CN114716522B - Application of KIN10 protein and related biological materials thereof in saline-alkali tolerance of plants - Google Patents

Abstract

The invention discloses application of KIN10 protein and related biological materials thereof in plant salt and alkali tolerance. One technical scheme to be protected by the invention is the application of protein-related biological materials in regulating and controlling plant stress tolerance, or in preparing products for improving plant stress tolerance, or in plant breeding. The protein is protein which is derived from the protein, has the same function, has the same identity with the protein by more than 80% and has the same function, and the amino acid sequence is the protein of the sequence 2 in the sequence table. Experiments prove that the KIN10 over-expression strain shows remarkable saline-alkali stress tolerance compared with the wild type control B73-329. Therefore, the function of the KIN10 protein is to positively regulate and control the stress tolerance of plants to high salt and alkali, and can be applied to plant salt and alkali tolerance breeding.

Description

Application of KIN10 protein and related biological materials thereof in saline-alkali tolerance of plants

Technical Field

The invention relates to the field of biotechnology, in particular to application of KIN10 protein and related biological materials thereof in plant saline-alkali tolerance.

Background

Previous studies have found that the KIN10 protein is involved in the autophagy process, but other functions of KIN10 are not known.

Soil salinization is an important factor restricting agricultural production. In alkaline earth environment, na ⁺ As the alkali metal element with the highest content in the nature, the alkali metal element is enriched in alkaline earth to cause salinization of soil, so that Na with high concentration in the soil is caused ⁺ And too high a pH. Along with the accumulation of salt ions in plant cells, ion toxic effects are generated on the cells, and normal functions of the cells are affected. Most of the metal elements except the alkali metal elements can form insoluble salts, so that the effectiveness of the metal elements is reduced, and the absorption of the plant to the nutrient elements is not facilitated.

Disclosure of Invention

The invention aims to solve the technical problem of improving the salt and alkali resistance of plants.

In order to solve the above technical problems, the present invention provides, first, any one of the following applications of protein-related biomaterials:

q1, the application of the biological material in regulating and controlling plant stress tolerance,

q2, the application of the biological material in preparing a product for improving plant stress tolerance,

q3, the application of the biological material in plant breeding.

The protein is the protein of A1), A2) or A3) as follows:

a1 Amino acid sequence is protein of sequence 2 in the sequence table;

a2 A protein which is obtained by substituting and/or deleting and/or adding one or more amino acid residues of the amino acid sequence shown in the sequence 2 in the sequence table, is derived from A1) and has the same function or has more than 80 percent of the same with the protein shown in A1) and has the same function;

a3 Fusion proteins obtained by ligating protein tags at the N-terminus or/and the C-terminus of A1) or A2).

The biomaterial is any one of the following B1) to B9):

b1 A nucleic acid molecule encoding said protein;

b2 An expression cassette comprising the nucleic acid molecule of B1);

b3 A recombinant vector comprising the nucleic acid molecule of B1) or a recombinant vector comprising the expression cassette of B2);

b4 A recombinant microorganism comprising the nucleic acid molecule of B1), or a recombinant microorganism comprising the expression cassette of B2), or a recombinant microorganism comprising the recombinant vector of B3);

b5 A transgenic plant cell line comprising the nucleic acid molecule of B1) or a transgenic plant cell line comprising the expression cassette of B2);

b6 A transgenic plant tissue comprising the nucleic acid molecule of B1) or a transgenic plant tissue comprising the expression cassette of B2);

b7 A transgenic plant organ comprising the nucleic acid molecule of B1) or a transgenic plant organ comprising the expression cassette of B2);

b8 A nucleic acid molecule that promotes or enhances gene expression of the above protein;

b9 An expression cassette, a recombinant vector, a recombinant microorganism or a transgenic plant cell line containing the nucleic acid molecule of B8).

In the protein, the sequence 1 in the sequence table consists of 741 amino acid residues.

The protein can be synthesized artificially or obtained by synthesizing the coding gene and then biologically expressing.

Among the above proteins, the protein tag (protein-tag) refers to a polypeptide or protein that is fusion expressed together with a target protein by using a DNA in vitro recombination technique, so as to facilitate the expression, detection, tracing and/or purification of the target protein. The protein tag may be a Flag tag, his tag, MBP tag, HA tag, myc tag, GST tag, and/or SUMO tag, etc.

In the above proteins, the identity refers to the identity of amino acid sequences. The identity of amino acid sequences can be determined using homology search sites on the internet, such as BLAST web pages of the NCBI homepage website. For example, in advanced BLAST2.1, the identity of a pair of amino acid sequences can be searched for by using blastp as a program, setting the Expect value to 10, setting all filters to OFF, using BLOSUM62 as Matrix, setting Gap existence cost, per residue gap cost and Lambda ratio to 11,1 and 0.85 (default values), respectively, and calculating, and then obtaining the value (%) of the identity.

In the above protein, the 80% or more identity may be at least 81%, 82%, 85%, 86%, 88%, 90%, 91%, 92%, 95%, 96%, 98%, 99% or 100% identity.

In the above biological material, the nucleic acid molecule of B1) is a gene encoding the protein as shown in B1), B2) or B3) below:

b1 A coding sequence of the coding chain is a cDNA molecule or a DNA molecule of a nucleotide of a sequence 1 in a sequence table;

b2 Nucleotide is cDNA molecule or DNA molecule of sequence 1 in sequence table;

b3 A cDNA molecule or a DNA molecule which hybridizes with the cDNA or DNA molecule defined in b 2) and which codes for a protein having the same function.

In the above biological material, the expression cassette containing a nucleic acid molecule of B2) refers to a DNA capable of expressing the protein of the above application in a host cell, and the DNA may include not only a promoter for promoting transcription of a gene encoding the protein but also a terminator for terminating transcription of the gene encoding the protein. 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.

Recombinant expression vectors containing the protein-encoding gene expression cassettes can be constructed using existing plant expression vectors. The plant expression vector comprises a binary agrobacterium vector, a vector which can be used for plant microprojectile bombardment and the like. Such as pAHC25, pWMB123, pBin438, pCAMBIA1302, pCAMBIA2301, pCAMBIA1301, pCAMBIA1300, pBI121, pCAMBIA1391-Xa or pCAMBIA1391-Xb (CAMBIA Co.). The plant expression vector may also comprise the 3' -untranslated region of a foreign gene, i.e., comprising a polyadenylation signal and any other DNA segments involved in mRNA processing or gene expression. The polyadenylation signal may direct the addition of polyadenylation to the 3 'end of the mRNA precursor and may function similarly to the 3' transcribed untranslated regions of Agrobacterium tumefaciens induction (Ti) plasmid genes (e.g., nopaline synthase gene Nos) and plant genes (e.g., soybean storage protein genes). When the gene of the present invention is used to construct a plant expression vector, enhancers, including translational or transcriptional enhancers, may be used, and these enhancers may be ATG initiation codon or adjacent region initiation codon, etc., but must be identical to the reading frame of the coding sequence to ensure proper translation of the entire sequence.

In the above biological material, the recombinant microorganism may specifically be yeast, bacteria, algae and fungi.

In order to solve the technical problems, the invention also provides any one of the following applications of the protein:

p1, application of the protein in regulating plant stress tolerance,

p2, the application of the protein in preparing a product for improving stress tolerance of plants,

p3, the application of the protein in plant breeding.

The modulation of plant stress tolerance described above may be an increase in plant stress tolerance.

The stress tolerance described above may be salt and alkali tolerance.

The plant described above may be any of the following:

1) Dicotyledonous plants;

2) A monocotyledonous plant, which is selected from the group consisting of,

3) The plant of the order Gramineae,

4) A plant of the Gramineae family,

5) A plant of the genus zea,

6) Corn.

In order to solve the technical problems, the invention also provides a method for cultivating saline-alkali tolerant plants, which comprises the steps of improving the activity of the protein in target plants or/and the expression level of the encoding gene of the protein to obtain the saline-alkali tolerant plants; the saline-alkali tolerance of the saline-alkali tolerant plant is higher than that of the target plant.

The improvement of the activity of the above-described protein in the target plant and/or the expression level of the above-described gene encoding the protein is achieved by introducing the above-described gene encoding the protein into the target plant.

In the method, the encoding gene of the protein can be modified as follows and then introduced into a target plant so as to achieve better expression effect:

1) Ligating to promoters expressed by various plants to facilitate expression thereof in plants; the promoter may include constitutive, inducible, chronologically regulated, developmentally regulated, chemically regulated, tissue-preferred, and tissue-specific promoters; the choice of promoter will vary with the time and space of expression requirements and will also depend on the target species; for example, a tissue or organ specific expression promoter, depending on the desired time period of development of the receptor; although many promoters derived from dicots have been demonstrated to be functional in monocots and vice versa, it is desirable to select dicot promoters for expression in dicots and monocot promoters for expression in monocots;

2) The expression efficiency of the gene of the invention can be improved by connecting with a proper transcription terminator; e.g., tml derived from CaMV, E9 derived from rbcS; any available terminator known to function in plants may be ligated to the gene of the present invention;

3) Enhancer sequences such as intron sequences (e.g., derived from Adhl and bronzel) and viral leader sequences (e.g., derived from TMV, MCMV and AMV) are introduced.

