CN1566146A - Paddy rice stalk extension gene, coded protein and application thereof - Google Patents
Paddy rice stalk extension gene, coded protein and application thereof Download PDFInfo
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Abstract
本发明提供了一种新的植物基因－水稻茎杆伸长基因，及其编码蛋白和启动子。本发明还公开了编码这种茎杆伸长基因的的用途，尤其是在植物生长改良和杂交育种中用于改变植物的茎杆伸长性和抗衰老性。The invention provides a new plant gene-rice stem elongation gene, its encoded protein and its promoter. The invention also discloses the application of the coding stem elongation gene, especially for changing the stem elongation and anti-aging properties of plants in plant growth improvement and hybrid breeding.
Description
Technical field
The invention belongs to the phytology field, specifically, the present invention relates to new paddy rice (Oryza sativa) cane elongation gene, and proteins encoded and promotor.The invention also discloses this cane of coding elongation gene purposes, especially in plant-growth improvement and cross-breeding, be used to change cane extensibility and the anti-ageing property of plant.
Background technology
The quick growth of plant comprises that for crop, production of forestry and opposing bad climate condition drought and water shortage, flood, low temperature etc. have crucial effect, also is simultaneously the differentiation of research vegetable cell, hormone regulating and controlling, the physiological good model of molecule.
Yet, still do not disclose at present and the plant related gene of growing fast.Therefore, in order to change the quick growth of plant variety effectively, pointedly, this area presses for exploitation and proteins associated of growth property fast and encoding gene thereof.
Summary of the invention
An object of the present invention is to provide a kind of new stem elongation albumen (being rice stem stem elongation albumen) and fragment, analogue and derivative.
Another object of the present invention provides the polynucleotide of these polypeptide of coding.
Another object of the present invention provides the method for producing these polypeptide and the purposes of this polypeptide and encoding sequence, is especially changing the plant purposes aspect the growth property (delaying as longer blooming period or fruit maturation) fast.
In a first aspect of the present invention, novel isolated rice stem stem elongation albumen is provided, this peptide source is from paddy rice, and it comprises: polypeptide or its conservative property variation polypeptide or its active fragments or its reactive derivative with SEQ ID NO:3 aminoacid sequence.Preferably, this polypeptide is selected from down group: the polypeptide that (a) has SEQ ID NO:3 aminoacid sequence; (b) SEQ ID NO:3 aminoacid sequence is formed through replacement, disappearance or the interpolation of one or more amino-acid residues, and have promote plant stem elongation function by (a) polypeptides derived.
In a second aspect of the present invention, the polynucleotide of isolating these polypeptide of coding are provided, these polynucleotide comprise a nucleotide sequence, and this nucleotide sequence is shown at least 70% homogeny with a kind of nucleotides sequence that is selected from down group: (a) the proteic polynucleotide of the above-mentioned rice stem stem elongation of coding; (b) with polynucleotide (a) complementary polynucleotide.Preferably, this polynucleotide encoding has the polypeptide of aminoacid sequence shown in the SEQ ID NO:3.More preferably, the sequence of these polynucleotide is be selected from down group a kind of:
(a) has the sequence of 1-5900 position among the SEQ ID NO:1;
(b) SEQ ID NO:1 has lacked the formed sequence in 2927-3169 position or 3153-3157 position;
(c) has the sequence of 1-2172 position among the SEQ ID NO:2.
In a third aspect of the present invention, the carrier that contains above-mentioned polynucleotide is provided, and has been transformed or host cell of transduceing or the host cell that is directly transformed or transduce by above-mentioned polynucleotide by this carrier.
In a fourth aspect of the present invention, provide preparation rice stem stem elongation proteic method, this method comprises: (a) under conditions suitable for the expression, cultivate the above-mentioned host cell that is transformed or transduce; (b) from culture, isolate rice stem stem elongation albumen.
In a fifth aspect of the present invention, provide and above-mentioned rice stem stem elongation protein-specific bonded antibody.The nucleic acid molecule that can be used for detecting also is provided, and it contains a successive 10-2226 Nucleotide in the above-mentioned polynucleotide.
In a sixth aspect of the present invention, provide and whether had the proteic method of rice stem stem elongation in the proteic antibody of anti-the present invention and a kind of test sample, it comprises: sample is contacted with the proteic specific antibody of rice stem stem elongation, observe whether form antibody complex, formed antibody complex and just represented to exist in the sample rice stem stem elongation albumen.
