgagcat 9180

agtgcatgtt caaagtttgt aaatttcagg tttcgcatat tcaccccctc taagcgactt 9240

tcaataagca aatgcaacaa gaatacgatg gttgatgatc atgagaagga ag 9292

<210> 3

<211> 549

<212> DNA

<213> Zea mays L.

<400> 3

atggcagggc cggcgccgga tcgggccgcc ctgactgtgg ggccgggcat ggacatgcca 60

atcatgcacg acagcgaccg gtacgagctc gtgcgcgaca tcggctccgg caacttcggc 120

gttgcccgcc tcatgcgcga ccgccgcacc agcgaactcg tcgccgtcaa gtacatcgag 180

cgcggcgaga agatagatga gaatgtccag cgtgagataa ttaaccatag atcattgaag 240

caccctaaca ttattaggtt taaagaggtt attttaacac cgacccatct tgctattgtc 300

atggaatatg cctctggcgg tgagcttttt gagaggatat gtaagaacgt gcgattcagt 360

gaagatgagg ctcgctactt cttccagcag cttatttcag gagtaagcta ctgccattca 420

atgcaagtat gccaccgtga tttgaagttg gagaacacac ttctagatgg gagtgacgct 480

cctcgcttga agatttgcga tttcggttat tccaaggttc ttacccatgt caacttcatc 540

ttgttctag 549

Claims (10)

1. A protein, which is any one of:

(A1) protein with an amino acid sequence of SEQ ID No. 1;

(A2) the amino acid sequence shown in SEQ ID No.1 is substituted and/or deleted and/or added by one or more amino acid residues and is derived from the protein with the same function of corn;

(A3) a protein having 99% or more, 95% or more, 90% or more, 85% or more, or 80% or more identity to the amino acid sequence defined in any one of (A1) - (A2) and derived from maize having the same function;

(A4) a fusion protein obtained by attaching a protein tag to the N-terminus and/or C-terminus of the protein defined in any one of (A1) to (A3).

2. A nucleic acid molecule encoding the protein of claim 1.

3. The nucleic acid molecule of claim 2, wherein: the nucleic acid molecule is any one of the following:

(B1) DNA molecule shown in SEQ ID No. 2;

(B2) a DNA molecule shown in the 843-4266 position of SEQ ID No. 2;

(B3) a DNA molecule shown as SEQ ID No. 3;

(B4) a DNA molecule which hybridizes to a DNA molecule defined in any one of (B1) - (B3) under stringent conditions and which encodes the protein of claim 1;

(B5) a DNA molecule which has 99% or more, 95% or more, 90% or more, 85% or more or 80% or more identity to the DNA sequence defined in any one of (B1) to (B4) and which encodes the protein of claim 1.

4. An expression cassette, recombinant vector, recombinant bacterium or transgenic cell line comprising the nucleic acid molecule of claim 2 or 3.

5. Use of a protein according to claim 1 or a nucleic acid molecule according to claim 2 or 3 or an expression cassette, recombinant vector, recombinant bacterium or transgenic cell line according to claim 4 for modulating drought resistance in a plant.

6. Use according to claim 5, characterized in that: an increase in the expression level and/or activity of the protein according to claim 1 in the plant, thereby increasing drought resistance of the plant; and/or

The protein according to claim 1, wherein the expression level and/or activity of the protein is decreased, and the drought resistance of the plant is decreased.

7. A method for producing a plant having an increased drought resistance, which comprises the step of increasing the expression level and/or activity of the protein according to claim 1 in a recipient plant.

8. A method of breeding a transgenic plant with improved drought resistance comprising the steps of: introducing into a recipient plant a nucleic acid molecule capable of expressing the protein of claim 1 to produce a transgenic plant; the transgenic plant has increased drought resistance compared to the recipient plant.

9. The method of claim 8, wherein: a nucleic acid molecule capable of expressing the protein of claim 1 is introduced into the recipient plant by means of a recombinant expression vector.

10. Use or method according to any of claims 5-9, wherein: the plant is a monocotyledon or a dicotyledon;

further, the monocotyledon is a gramineous plant;

still further, the gramineous plant is corn.

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