In the above method, the stress-sensitive plant may be a transgenic plant, or a plant obtained by conventional breeding techniques such as crossing.

In the above methods, the transgenic plants are understood to include not only first to second generation transgenic plants but also their progeny. For transgenic plants, the gene may be propagated in that species, and may be transferred into other varieties of the same species, including particularly commercial varieties, using conventional breeding techniques. The transgenic plants include seeds, calli, whole plants and cells.

Any of the following applications of the above method also fall within the scope of the present invention:

d1, application in regulating plant stress tolerance,

d2, the application in preparing the product for improving the stress tolerance of plants,

d3, application in plant breeding.

The regulation of plant stress tolerance may be an improvement in plant stress tolerance.

The stress tolerance may be salt and alkali tolerance.

The plant described above and/or the plant of interest is any one of the following:

1) Dicotyledonous plants;

2) A monocotyledonous plant, which is selected from the group consisting of,

3) The plant of the order Gramineae,

4) A plant of the Gramineae family,

5) A plant of the genus zea,

6) Corn.

The biological material described above or the protein described above also fall within the scope of the present invention.

Experiments of the invention prove that the KIN10 over-expression strain shows remarkable salt and alkali stress tolerance compared with the wild type control B73-329. Therefore, the KIN10 protein has the function of improving the stress tolerance of plants to high salt and alkali, and can be applied to plant salt and alkali tolerance breeding.

Drawings

FIG. 1 shows that wild-type maize B73-329 and KIN10 overexpressing strains are in NaHCO-containing form ₃ Phenotype in treated soil.

Detailed Description

The following detailed description of the invention is provided in connection with the accompanying drawings that are presented to illustrate the invention and not to limit the scope thereof. The examples provided below are intended as guidelines for further modifications by one of ordinary skill in the art and are not to be construed as limiting the invention in any way.

The experimental methods in the following examples, unless otherwise specified, are conventional methods, and are carried out according to techniques or conditions described in the literature in the field or according to the product specifications. Materials, reagents and the like used in the examples described below are commercially available unless otherwise specified.

In the following examples, 5 or more repeated tests were used for quantitative tests, unless otherwise specified.

Corn B73-329 in the examples below was obtained from the national agricultural university crop functional genome and molecular breeding research center.

Example 1 application of KIN10 Gene in regulating salt-tolerant alkalinity of plants

1. Acquisition of maize KIN10 overexpression lines construction of 1.1 maize KIN10 overexpression vectors

CDS sequence of cDNA gene of maize KIN10 gene is shown as sequence 1 in sequence table, amino acid sequence of coded protein is shown as sequence 2 in sequence table, nucleotide sequence of genome gene is shown as sequence 3 in sequence table.

The pBCXUN vector is an expression vector obtained by replacing the HYG gene (hptII, hygromycin resistance gene) of pCXUN (GenBank: FJ905215.1, 06-JUL-2009) with the Bar gene (encoding phosphinothricin acetyltransferase) (the nucleotide is sequence 4 in the sequence table) and keeping the other nucleotides of pCXUN unchanged.

The DNA molecule with the nucleotide sequence of sequence 1 in the sequence table is inserted into a pBCXUN vector to obtain an expression recombinant vector of the KIN10 gene, which is named pBCXUN-KIN10. In pBCXUN-KIN10, the DNA molecule shown in the sequence 1 in the sequence table is started by the promoter of the maize ubiquitin gene Ubi and is stopped by a Nos terminator, so that the KIN10 protein can be expressed (the amino acid sequence is shown as the sequence 2 in the sequence table).

1.2 obtaining of pBCXUN-KIN10/EHA105 recombinant Agrobacterium

The pBCXUN-KIN10 constructed in 1.1 is transformed into competent agrobacterium EHA105 strain by a heat shock method, positive clones are identified by colony PCR, sequencing of the positive clones is selected, and primers for colony PCR and sequencing identification are as follows:

f (corresponding to the Ubi promoter) 5'-TTTTAGCCCTGCCTTCATACGC-3',

r (corresponds to the terminator of the Nos): 5'-AGACCGGCAACAGGATTCAATC-3'.

The positive clone containing pBCXUN-KIN10 is recombinant Agrobacterium and is named pBCXUN-KIN10/EHA105.

Acquisition of 1.3KIN10 overexpressing lines

1.3.1 activation of recombinant Agrobacterium pBCXUN-KIN10/EHA105

The recombinant Agrobacterium pBCXUN-KIN10/EHA105 obtained in 1.2 was inoculated into 2-3mL of a liquid medium containing 100. Mu.g/mL kanamycin and 50. Mu.g/mL rifampicin, and cultured overnight at 28℃with shaking for activation.

1.3.2 shaking of recombinant Agrobacterium pBCXUN-KIN10/EHA105

Recombinant Agrobacterium pBCXUN-KIN10/EHA105 activated in 1.3.1 was transferred to a large volume of liquid medium containing 100. Mu.g/mL kanamycin and 50. Mu.g/mL rifampicin and shake cultured to OD of the bacterial solution ₆₀₀ Recombinant Agrobacterium suspensions were obtained at a time between 0.8 and 1.0.

1.3.3 transformation of maize immature embryo by recombinant Agrobacterium

Infecting B73-329 maize young embryo scraped under aseptic condition with the recombinant agrobacterium suspension obtained in 1.3.2, and screening the induced callus by using herbicide glufosinate. After the callus is differentiated into seedlings, extracting genome DNA of transgenic plant leaves, and adopting the primer pair PCR in the step 1.2 to identify and screen, finally obtaining the KIN10 transgenic over-expression positive plants. And (3) carrying out subsequent experiments on the KIN10 transgenic overexpression positive plant to obtain a KIN10 gene overexpression OX strain homozygous for the T3 generation after selfing propagation.

2. Application of KIN10 gene in plant salt and alkali tolerance

35 seeds of each of the T3 generation 4 independent KIN10 gene overexpressing strains OX-1, OX-2, OX-3 and OX-4 obtained in wild-type control B73-329 and step 1.3.3 were planted in small pure vermiculite pots without nutrient soil (4 seeds were planted in each pot), and the culture room (temperature 25 ℃ C., humidity 40%) was grown normally and irrigated with 1/2Hoagland's nutrient solution. After 10 days of corn growth (trefoil stage) the corn was irrigated with 100mM NaHCO ₃ (saline-alkaline stress) 1/2Hoagland's nutrient solution (NaHCO was added to 1/2Hoagland's nutrient solution ₃ To NaHCO ₃ Liquid obtained at 100mM content), once every 10 days, two liters per dish, and the phenotype of maize growth and leaf yellowing degree under saline-alkali stress conditions was observed for 40 days of growth.

The phenotypic results are shown in FIG. 1, at 100mM NaHCO ₃ Under the treatment conditions, 4 independent KIN10 overexpressing strains OX-1, OX-2, OX-3 and OX-4 showed stronger salt tolerance than wild-type control B73-329, greener leaves and low yellowing; the strain height is higher, the growth vigor is better, and the strain height of the KIN10 over-expression strain is 1.2-1.4 times of the wild type control B73-329 strain height. Therefore, KIN10 positively regulates the salt and alkali resistance process of corn, and can be applied to salt and alkali resistance breeding of plants.

The present invention is described in detail above. It will be apparent to those skilled in the art that the present 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 respect to specific embodiments, it will be appreciated that the invention may 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 application of some of the basic features may be done in accordance with the scope of the claims that follow.

Sequence listing

<110> Chinese university of agriculture

<120> KIN10 protein and application of related biological material thereof in saline-alkali tolerance of plants

<130> GNCSQ203345

<160> 3

<170> PatentIn version 3.5

<210> 1

<211> 2226

<212> DNA

<213> Artificial sequence (Artificial Sequence)