In a seventh aspect of the present invention, provide a kind of change plant fast growth property and or the method for anti-ageing property, it comprises step:
(1) provide the Agrobacterium of carrying expression vector, described expression vector contains rice stem stem elongation protein D NA encoding sequence, and described rice stem stem elongation albumen is selected from down group:
(a) has the polypeptide of SEQ ID NO:3 aminoacid sequence;
(b) SEQ ID NO:3 aminoacid sequence is formed through replacement, disappearance or the interpolation of one or more amino-acid residues, and have promote plant stem elongation function by (a) polypeptides derived;
(2) vegetable cell or tissue or organ are contacted with Agrobacterium in the step (1), thereby make rice stem stem elongation protein D NA encoding sequence change vegetable cell over to, and be incorporated on the karyomit(e) of vegetable cell;
(3) select vegetable cell or tissue or the organ that changes rice stem stem elongation protein D NA encoding sequence over to;
(4) vegetable cell in the step (3) or tissue or neomorph are become plant.
Others of the present invention are because disclosing of the technology of this paper is conspicuous to those skilled in the art.
Description of drawings
Fig. 1 has shown the g and D of Eui and mutant (eui) controlling plant cane thereof.
Fig. 2 has shown that eui can eliminate the bag neck of rice sterile line.
Fig. 3 has shown the merismatic cell fission of Eui Gene Handling internode.
Fig. 4 has shown that the eui gene can produce or accumulate more Plant hormones regulators,gibberellins (being abbreviated as GA or GA3).
Fig. 5 has shown seedling height under different GA3 concentration are handled, and top curve is the height of band mutator gene eui plant, and lower curve is the height of wild-type Eui gene plant.
Fig. 6 shown in different GA3 concentration and handled plant height at following heading stage, and top curve is the height of band mutator gene eui plant, and lower curve is the height of wild-type Eui gene plant.Fig. 5 and 6 shows that Eui has controlled the susceptibility of plant to GA3.
Fig. 7 has shown that the eui plant has antisenility function.
Fig. 8 has shown the normal elongation of Eui transgenosis eui mutant plant answer.
Embodiment
The inventor is through extensive and deep research, from paddy rice (Oryza sativa.L), obtained first to grow fast and the mutant eui (Elongated upper mostinternode) that highly extends between a first segment, this mutant shows as single-gene recessive inheritance, can extend fast between first segment at heading stage, and it is extremely sensitive to GA3, elongation under the adverse circumstance situation is more rapid, this gene early stage by Primary Location in paddy rice the 5th chromosomal long arm end.And be considered to the CA++ of paddy rice cross breeding breeding.The inventor utilizes the method Fine Mapping of map based cloning and has cloned this functional gene, and the result shows the key enzyme albumen of Eui coding adjusting plant hormone and cell growth.This eui gene is imported long-grained nonglutinous rice, can improve all kinds of hybridisation rice sterile line bag neck proterties.This shows that gene of the present invention can be applicable to the quick growth of plant, transgenosis anti-ageing, the novel plant type is created and the molecular breeding of hybridisation rice kind.
In the present invention, term " paddy rice cane elongation albumen ", " rice stem stem elongation polypeptide ", " Eui albumen ", " Eui polypeptide " etc. are used interchangeably, and all refer to have the albumen or the polypeptide of rice stem stem elongation Argine Monohydrochloride sequence (SEQID NO:3).
In the present invention, term " mutant paddy rice cane elongation albumen ", " albumen between the paddy rice meropodium ", " mutant rice stem stem elongation polypeptide ", " eui albumen ", " eui polypeptide " etc. are used interchangeably, and all refer to have mutant rice stem stem elongation Argine Monohydrochloride sequence.Eui albumen does not have corresponding activity of Eui albumen or loss of activity.In the present invention, " albumen of the present invention or polypeptide " comprises " Eui albumen " and " eui albumen "; " gene of the present invention " refers to the polynucleotide of coding " Eui albumen " and " eui albumen ".
When not particularly pointing out, term " stem elongation albumen " comprises mutant and wild-type stem elongation albumen.
As used herein, " isolating " is meant that material separates (if natural substance, primal environment promptly is a natural surroundings) from its primal environment.Do not have separation and purification as polynucleotide under the native state in the active somatic cell and polypeptide, but same polynucleotide or polypeptide as from native state with in other materials that exist separately, then for separation and purification.
As used herein, " isolating rice stem stem elongation albumen or polypeptide " is meant that rice stem stem elongation albumen is substantially free of natural relative other albumen, lipid, carbohydrate or other material.Those skilled in the art can extend albumen with the purified technology of protein purifying paddy rice stem of standard.Basically pure polypeptide can produce single master tape on non-reduced polyacrylamide gel.