<400> 1

atggagatgg tggatgacga cggtggtcct gttcgtggag ccggcgcaac agtccagccc 60

gaggggcgcg tccgcggcgt cgctgctggc cgagccgtcg tacgacggct cgccgctccg 120

ccgccaggcg ttccagagct acctcaccaa gagcggcgtc atgtcgtcgt ccccgaccag 180

cacgctggtc tcgcccccga ggtcgccgcc gtcggagtcc ccgccaatgt cgccggacgc 240

cgccgccgcg gcgcgtggcc gggtgtcggc ggctcgcccg tcaacgaggt cctcgcctcg 300

ctgcgccagc tccgcctcgg cggcggctcg ccgaggtcgg cgccttccgg cgggtcgttc 360

ttggcaggcg gctacccgtt cgggtccctg aagtcgccgg ccggtctgta cagtctcccg 420

tccacaccac caacccggtc gtgcccagtg atggtgacca ccgcctccgg cgccaccatc 480

atgaccgtgg aacgcctcaa cctcgggctc ctcgtggacg aggagccggt catggagcgg 540

gtcgagtccg ggagagccct ccgcgagaag gtgttcgaac ggctcagcaa agaaagcgcc 600

gtgcccaacg acaccgtcgc ctccgccaac gtagaggcgc actccgcctc tcctctctcc 660

gtcgctgggc agctgggctt ctctcccgac tcccgagcag ccgagctccc cctgctcggc 720

ctcgcctccg ccgcgaaccg cgcctccggc tccgcctgct cgggcgggcg gcacccagct 780

ccctgcatct tcgctgcggc gaggggttgg ggatttgggg tcggggcgct tcagcttcag 840

cttcagcttc gttttctaca gcccaaggcc aaccggctct tgttttccct ttcccctcaa 900

attagtatga tatgctttgg ttttgattca gggtacacga tggagggagc tggaagagat 960

gccaaccctt tgggcggtta ccgaattggc aaaaccctag gaatcgggtc gttcggtaaa 1020

gtgaagatcg cggaacatat attgactggc cataaggtgg caatcaagat cctcaatcgc 1080

cgtaagatca gaagcatgga aatggaagag aaagtgaaga gagaaatcaa gatactgaga 1140

ttatttatgc atcctcatat catacgcctt tatgaggtga tagatacacc tgctgatatt 1200

tatgttgtta tggagtatgt taaatctgga gagttgtttg attacattgt tgagaaggga 1260

agactacatg aagaagaagc ccggcgcttt tttcagcaga tcatatctgg tgttgaatat 1320

tgccatagga acatggttgc tcaccgtgat ttaaagccag agaatcttct tttggattcg 1380

aaatgcaaca ttaagattgc tgattttggc ttaagtaatg ttatgcgtga tggtcacttt 1440

cttaagacga gttgtggtag cccgaattat gcagcacctg aggtcatatc tggtaaacta 1500

tatgctggtc ctgaagttga tgtctggagc tgcggcgtta ttctttatgc tcttctttgt 1560

ggcactcttc catttgacga tgagaatatt ccaaaccttt ttaagaaaat aaagggtgga 1620

atatataccc ttcctagtca tttgtcacct tcagcgaggg acttgattcc tagaatgctg 1680

gttgttgatc cgatgaaaag gattacaata cgtgaaatcc gtgaacatat gtggttcaaa 1740

attcaacttc cgcgctattt ggctgtaccg cctccagaca ctgcccaaca agttaagaag 1800

ctggacgagg aaacccttaa tgatgttatt aagatgggtt ttgacaagaa tcagctaatt 1860

gaatctctgc acaacagatt gcagaatgag gcaacagttg cctattattt actcatggac 1920

aataggcttc gtacaaccag tggttatctt ggatctgagt ttcaagaatc tatggaccca 1980

tctttctctc aagtacatgc tgaaacacca acttcagcaa ctgaacatcg gcagcatgtg 2040

tttacagaat ctccggggtc tggtttgagg cagcattttg catctgaaag gaaatgggcc 2100

cttggtcttc agctttacaa aacgagggat gagaagtacc ttcttgacct gcaaagggtc 2160

agtggaccac aactcctctt tctggacttg tgttccgcct ttctaacgca gctgagagtt 2220

ctttga 2226

<210> 2

<211> 741

<212> PRT

<213> corn (Zea mays)

<400> 2

Met Glu Met Val Asp Asp Asp Gly Gly Pro Val Arg Gly Ala Gly Ala

1 5 10 15

Thr Val Gln Pro Glu Gly Arg Val Arg Gly Val Ala Ala Gly Arg Ala

20 25 30

Val Val Arg Arg Leu Ala Ala Pro Pro Pro Gly Val Pro Glu Leu Pro

35 40 45

His Gln Glu Arg Arg His Val Val Val Pro Asp Gln His Ala Gly Leu

50 55 60

Ala Pro Glu Val Ala Ala Val Gly Val Pro Ala Asn Val Ala Gly Arg

65 70 75 80

Arg Arg Arg Gly Ala Trp Pro Gly Val Gly Gly Ser Pro Val Asn Glu

85 90 95

Val Leu Ala Ser Leu Arg Gln Leu Arg Leu Gly Gly Gly Ser Pro Arg

100 105 110

Ser Ala Pro Ser Gly Gly Ser Phe Leu Ala Gly Gly Tyr Pro Phe Gly

115 120 125

Ser Leu Lys Ser Pro Ala Gly Leu Tyr Ser Leu Pro Ser Thr Pro Pro

130 135 140

Thr Arg Ser Cys Pro Val Met Val Thr Thr Ala Ser Gly Ala Thr Ile

145 150 155 160

Met Thr Val Glu Arg Leu Asn Leu Gly Leu Leu Val Asp Glu Glu Pro

165 170 175

Val Met Glu Arg Val Glu Ser Gly Arg Ala Leu Arg Glu Lys Val Phe

180 185 190

Glu Arg Leu Ser Lys Glu Ser Ala Val Pro Asn Asp Thr Val Ala Ser

195 200 205

Ala Asn Val Glu Ala His Ser Ala Ser Pro Leu Ser Val Ala Gly Gln

210 215 220

Leu Gly Phe Ser Pro Asp Ser Arg Ala Ala Glu Leu Pro Leu Leu Gly

225 230 235 240

Leu Ala Ser Ala Ala Asn Arg Ala Ser Gly Ser Ala Cys Ser Gly Gly

245 250 255

Arg His Pro Ala Pro Cys Ile Phe Ala Ala Ala Arg Gly Trp Gly Phe

260 265 270

Gly Val Gly Ala Leu Gln Leu Gln Leu Gln Leu Arg Phe Leu Gln Pro

275 280 285

Lys Ala Asn Arg Leu Leu Phe Ser Leu Ser Pro Gln Ile Ser Met Ile

290 295 300

Cys Phe Gly Phe Asp Ser Gly Tyr Thr Met Glu Gly Ala Gly Arg Asp

305 310 315 320

Ala Asn Pro Leu Gly Gly Tyr Arg Ile Gly Lys Thr Leu Gly Ile Gly

325 330 335

Ser Phe Gly Lys Val Lys Ile Ala Glu His Ile Leu Thr Gly His Lys

340 345 350

Val Ala Ile Lys Ile Leu Asn Arg Arg Lys Ile Arg Ser Met Glu Met

355 360 365

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

370 375 380

Pro His Ile Ile Arg Leu Tyr Glu Val Ile Asp Thr Pro Ala Asp Ile

385 390 395 400

Tyr Val Val Met Glu Tyr Val Lys Ser Gly Glu Leu Phe Asp Tyr Ile

405 410 415

Val Glu Lys Gly Arg Leu His Glu Glu Glu Ala Arg Arg Phe Phe Gln

420 425 430

Gln Ile Ile Ser Gly Val Glu Tyr Cys His Arg Asn Met Val Ala His

435 440 445

Arg Asp Leu Lys Pro Glu Asn Leu Leu Leu Asp Ser Lys Cys Asn Ile

450 455 460

Lys Ile Ala Asp Phe Gly Leu Ser Asn Val Met Arg Asp Gly His Phe

465 470 475 480

Leu Lys Thr Ser Cys Gly Ser Pro Asn Tyr Ala Ala Pro Glu Val Ile

485 490 495

Ser Gly Lys Leu Tyr Ala Gly Pro Glu Val Asp Val Trp Ser Cys Gly

500 505 510

Val Ile Leu Tyr Ala Leu Leu Cys Gly Thr Leu Pro Phe Asp Asp Glu

515 520 525

Asn Ile Pro Asn Leu Phe Lys Lys Ile Lys Gly Gly Ile Tyr Thr Leu

530 535 540

Pro Ser His Leu Ser Pro Ser Ala Arg Asp Leu Ile Pro Arg Met Leu

545 550 555 560

Val Val Asp Pro Met Lys Arg Ile Thr Ile Arg Glu Ile Arg Glu His

565 570 575

Met Trp Phe Lys Ile Gln Leu Pro Arg Tyr Leu Ala Val Pro Pro Pro

580 585 590

Asp Thr Ala Gln Gln Val Lys Lys Leu Asp Glu Glu Thr Leu Asn Asp

595 600 605

Val Ile Lys Met Gly Phe Asp Lys Asn Gln Leu Ile Glu Ser Leu His

610 615 620

Asn Arg Leu Gln Asn Glu Ala Thr Val Ala Tyr Tyr Leu Leu Met Asp

625 630 635 640

Asn Arg Leu Arg Thr Thr Ser Gly Tyr Leu Gly Ser Glu Phe Gln Glu

645 650 655

Ser Met Asp Pro Ser Phe Ser Gln Val His Ala Glu Thr Pro Thr Ser

660 665 670

Ala Thr Glu His Arg Gln His Val Phe Thr Glu Ser Pro Gly Ser Gly

675 680 685

Leu Arg Gln His Phe Ala Ser Glu Arg Lys Trp Ala Leu Gly Leu Gln

690 695 700

Leu Tyr Lys Thr Arg Asp Glu Lys Tyr Leu Leu Asp Leu Gln Arg Val

705 710 715 720

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

725 730 735

Gln Leu Arg Val Leu

740

<210> 3

<211> 22738

<212> DNA

<213> corn (Zea mays)

<400> 3

gcccctgcgc cccaaatctc gccgacgtgc tcgaccacct caagccccac actatcgctc 60

tcctccatct tctctacagc cgctgcccac ccctagctcc gcctcctctc tatgcgaaat 120

gggcgcgtgc gcactgcggc cctagcgaag gcgttccaga tccgcaggag caggcgacgc 180

ttgcctacct ctgcgccccg ttcgaatccg atagcatcat cgatctgttc gtacggacga 240

tacctcaatg gagatggtgg atgacgacgg tggtcctgtt cgtggagccg gtatgcgacc 300

gcgttcgaag tccaaggaag agaaggggag gctcgggcgg tcctgttcgt aacatctgac 360

aatctccatg gcggctgttt aggtcctgcg cctgcgaccg cgtgcgagct ggcgcacggg 420

gtgttcgagt gctggctcca cccgttgcgc taacggatcg cagccctgca aggacggcac 480

cgggtgccgc cgccgcgtct gcttcttcgc gcacacgccg gaccagcttc gcgtgccaca 540

ggcgcaacag tccagcccga ggggcgcgtc cgcggcgtcg ctgctggccg agccgtcgta 600

cgacggctcg ccgctccgcc gccaggcgtt ccagagctac ctcaccaaga gcggcgtcat 660

gtcgtcgtcc ccgaccagca cgctggtctc gcccccgagg tcgccgccgt cggagtcccc 720

gccaatgtcg ccggacgccg ccgccgcggc gcgtggccgg gtgtcggcgg ctcgcccgtc 780

aacgaggtcc tcgcctcgct gcgccagctc cgcctcggcg gcggctcgcc gaggtcggcg 840

ccttccggcg ggtcgttctt ggcaggcggc tacccgttcg ggtccctgaa gtcgccggcc 900

ggtctgtaca gtctcccgtc cacaccacca acccggtcgt gcccagtgat ggtgaccacc 960

gcctccggcg ccaccatcat gaccgtggaa cgcctcaacc tcgggctcct cgtggacgag 1020

gagccggtca tggagcgggt cgagtccggg agagccctcc gcgagaaggt gttcgaacgg 1080

ctcagcaaag aaagcgccgt gcccaacgac accgtcgcct ccgccaacgt agaggtagcg 1140

gctccgaacg tcagttgggt ctccgacctc atcaactgag ggaatgaaac tcctgaaact 1200

atcagcatca gccgtgcctc ctctgaacgg tgcttatcca tggtaagtgc gtgagggaag 1260

cttagaggcg tcactgcaca gcagcaggag aagaaagttc gtgggaaaaa ccaaaaaaaa 1320

gatttttatt tttctagatt tgtactaaga tgatccttgt acatattttc ctccatgtgt 1380

gaatattctc ggatggctgt tgcctgttgg agggagggga ggcagggaaa ggttgatttg 1440

ttctggggtt tcagatgatg gcatgccagt ggaggtagaa ggggtggcca gtaaaacaca 1500

aaaggatcaa aggagacatg gtaggcgacc agaccatgca tgaggtgtct agatgtacat 1560

agcgtctgtg cgtacctgtg tgttgtgtgt gtgtggaggt agagcgtggg agaggcgacc 1620

atgtcggcga tccaagacag gagtaaagat gacaggtttt tttatatatc tggcgccttg 1680

tcatagcctt tgttttataa ttttttaaca actctaaggt aaatatatcg aacatagtga 1740

tgttcagtct tgctcgatgc tgtgctaaag gttcaaaaat cagagctgaa agctgattcg 1800

aatgcctgaa cactgtgtag gggacttgct ccttggtgac gacatcttca acaccgacgg 1860

ggagctgtgg aggaagtaga ggaagacgac gtttttcaag ttcacctcta agaccctgag 1920

gtatttcaac gccaatatgt tcagagtact tagatcaaca caccatgttt ttttgttgca 1980

tgccatttgt agtagttcag gaactcagat cagcacacca tgttcttctg ttgcatgcca 2040

tgtgagtagt tcagatactc agatcagcaa taggatgact tttagcagtt caaagtcatt 2100

ttgttttgtt gcatgccatg tgtagttcta catatgtgag tagttgacgg ccacaacaaa 2160

tttacaaagt cacctccatg ggtgaattag gaggtgaatg tgcttgcaat acatgcgggt 2220

gtaatctagc gttttagcgg attccaggtt tccatggagt aagattcctc tatgcgtttt 2280

attcgatttc tttgtatatt gttctttcta tttttttaat tcagtaacaa gatcacaaac 2340

tgatttccca atttgcaggt ccaattcgcg tgtggcatag gaatcccatc ctagattcaa 2400

tagaggggct gcactgccag cattgccata gctgaaatcg agcgttgacg gacaccgggc 2460

gccgagatgg gccattctgt ccactgccgc aagaacctca tgcggtggag ccatccttac 2520

acctgctcct acttccacgc tagcagccca acacgccgac gacacgagct cctccctata 2580

aaagcaacat ctttactgta tagataaaga atggttataa ttatttattc ttgtgtccaa 2640

gtggtattgg atttttttct agcttggggt taaagaaaag acagtcatgt actttgcata 2700

ttttttattt atattttatg ctaaattgtg gagtaacact atattagttg ttagcaaaga 2760

attttttctt tgtgagatat taaggttgat gataatatat attgttattt catgtaatca 2820

ttgtgtatta atattttatt gaataattgg tgtaccgtcg caacgcacgg gctctatact 2880

agtttcttta taaatttaag aagaaatttg ccaaatagat aatttttata gcagcttatc 2940

agtttagctg tttctcaaaa tacaataaaa acagtcaaca agccgaagcg gtttttcact 3000

gcatcacgca ccaaaacagg gcctaaggct gtctccagca acgtcctcta tattcatcct 3060

ctatatccgt cctttacagt cttctctaaa agattccacc ctctatatct ccttcctctc 3120

taacaacgtt ctctaaatca cgtcctctat actcaaatat ctatattaga aacatttttt 3180

atttttattt tttgtacata cgtatttgtc atactctcaa atgtattgta catattttag 3240

ttttgctaaa ccggttattt aaagtattga aatggataga agatcgttta gaggaactct 3300

atatatagag aatccagcag cgtcctctaa atttagagaa ccgtttagag gacgcttctg 3360

gagggcgtag aggaccattc gatcctctat atttagggta cagaaccatt taaggtctct 3420

tgttggagct agtctaatca ggagcccgat tacgccgtcg cggtggggtc ccacgtgagc 3480

gtcgcctttc tttcctcccc atttttctct ctctggcgtc actcatgtgt cagactgtca 3540

ggcgcactcc gcctctcctc tctccgtcgc tgggcagctg ggcttctctc ccgactcccg 3600

agcagccgag ctccccctgc tcggcctcgc ctccgccgcg aaccgcgcct ccggctccgc 3660

ctgctcgggc gggcggcacc cagctccctg catcttcgct gcggcgaggt acgccccccc 3720

cccccccccc ccccccatcg cctcatccct ccgcgcctgc ctctgcgtct ctgcctgtct 3780

ctttgtcctc ccagcgtacc aattgggatg cgcgcatgtc tgggtctttg cgctgtgaag 3840

aagctgcggt ctgtggctcg tgtcccgttg ctagcggaga gggatttcgt ggctccggct 3900

agctccgcca gatccgaggc ctcgcgtcgt gggatagcgc gcttagcggt tcggctgtgc 3960

aggggttggg gatttggggt cggggcgctt cagcttcagc ttcagcttcg ttttctacag 4020

cccaaggcca accgggtgcg gatagattct gaaagatagc ttgaggattt tggtaacggc 4080

attctggttc gcaactcccg cctctccgcg aagtagattg aggcgtgctg ttttggccac 4140

atgctgcctc atgccgctca tccctatgtg atcagaaaca tggttgcggg ctgtagtgtt 4200

agtttgctgt tttgtttttt ttaatctgtc ttatcttgta attttgttgc tggagtaatg 4260

aagttcggtt ttaccgttgt ggaaaaaaaa tgtttgaggt gtgttgtgtc tttgcttagc 4320

ctgaatcggt attacttgtc tcggtcgtct aggtccagcc ttcgaattgt atcagagcgc 4380

gcatagtttt aggtctgctt gcacaatagt cgttacctgg ttttagctct tgttttccct 4440

ttcccctcaa attagtatga tatgctttgg ttttgattca gggtacacga tggagggagc 4500

tggaagagat gccaaccctt tgggcggtta ccgaattggc aaaaccctag gaatcgggtc 4560

gttcggtaaa gtgaagatcg cggaacatat attgactggc cataaggtgg caatcaagat 4620

cctcaatcgc cgtaagatca gaagcatgga aatggaagag aaaggtcagt gcttattcct 4680

tccgtacaga acattggctt gtggcgctta agcttggcat gtgatgtatg gattatgtac 4740

cttatgtaca cgcagcagag ctccttatta tgtcttgaca attttttatc tgaagtactc 4800

aacttaattc tctcttacta tgtacctgca gtgaagagag aaatcaagat actgagatta 4860

tttatgcatc ctcatatcat acgcctttat gaggtgatag atacacctgc tgatatttat 4920

gttgttatgg agtatgttaa atctggagag ttgtttgatt acattgttga gaagggaaga 4980

ctacatgaag aagaagcccg gcgctttttt cagcaggtgg ctttcttgca tattgttcat 5040

tccaaagatt gcacaatgca ttgcaatcac tgttttgaag aactgtagta tggtactatg 5100