Polypeptide of the present invention can be recombinant polypeptide, natural polypeptides, synthetic polypeptide, preferred recombinant polypeptide.Polypeptide of the present invention can be the product of natural purifying, or the product of chemosynthesis, or uses recombinant technology to produce from protokaryon or eucaryon host (for example, bacterium, yeast, higher plant, insect and mammalian cell).The host used according to the recombinant production scheme, polypeptide of the present invention can be glycosylated, maybe can be nonglycosylated.Polypeptide of the present invention also can comprise or not comprise initial methionine residues.
The present invention also comprises the proteic fragment of rice stem stem elongation, derivative and analogue.As used herein, term " fragment ", " derivative " are meant with " analogue " and keep identical biological function or the active polypeptide of natural paddy rice stem elongation albumen of the present invention basically.Polypeptide fragment of the present invention, derivative or analogue can be that (i) has one or more conservative or substituted polypeptide of non-conservation amino-acid residue (preferred conservative amino acid residue), and the amino-acid residue of such replacement can be also can not encoded by genetic code, or (ii) in one or more amino-acid residues, has a polypeptide of substituted radical, or (iii) mature polypeptide and another compound (such as the compound that prolongs the polypeptide transformation period, polyoxyethylene glycol for example) merges formed polypeptide, or (iv) additional aminoacid sequence is fused to this peptide sequence and the polypeptide that forms (as leader sequence or secretion sequence or be used for the sequence or the proteinogen sequence of this polypeptide of purifying or fusion rotein).According to the instruction of this paper, these fragments, derivative and analogue belong to the known scope of those skilled in the art.
In the present invention, term " rice stem stem elongation albumen " refers to have the SEQ ID NO.3 polypeptide of sequence of rice stem stem elongation protein-active.This term also comprises having and variant form rice stem stem elongation albumen identical function, SEQ ID NO.3 sequence.These variant forms comprise (but being not limited to): several (are generally 1-50, preferably 1-30, more preferably 1-20,1-10 best) amino acid whose disappearance, insertion and/or replacement, and add one or several at C-terminal and/or N-terminal and (be generally in 20, preferably being in 10, more preferably is in 5) amino acid.For example, in the art, when replacing, can not change proteinic function usually with the close or similar amino acid of performance.Again such as, add one or several amino acid at C-terminal and/or N-terminal and also can not change proteinic function usually.This term also comprises proteic active fragments of rice stem stem elongation and reactive derivative.
The variant form of this polypeptide comprises: homologous sequence, conservative property varient, allelic variant, natural mutation, induced mutation body, under high or low tight degree condition can with coded albumen of the DNA of rice stem stem elongation protein D NA hybridization and polypeptide or the albumen that utilizes the proteic antiserum(antisera) of anti-rice stem stem elongation to obtain.The present invention also provides other polypeptide, as comprises rice stem stem elongation albumen or its segmental fusion rotein.Except the polypeptide of total length almost, the present invention has also comprised the proteic soluble fragments of rice stem stem elongation.Usually, this fragment have rice stem stem elongation protein sequence at least about 10 continuous amino acids, usually at least about 30 continuous amino acids, preferably at least about 50 continuous amino acids, more preferably at least about 80 continuous amino acids, best at least about 100 continuous amino acids.
Invention also provides the analogue of rice stem stem elongation albumen or polypeptide.It can be difference on the aminoacid sequence that these analogues and natural paddy rice stem extend proteic difference, also can be the difference that does not influence on the modified forms of sequence, perhaps haves both at the same time.These polypeptide comprise natural or the inductive genetic variant.The induce variation body can obtain by various technology, as by radiation or be exposed to mutagenic compound and produce random mutagenesis, also can pass through site-directed mutagenesis method or the biological technology of other known moleculars.Analogue also comprises having the analogue that is different from the amino acid whose residue of natural L-(as D-amino acid), and has non-natural analogue that exist or synthetic amino acid (as β, gamma-amino acid).Should be understood that polypeptide of the present invention is not limited to the above-mentioned representational polypeptide that exemplifies.
(the not changing primary structure usually) form of modification comprises: the chemically derived form such as the acetylize or carboxylated of the polypeptide that body is interior or external.Modify and also comprise glycosylation.Modified forms also comprises have the phosphorylated amino acid residue sequence of (as Tyrosine O-phosphate, phosphoserine, phosphothreonine).Thereby also comprise the polypeptide that has been improved its anti-proteolysis performance or optimized solubility property by modifying.