gttggcataa ccttttttgt ggctgataaa taattgaaat ctgttaaccg tcatgcttct 5160

gtcatatata tatctgtgtc agtaaattga ttgtttgtgt ttatttggtt cattgagttt 5220

agtttgataa ttttcttctt attacttttt atgcagatca tatctggtgt tgaatattgc 5280

cataggaaca tggttgctca ccgtgattta aagccagaga atcttctttt ggattcgaaa 5340

tgcaacatta agattgctga ttttggctta agtaatgtta tgcgtgatgg tcactttctt 5400

aagacgagtt gtggtagccc gaattatgca gcacctgagg tatacatatg acattattta 5460

caattagcat tcttcatggt cacatttaca tatataactt gctatgacat gattcttgat 5520

ttcttgttta ttttcttagg tcatatctgg taaactatat gctggtcctg aagttgatgt 5580

ctggagctgc ggcgttattc tttatgctct tctttgtggc actcttccat ttgacgatga 5640

gaatattcca aaccttttta agaaaataaa ggtgagttat catctccttg atttatcgag 5700

catcttctaa taaaaatgat gaggatctct atgaaccaat ttggtttcat gtccaaaaga 5760

ttaaccatgg aagctatttt cttaataaga caagttatgg agtagtatag agagcagaag 5820

aagaacctac acatggtttt cattgagttg gagaagattt ttgacaaaat accaataaat 5880

gttgtgcggc gggctttgaa caaacataaa gttccaacaa aatacattag agccattaag 5940

tacatgtaca actatattgt ggctagtgtt agaacaagtg atggagacac gaatgactgg 6000

ccgatgagaa taggactaca ccaagggtta gccttgagcc cttacctttt tgtcttggtg 6060

atggataaaa tcacaagcga catataaggg gatatatgct tttgcgaata atgttgtact 6120

agttgatgaa tggcaggcat gattaaatag gaaacttgag ttgtggttga agactataga 6180

gtgcaggctc aatagaacta aaaccgaata tatgagatat gactttgaca ctacatatga 6240

gaaagaagat attagtttgg aaggtcaagt agtgcccagg aaggacactt ttcggtattt 6300

gtgatcaata ctatagaaag acggggatat cgatgaagat gttagtcata gaataaaaac 6360

agggtggatc aagtgacatc aaccagcgtt ctatgtgaca agaagggtag cacataagct 6420

aaaaggagag ttttatagga tgacgattat acctgctatg ttgtatggtg cagaatgttg 6480

gctaataaaa gacgatatgt tcaacagtta agtagcggaa atgcatatgt tgtgttgaat 6540

ttatggcata aaaaaaggat caagtccaga atgaaggtat atgtgataag ctaggggtag 6600

cacaaattta agaaaagctt gtctaacacc ggttgagatg gtttagacat gtccaacgga 6660

gacatccaaa aggcggttta aaaggaaaga ctgaaattgg catgggaaga ggcagtaaaa 6720

ggtgatttaa aaggatgaaa tgtacataaa gatttagcct tgaatacgag tgcatggaaa 6780

acggctatcg acatgcctga accttgaatt gtgtcttttg ttaggtttta actatagcct 6840

accccaactt gctagggatt aaaaggcttt cttgttgctg ctgctgctgt atcatctcat 6900

tgatttacct gaaaaaaaga ttcctttata ttttactctt aatatgctcg ttaaattctt 6960

atatctaatt gtagggtgga atatataccc ttcctagtca tttgtcacct tcagcgaggg 7020

acttgattcc tagaatgctg gttgttgatc cgatgaaaag gattacaata cgtgaaatcc 7080

gtgaacatat gtggttcaaa attcaacttc cgcgctattt ggctgtaccg cctccagaca 7140

ctgcccaaca agttaagaag gtataatgat attgtgtgta tggatggcgc tattctgtaa 7200

aattcttatt ccacgttttc tacttcagaa aatgaattga cattgttttg aattttgtag 7260

ctggacgagg aaacccttaa tgatgttatt aagatgggtt ttgacaagaa tcagctaatt 7320

gaatctctgc acaacagatt gcagaatgag gtatctgctt cactctcagt ttcctttggt 7380

cgttttatgg aatcatgatg gtttggctgt gcaggcaaca gttgcctatt atttactcat 7440

ggacaatagg cttcgtacaa ccagtggtta tcttggatct gagtttcaag aatctatggt 7500

gagtggcttt cttatctctt gcatttccaa atgttggttg agttattcag tacttgtgat 7560

gctataaatt tcttgctaat tttctgtttg cgttttgtga tctctaggac ccatctttct 7620

ctcaagtaca tgctgaaaca ccaacttcag caactgaaca tcggcagcat gtgtttacag 7680

aatctccggg gtctggtttg aggcagcatt ttgcatctga aaggaaatgg gcccttggtc 7740

ttcaggtaat tttcatgtta agtacatcgt atgatgtctt tcctgcatta ttatgattca 7800

attatctgta attgtctaac agtctcgagc acatccacga gaaataataa gtgaagtgct 7860

taaagcactg caagaactga atgtttactg gaaaaagatt ggacactaca acatgaaatg 7920

cagatggagt cctggctgcc ttgagagtat gatgcataac tgtgatggct ttagtgcaga 7980

gtctgctata attgaaactg atgatctcat agcgaaatca acccgtatag tgaagtttga 8040

gattcaggta tcatttggaa tcttaatcat gcacgcttaa ataaatgtta tgtaacctag 8100

tttttgaaag ggaaatgtgc ctttgggcca tttctaagta ttttggtgat taagtgccaa 8160

cacaagtgct taaatgtaaa tcaatgccca tggatgaaca aagtgcaaat cttcaacaaa 8220

ggtatgtttc taagtcttag tacattggtt ttgtgtacta atatatttgt ctaagtgtta 8280

gaaacagaaa gaagaagaga agagaagact tggctgtgtg cagccaaggg gctgcttcgg 8340

tctagggcac cggactgtcc ggtggtgcac cggacagtgt ccgatggtgc accggacagt 8400

gtccggtgcg ccaggccgcc ttggccgaag aggccgctct cgggtttttc tccggcgact 8460

tcggctaaaa ttcaccggac tgtccggtgt gcaccggact gtccggtgag ccaacggtcg 8520

gccgggccaa cggtcggccg cgtgatcggc gcgcgacacg tggccgagcc aacggtcgga 8580

gggaagcacc ggactgtccg gtgtgcaccg gactggccgg tgcgccagat ctgcaacggt 8640

cggcaacggt cggcttcgcc aattaaggaa aggaatcggg caccggacac tgtccggtgt 8700

gcaccggact gtccggtgcg cccgacgaca gaaggcaaga atagccttcc agatgtgctc 8760

tcaacggctc ctaggtgcct tggggctata aaagggaccc cttggcgcat ggaggagtac 8820

aaccaagcat tcctacaacc tttctaagca ccaagacatt gatctagcgc attcgattca 8880

ttgtgatagc atatagagct cttgtggagt tgtgtactct ttgagttgtg ttgcgagctc 8940

ttattgctgc ttgtgtgcgt gttgctctga tcttttgaag tcttgtgtgc gtcgctcatt 9000

ccctcccctg ctccgtgatt ctctgtgaac atcttttgta agggcgagag gctccaaatt 9060

gtggagattc ctcgcaagcg ggattgagaa aagaaaagca agaacaccgt ggtattcaag 9120

ttgatcattg gatcacttga gaggagttga gtgcaactct cgtccgttgg gacgccacaa 9180

cgtggaagta ggcaagtttt gtacttggcc gaaccacggg ataaccaccg tgtcatctct 9240

gtgattgatt tcttgtggtt attgtgtttt gactcctctc tagccacttg gccataattg 9300

tgctaacact taacaagttt ttgtggctta agttttgaag ttattcagga tcacctattc 9360

acccccccct ctaggtgctc tcaattggta tcagagccgt tctcttcaag aaagggacta 9420

accgcccgaa aagatggatc ctaaggggaa gggaattgtg atcaacgaca aggagaagga 9480

gtccttcgtc aacgagccaa gggatgacaa gtccaacgac tcgggctcgg gccacaggcg 9540

caaagatggg aagaagaaga agacaaggcg catcaaggag atcatctact acgacgatag 9600

tgatgaatct acttcttccc aaaaggatgg tgacaacgac tacaaaaaga cggtcaattc 9660

gaacttttca tttgattatt ctcgtattca acatagttcg aattcgcatt tgctttccat 9720

tcctcttggc aaacccccac actttgatgg ggaggactac ggattttgga gtcacaaaat 9780

gcgtagtcac ttattctctc tccatccaag catatgggag attgtggaga atggaatgaa 9840

atttgatagc tcggatagtc ctatgcttat aaatgaacaa attcatagaa atgcacaagc 9900

tactactgtt ctcttagcat ctttgtgcag ggaagaatac aataaggtga gcggcttgga 9960

caatgccaag cagatatggg acaccctcaa gatctctcac gaggggaacg acatcaccat 10020

gctcaccaag atggagttgg tggagggcga gcttggacga ttcgccatga taaggggaga 10080

ggagccaact caaacttaca accggctcaa gacccttatc aacaaaataa ggagctacgg 10140

aagcacgcgt tggacggacc acgacgtcgt ccgcctacta ctcaggtcct ttaccgttct 10200

tgatcctcat ctcgtgaaca atattcgtga gaattccagg tacacgatga tgacgcccga 10260

ggaagtactt ggaaaattcg tgagcgggcg aatgatgatc aaggaggcaa ggtacgtcga 10320

tgacgcgttg aacggtccaa tccacgagcc tcttgctctc aaggcatcga gaagcaagga 10380

ggcactacca agcaaggtgg cgcaaattga ggcggccggg cttaatgatg aagaaatggc 10440

tctcatcatt aagcgcttca agacggcgtt aaggggtcgc aaggagcatc ccaacaagac 10500

caagacgaag gggaagcgct catgcttcaa gtgtggtaag attggtcact ttatcgctaa 10560

ttgtcccgat aatgatagtg accaggaaaa gaagagtgga aaatgggaaa agaagaagaa 10620

ttacaagaag gcaaagggcg aggctcatat tggaaaagag tgggattcgg attgctcctc 10680

gtccgactcc gacaacgaag gactcgccgc caccgccttc aacaagtcat ccctctttcc 10740

caacgaacat cacacttgcc tcatggcaag ggaaaagaag gtaatcactc gtaatgctaa 10800

tacttatgat tcttctagtg atgatgaatc tagtgaggat gaagtagatt attcatgttt 10860

attcaagggc ttagatagat ctaagataga caaaattaat gaattaattg atgccttgaa 10920

tgaaaagaat atacttttag aaaagcaaga ggatttgttg tatgaagaac atgataagtt 10980

tgtagaggct caaaaatccc ttgcattaga aattaagagg aatgaaatgc ttgcttgtga 11040

agtgtcatca tgccatgatt ctatttctaa cttaaagagc ataaatgatg atttaaatgc 11100

taaactcgta gaagcaaata aatccaactc ctgtggtgaa actgttgtta tttgcaatag 11160

atgtaaagac tttgatattg atgcttgtag tgaacacata gcttctattg caaagttaaa 11220

taatgaagtg gctagtctta atgcccaact taagactagc aaaagtgaat ttgataaact 11280

aaaatttgca agggatgcct acactattgg tagacacccc tcaattaagg atggacttgg 11340

cttcaagagg gaagtcaaga acttaacaag ccataagact cccattccca ctaaggagaa 11400

agggaaggct cctatggcta ctagtgctaa aaagaaccat gcctttttat atcatgatag 11460

gagatattct agaaatgctt ttaaaggtca tgatgttttt ggttcacatg cttatgactc 11520

ttatgctatg actgcttcta gttctcatgt tatgcctggt agaaatgtgc ttagaagaaa 11580

tgttgttcat caaatgccta ggagaaatga tgctcatagt gctcctagga aagtagtaaa 11640

tgaatcttct acaatttatt gtgctttaaa tgcttccttt gctatttata gaaaggataa 11700

gaaaattgtt gctaggaaga taggggcaaa atgcaaggga gacaaaactt gcatttgggt 11760

ccctaaggat atttgcacta accttgtagg acccaacatg agttgggtac ctaagtccca 11820

agcctaaatt tgccttgcag gtttatgcat ccgggggttc aagctggatt atagacagcg 11880

gatgcacaaa ccatatgacg ggggagaaga agatgttcac ctcctacgtc aagaaaaagg 11940

attcccaaga ttcaattata ttcggtgatg ggaatcaagg caaggtaaaa gggctaggta 12000

aaattgcaat ttctaatgag cactctatat ctaatgtgtt tttagtagag tctcttggtt 12060

ataatttgct ttctgttagt caattatgca acatggggta taactgtcta tttacaaatg 12120

tagatgtgtc tgtctttaga agaagtgatg gttcactagc ttttaagggt gtattagacg 12180

gcaaacttta tttagttgat tttgcaaaag aagaggccga tctagatgca tgcttaatag 12240

ctaagactag catgggctgg ctgtggcatc gccgcttagc acatgtgggg atgaagaacc 12300

ttcacaagct taagggagaa cacgtgatag gtctaactaa tgttcaattc gaaaaagata 12360

gaccttgtgc agcttgtcaa gcagggaaac aggtgggagg ctctcatcac accaaaaatg 12420

tgatgacgac atcaagaccc ttggagatgc tgcatatgga cctttttgga cccgtcgcct 12480

atctgagcat aggaggaagt aagtatggtt tagttattgt tgatgacttt tcccgcttca 12540

cttgggtgtt ctttttgcag gaaaagtccg aaacccaagg gaccctcaaa cgcttcctca 12600

ggagatctca aaatgagttt gagctcaaag tgaagaagat aaggagcgac aacgggtccg 12660

agttcaagaa tcttcaagtg gaggaattcc ttgaagaaga agggatcaag catgagttct 12720

ccgctcccta cacaccacag caaaatggtg tggtagagag gaagaacagg acgctcatcg 12780

acatggcgag gactatgctt ggagagttca agacccccga gcgtttttgg tcggaagccg 12840

tgaacacggc ttgccacgcc atcaacaggg tctaccttca tcgcctcatc aagaagacgt 12900

cgtatgagcc actaaccggt aacaaaccca atgtatctta ctttcgtgta tttgggagca 12960

agtgctacat tctagtgaag aagggtagaa attctaagtt tgctcccaaa gctgtagaag 13020

ggtttttgtt aggttatgac tcaaatacaa aggcgtatag agtcttcaac aaatcatcgg 13080

gtttggttga agtctctagc gacgttgtat ttgatgagac taatggctct ccaagagagc 13140

aagttgttga ttgtgatgat gtagatgaag aagatgttcc aacggccgct atgcgcacca 13200

tggcgattgg tgatgtgcga ccacaggaac acttggagca agatcagcct tcttcctcaa 13260

ccatagtgca tcccccaacc caagatgacg aacaggtacc tcaagtggag gcgggcgatc 13320

aagggggagc acaagatgat catgtgatga aggaaaaagc gcaaccggca cctccaaccc 13380

aagttcgagc aatgattcaa agggatcatc ccgtcgacca aattctgggt gacatcagca 13440

agggagtaac tactcgttct cgattagtta atttttgtga gcattactct tttgtctcct 13500

ctattgagcc tttcagggta gaggaggcct tgctagatcc ggactgggtg ttggccatgc 13560

aggaggagct caacaacttc aagagaaatg aagtttggac gctggtgcct cgtccgaagc 13620

aaaatgttgt gggaaccaag tgggtgttcc gcaacaaaca ggacgagcac ggggtggtga 13680

cgagaaacaa ggctcgactt gtggcaaaag gttatgccca agtcgcaggt ttggattttg 13740

aggagacttt tgctcctgtg gctaggctag agtcaattcg aatcttgcta gcatatgccg 13800

ctcaccattc tttcaggttg taccaaatgg atgtgaagag cgctttcctc aacgggccaa 13860

tcaaggagga ggtgtacgtg gagcaacccc ctggcttcga ggatgaacgg taccccgacc 13920

acgtgtgtaa gctctctaag gcgctctatg gacttaagca agccccaaga gcatggtatg 13980

aatgccttag agatttctta attgctaatg ctttcaaggt tgggaaagcc gatccaactc 14040

ttttcactaa gacatgtgat ggtgatttgt tcgtgtgcca aatttatgtc gatgacataa 14100

tatttggttc tactaaccaa aagtcttgtg aagagtttag cagggtgatg acgcaaaaat 14160

tcgagatgtc gatgatgggc gagttgaact acttccttgg gttccaagtg aagcaactca 14220

aggacggcac cttcatctcc caaacgaagt acacgcaaga tctgctaaag cggtttggga 14280

tgaaggacgc caagcccgca aagactccga tggggaccga cggacacacc gacctcaaca 14340

aaggaggtaa gtccgttgat caaaaagcat accggtcaat gatagggtca ttactttatt 14400

tatgtgctag tagaccggat attatgctta gcgtatgcat gtgtgctaga tttcaatccg 14460

atcctaagga atgtcactta gtggcggtga agcggattct tagatatttg gttgctacgc 14520

cttgcttcgg gctctggtat ccaaaggggt ctacctttga cttggttgga tactcagact 14580

ccgactatgc tggatgtaag gtcgatagga agagtacatc ggggacgtgc cagttcttag 14640

gaaggtccct ggtgtcatgg aattctaaga aacaaacttt cgttgcccta tccaccgctg 14700

aggccgagta tgttgccgca ggacagtgtt gcgcgcaact actttggatg aggcaaaccc 14760

tcagggactt tggctacaat ctgagcaaag tcccactcct atgtgataat gagagtgcta 14820

tccgcatggc ggaaaatcct gttgaacaca gccgcacaaa gcacatagac atccgacatc 14880

actttttgag agaccaccag caaaagggag atatcgaagt gtttcatgtt agcaccgaga 14940

accagctagc cgatatcttt accaagcctc tagatgagaa gaccttttgc aggttgcgta 15000

gtgagctaaa tgtcatagat tcgcggaacc tggattgaat tgtagcatac atgtacttat 15060

gcttttgatc atgttccttt ttgcattttg ttgcttatta tggtgctcaa gttgtaccaa 15120

cactccctgg acctcacaaa tccgttgcaa agtgatgcac atgtttaggg ggagatgtgt 15180

tacaacttga ccctttgaga ctaaccattt gcttgagttt gcttgattga gtctcgaagg 15240

aggattgaaa gggaaaaggt ggacttggac catgaaagac ttccactgca ctccgatgag 15300

aggctaacta attccaagtt catctcatga aatcttattg ccatttgctc ttaattgaag 15360

actttggtga ggcaatgggg ttaagaggcc aagattaatc ccgttttggt gcttgatgcc 15420

aaagggggag aaaataaagg ccaaagtgat aaatggatca gctaccactt gagagatttt 15480

gaaaatagta gaatagagtt tttgttttgt caaaagcttt tattgtctct tattgtctct 15540

attgtcaaaa gttggcttct tgtggggaga agtgttgatt atgggaaata gggggagttt 15600

ttgaaatcct tgatcaatct cttttggaat gactctcttt atgcttcaac atgtgtgttt 15660

gacttagaga tagagatttg agtttgattt gcaaaaacaa accaagtggt ggcaaaggat 15720

gatccatata tgccaaaatt gaatcaaaac caatttgagt ttttatttga agtgattttg 15780

cacttgttct agttgcttta tgtggtgttg gcataaatca ccaaaaaggg ggagattgaa 15840

agggaaatgt gcctttgggc catttctaag tattttggtg attaagtgcc aacacaagtg 15900

cttaaatgta aatcaatgcc catggatgaa caaactgcaa atcttcaaca aaggtatgtt 15960

tctaagtctt agtacattgg ttttgtgtac taatatattt gtctaagtgt tagaaacaga 16020

aagaagaaga gaagagaaga cttggctgtg tgcagccaag gggctgcttc ggtctagggc 16080

accggactgt ccggtggtgc accggacagt gtccgatggt gcaccggaca gtgtccggtg 16140

cgccaagccg ccttggccga agaggccgct ctcgggtttt tctccggcga cttcggctaa 16200

aattcaccgg actgtccggt gtgcaccgga ctgtccggtg agccaacggt cggccgggcc 16260

aacagtcggc cgcgtgatcg gcgcgcgaca cgtggccgag ccaacggtcg gagggaagca 16320

ccggactgtc cggtgtgcac cggactggcc ggtgcgccag atctgcaacg gtcggcaacg 16380

gtcggcttcg ccaattaagg aaaggaatcg ggcaccggac actgtccggt gtgcaccgga 16440

ctgtccggtg cgcccgacga cagaaggcaa gaatagcctt ccagatgtgc tctcaacggc 16500

tcctaggtgc cttggggcta taaaagggac cccttggcgc atggaggagt acaaccaagc 16560

attcctacaa cctttctaag caccaagaca ttgatctagc gcattcgatt cattgtgata 16620

gcatatagag ctcttgtgga gttgtgtact ctttgagttg tgttgcgagc tcttattgct 16680

gcttgtgtgc gtgttgctct gatcttttga agtcttgtgt gcgtcgctca ttccctcccc 16740

tgctccgtga ttctctgtga acatcttttg taagggcgag aggctccaaa ttgtggagat 16800

tcctcgcaag cgggattgag aaaagaaaag caagaacacc gtggtattca agttgatcat 16860

tggatcactt gagaggagtt gagtgcaact ctcgtccgtt gggacgccac aacgtggaag 16920

taggcaagtt ttgtacttgg ccgaaccacg ggataaccac cgtgtcatct ctgtgattga 16980

tttcttgtgg ttattgtgtt ttgactcctc tctagccact tggccataat tgtgctaaca 17040

cttaacaagt ttttgtggct taagttttga agttattcag gatcacctat tcaccccccc 17100

ctctaggtgc tctcagtttt gagaagactt gctgtgcagg ctatgaacca cactatgcat 17160

aactctttaa catatgtcct tgacttcttt atgtgctatg tttatgcaac aaacataatt 17220

ttgttaggat tacatttgca tgccagatta tttctgaatt gtttcaatgt tcacaatccc 17280

ttaggaccaa gtatatttgt gattaaaatt agcatactcc ataatttgta cttccgttca 17340

tacctataga ttagacgctc cctttttgtt agcataaagt ctacttggaa aatgtaccca 17400

attctaatac acttccactg attgcagctt tacaaaacga gggatgagaa gtaccttctt 17460

gacctgcaaa gggtcagtgg accacaactc ctctttctgg acttgtgttc cgcctttcta 17520

acgcagctga gagttctttg agcctgatga catgattggc ttcccaagca caatccttga 17580

tgcaccccat ggactttcaa attacattcg tgctgtacat aatggccata tataatcctc 17640

cggtagatta tattttctgg agcatgtgga ccgtacaggt atgtcctggt tgccgagatg 17700

attgtcacct aaggtcctaa acaagaggcc acatgcgagc cccaggcctt cttcctcaat 17760

tagattatgc gagttttgtt cggcctttct ttagtccttg gacatgtttt tttctttgcc 17820

ttgtttggtt gtgtttggat caaaggagat tgagagctgg tttgatgacc agggaaatgg 17880

aggggattaa tggggagaaa tctccttcct atttgaaatt gaatagcaag tggatatctc 17940

tcccatgcat cctctctatt tcccgtatca ccaaattagc cctgaggctt tgtttggata 18000

ctctagtatt tatctcagta catatggatt gggatagatt aaaatgtaaa ttaaactaac 18060

ttacacctca atccatgtgc atagatgtaa atactagatt attcaaacaa agcttgaggg 18120

ggattaaatc cccttatatt caattttgac tagtgagaga tttaatcact ccaatcccat 18180

ttgatcccga tgccctaagg tgatcgtgct agcatggtac ctaaggctga aaatgttcca 18240

ttgaaaactg aaaagaaagt ttacccgttg atgtaccagt gtcaactctg acataacagg 18300

agagacagta attagatgtc acagcaagtg cacaatcatc acttcatcag ataaaattac 18360

tattttaggt attgtaagat gcatctccag aggatttatg agaaagggca cacgtgcaag 18420

cacaacaacg gaagagatta gggcagcaaa taactgaatc tgtgctctag ctcaacagca 18480

cgaggacata cgtaatgtac atgggtcagc atatgatttt actttttaca gtaggaaatg 18540

ctagaaaaga cttagctagc ctaaactata atgtacagcg gcaacaccga tggtgtggta 18600

tcagtaattc tcgggtgcat aagaagctga atcaagctgc cccatctgct cagtgatctg 18660

atcaacattg tcctggagac gaggacagca gcggtctacg gcagtcttca catcctcttt 18720

tgcttctatt gcctcgcttt taccctcggc ggcgtctgtg agaaaccgct cccagaatac 18780

atcgttcact ttggccggca catgggaacc tccagtttca ctggtggtat catcttgcga 18840

ggtgtcgccg gttgtggtgt cagagttcat atcgactcca gcagctgtcc tatgggcatg 18900

ttcatggagc tgtgccatcg gcagcatcgg gctctgggcg tagtagctcg ggggttctgc 18960

caacggagaa gacgactggt cttgccctga ggtcgaagcg aaggggttct ggcaagctga 19020

tggctgcaga tcgatctcgg tatccattgg tgccgagagc atctcgccta gagccagagc 19080

agggctgggt tcagcaacac cagcggcgta caacatgcct tgagcagcag acgtatgcgc 19140

tgttgccctc tgcaagaaac tctccagcga caccagggcc agctccatct tctcaaacgg 19200

ctcggcgctc agaaggcgga tcggagacat gcctggcgtc tccgcgccca ccgtctgcag 19260

gaacggcacc tgctgttctt cgaccttggg ttcgtcgacg aagaagtcca tcttcggaac 19320

cctcctcttc ttgctgaagt ggtccgcccc cgtcgccgac ccccacgaaa cgaccccgtg 19380

tccgaccggc agcaggatgt cgcgcaggga ggccacgatg ttcctctggc gctcctccat 19440

ctgcgccagc ctgcgttcca gcgactgcat ctgccagctg atcccgcacc gctgctggct 19500

ctgcctctgg aggtccgtca ggagcaggct cttctcgtac ttgagcctgc tgatctcgtc 19560

ctcgagctcg cgcctctccg actccgccag tgggccgttc acctgggtct gcggcgagtg 19620

gctgtgcacc ggcttgcgcc ggtggatgtt cttcaggagg tgcgtgtgcc ccctaacgaa 19680

gtcgtcgttg gcgaactccc atctctcggg atcgatcttt cggaaaccct ggaagtgaag 19740

aaccaagtta gcagcaggcg gaaagtgagg acagacatga ggagtgagga ccagctgtta 19800

aaaacagccg ccacttttaa aacatgggct tcctactata cagaatagga acagagcaga 19860

gagcacacag aatcccgtgc aaaaaaaatc aagaaccaac tttgaattga atgcccgtaa 19920

tgctcctgtt gctatttggg ttatgagaca aatacgtctc tcttcatgaa tagacgactc 19980

cacgatcagg tgcgctggtg ccaactgaac atgcgaaaaa tgccgaggaa aaaggcacat 20040

cgtcttgagg aagatagcag acaggaaatg gcaagaaata gacgcgcaac agcaacatgt 20100

cacacgcaca cagagagaga gattgagaga gggagattta gaggcactca cgtatgtgtt 20160

gagctgcctg atgaagctgg agaagttgtt gtgcttgaag tacttgggca gcaggtcccg 20220

tgagaagtcc ggcgggttcc acaccacgaa gcttgcgccg ccggggcccc acgacacgac 20280

gtggttggtc gccgggtcct ccaccatctc gtaggtcttg atgaggaacg gcggcggctg 20340

cgagctggaa ttggaaccct ccatgccggc catgcacctc ctcctgtctc tcctccgact 20400

gatgatgggc gaagaattaa ctaaccgacg ctcccccaac tctgtcttac ccaccggaat 20460

ggctcgcact caaccgcagc gagaaaaatt ctcgccaagc tgccgcaagg acgacgaaag 20520

caagggctac ggctactgaa ggggaagaaa cctgagcggt tgctaggatt tttgctcctc 20580

gcgttggcgt cagctcgggc gcggcagatg agcgagccga ggcagcccaa ggggcagagg 20640

agaaaacgag ggggagtggc gtcgcgcggc ggttttataa agggggtcgg gtcgcgcgtc 20700

gtgtcgtgtc gtgtgtccgg gtgcgcggcg ggccaccggg actcggacga cgacgcggac 20760

ggtgactggg ctgagcagct cgcgctggct cgctggcgcc cgcatcctgg gctgggtata 20820

catcggctgg cggctggtcg ttgcatgggg gcgtctgagg tttgactggc caatgcgttg 20880

gcggaccggg acccaggtcg cttctagaag aaggcgcgcg gaaacgggtc ccggcggcgg 20940

gccgtcaaag ttcgccgccc agcgcgcacg ggcaaaggct cggtgctgcc ggtcagcagg 21000

gcaagcgagc cagccgccag cgttcgatag ctgtcggtcg tctgggtgac tggaacaaca 21060

gcagcagcag gccagttttg gggccgagac gtcccgacct ctgtgcgcgt cccgtcacat 21120

actcgcatgt gctgctgctc cgacacttcc cttccctaag ctggcgaccg ttgctctcca 21180

gtgacgtcgg tcacctccag gagccaggaa tcttcccaac gcccctcgcc ctgcccctgc 21240

ggcaccccgc gcggccggtt ggagggtcga gcgagacgcc tttccatcgc ccgtcgggtc 21300

gtcgcgggca ggatgtgaac ggtctcggcg ggcgaccatt cctgccgaga attttcagcg 21360

gtgcgccggt gcccgcccgc gcgccgtgga aaaagagggc cgcggatcgc caggcgagtg 21420

gatgctagcg cacgcacgag cagattcgtt gggcagaaga accacgcgcc gacacgacgg 21480

cgacggcagg cttgaactat gtccgcgccg cgggcgctgc gtgtgtgtgt gtgtgccacc 21540

ttttcttcct taactttgtc cccaagaatg aacgaactga aggaaaaaaa gtctagtgtt 21600

ttaatcgatc tgcaagtgtg tcacgcagtc tcagatcaag agtcaatttc catgctgata 21660

ccgagaggca aggttaattt gggtcattga ataccagtcg gtagtatttg gtcatttgca 21720

tgtcgttgtt gttcttgcca cagatcaagt tgtgaggttt gtggtgctgc tagagatgtc 21780

caaacgggcc gcccggcccg gtccggcctg ggcccggtga agcccggccc gctaggcacg 21840

attaaaaaac cgggtcggac cggctaagca cgcgggctca atttcttgtc cgagtccggt 21900

ccgcaacggg cctaaatggg tcgggccagc acaaaaaaag cgggccggaa agcgggctat 21960

gcggaccgaa aaacacattt tagtgtaaaa aaagcgggct taacgggctt agagctaaac 22020

ggaccgtgcc gggctagcct accgtgtcta atttcctgtc cgagcccggc ccgcttattc 22080

gtgccgggcc gacccgggcc cgcatataac cgggccgtgc cgggctcgga ccgagcccca 22140

aaagcgggct tcgtgccggg ctcgcgggcc tcgtgcctat aggtgctgct acgttcccat 22200

caagtgcatg tgcatgtgaa ttgaggaagc aatcaagttc ctcttgcctt gctccctttt 22260

gatcggtaac gaagagaaaa aagaaaagag atcagatttc aacatatttg ccacacacgc 22320

ctgagcagat tagcgcaagg ttttaagttt attactaaat atacacaggt tatgtaatag 22380

ccatcatttt ttttttgcta gctggacgtc agcgaactga ggcctgagaa gcacggcaat 22440

aataatcctg tgcgacggag cgagccgcta gctggacgtc agctatggtg tcaccgcgcg 22500

gaaggggggc atgcaagggg cggggttgac ctcggcgtca ggttgttgct gaatgtttct 22560

gggggttcct atcattgctt cttctagact agatgagcat ggacttgtgg tgtggtggca 22620

tgggcatgca acatgcatgc aggatcaggc caggctacca gggttttttt ttgatctatc 22680

tgcacgagct gatatattag tgtacccagt ttcttcttct agattgttca acttgtgc 22738

Claims (6)

1. Any of the following applications of protein-related biomaterials:

q1, application of the biological material in improving the saline-alkali tolerance of plants;

q2, application of the biological material in preparing a product for improving salt and alkali resistance of plants;

the protein is the protein of A1) or A2) as follows:

a1 Amino acid sequence is protein of sequence 2 in the sequence table;

a2 Fusion proteins obtained by ligating protein tags at the N-terminus or/and the C-terminus of A1);

the biomaterial is any one of the following B1) to B7):

b1 A nucleic acid molecule encoding said protein;

b2 An expression cassette comprising the nucleic acid molecule of B1);

b3 A recombinant vector comprising the nucleic acid molecule of B1) or a recombinant vector comprising the expression cassette of B2);

b4 A recombinant microorganism comprising the nucleic acid molecule of B1), or a recombinant microorganism comprising the expression cassette of B2), or a recombinant microorganism comprising the recombinant vector of B3);

b5 A transgenic plant cell line comprising the nucleic acid molecule of B1) or a transgenic plant cell line comprising the expression cassette of B2);

b6 A transgenic plant tissue comprising the nucleic acid molecule of B1) or a transgenic plant tissue comprising the expression cassette of B2);

b7 A transgenic plant organ comprising the nucleic acid molecule of B1) or a transgenic plant organ comprising the expression cassette of B2);

the plant is corn.

2. The use according to claim 1, characterized in that: b1 The nucleic acid molecule is as shown in b 1) or b 2) below:

b1 A coding sequence of the coding chain is a cDNA molecule or a DNA molecule of a nucleotide of a sequence 1 in a sequence table;

b2 Nucleotide is cDNA molecule or DNA molecule of sequence 1 in sequence table.

3. Use of any one of the proteins described in claim 1:

p1, application of the protein in improving salt and alkali tolerance of plants;

p2, the application of the protein in preparing a product for improving the salt and alkali resistance of plants;

the plant is corn.

4. A method for cultivating saline-alkali tolerant corn, comprising increasing the expression level of the coding gene of the protein of claim 1 in target corn to obtain saline-alkali tolerant corn; the salt and alkali tolerance of the salt and alkali tolerant corn is higher than that of the target corn.

5. The method according to claim 4, wherein: the improvement of the expression level of the gene encoding the protein of claim 1 in the corn of interest is achieved by introducing the gene encoding the protein of claim 1 into the corn of interest.

6. Use of the method of claim 4 or 5 for any of the following:

d1, application in improving salt and alkali resistance of plants;

d2, application in preparing a product for improving salt and alkali resistance of plants;

the plant is corn.

Images