In the present invention, " rice stem stem elongation albumen conservative property variation polypeptide " refers to compare with the aminoacid sequence of SEQ ID NO:3, has 10 at the most, preferably at the most 8, more preferably at the most 5,3 amino acid is replaced by similar performance or close amino acid and is formed polypeptide at the most best.
Polynucleotide of the present invention can be dna form or rna form.Dna form comprises the DNA of cDNA, genomic dna or synthetic.DNA can be strand or double-stranded.DNA can be coding strand or noncoding strand.The coding region sequence of encoding mature polypeptide can be identical with the coding region sequence shown in the SEQ ID NO:1 or the varient of degeneracy.As used herein, " varient of degeneracy " is meant that in the present invention coding has the protein of SEQ ID NO:3, but with the differentiated nucleotide sequence of coding region sequence shown in the SEQ ID NO:1.
The polynucleotide of the mature polypeptide of coding SEQ ID NO:3 comprise: the encoding sequence of an encoding mature polypeptide; The encoding sequence of mature polypeptide and various additional code sequence; Encoding sequence of mature polypeptide (with optional additional code sequence) and non-coding sequence.
Term " polynucleotide of coded polypeptide " can be the polynucleotide that comprise this polypeptide of encoding, and also can be the polynucleotide that also comprise additional code and/or non-coding sequence.
The invention still further relates to the varient of above-mentioned polynucleotide, its coding has the polypeptide of identical aminoacid sequence or fragment, analogue and the derivative of polypeptide with the present invention.The varient of these polynucleotide can be the allelic variant of natural generation or the varient that non-natural takes place.These nucleotide diversity bodies comprise and replace varient, deletion mutation body and insert varient.As known in the art, allelic variant is the replacement form of polynucleotide, and it may be replacement, disappearance or the insertion of one or more Nucleotide, but can be from not changing the function of its encoded polypeptides in fact.
The invention still further relates to and above-mentioned sequence hybridization and two sequences between have at least 50%, preferably at least 70%, the polynucleotide of at least 80% homogeny more preferably.The present invention be more particularly directed under stringent condition and the interfertile polynucleotide of polynucleotide of the present invention.In the present invention, " stringent condition " is meant: (1) than hybridization under low ionic strength and the comparatively high temps and wash-out, as 0.2 * SSC, and 0.1%SDS, 60 ℃; Or (2) hybridization the time is added with denaturing agent, as 50% (v/v) methane amide, 0.1% calf serum/0.1%Ficoll, 42 ℃ etc.; Or (3) only at the homogeny between the two sequences at least more than 90%, be more preferably 95% and just hybridize when above.And the polypeptide of interfertile polynucleotide encoding has identical biological function and activity with the mature polypeptide shown in the SEQ ID NO:3.
The invention still further relates to nucleic acid fragment with above-mentioned sequence hybridization.As used herein, the length of " nucleic acid fragment " contains 15 Nucleotide at least, better is at least 30 Nucleotide, is more preferably at least 50 Nucleotide, preferably more than at least 100 Nucleotide.Nucleic acid fragment can be used for the amplification technique (as PCR) of nucleic acid to determine and/or the proteic polynucleotide of separation coding rice stem stem elongation.
Rice stem stem elongation pyrenoids thuja acid full length sequence of the present invention or its fragment can obtain with the method for pcr amplification method, recombination method or synthetic usually.For the pcr amplification method, can be disclosed according to the present invention about nucleotide sequence, especially open reading frame sequence designs primer, and with commercially available cDNA storehouse or by the prepared cDNA storehouse of ordinary method well known by persons skilled in the art as template, amplification and must relevant sequence.When sequence is longer, usually needs to carry out twice or pcr amplification repeatedly, and then the fragment that each time amplifies is stitched together by proper order.
In case obtained relevant sequence, just can obtain relevant sequence in large quantity with recombination method.This normally is cloned into carrier with it, changes cell again over to, separates obtaining relevant sequence then from the host cell after the propagation by ordinary method.
In addition, also the method for available synthetic is synthesized relevant sequence, especially fragment length more in short-term.Usually, by first synthetic a plurality of small segments, and then connect and to obtain the very long fragment of sequence.
At present, can be fully obtain the dna sequence dna of code book invention albumen (or its fragment, or derivatives thereof) by chemosynthesis.This dna sequence dna can be introduced in various existing dna moleculars as known in the art (or as carrier) and the cell then.In addition, also can will suddenly change and introduce in the protein sequence of the present invention by chemosynthesis.
The present invention also relates to comprise the carrier of polynucleotide of the present invention, and the host cell that produces through genetically engineered with carrier of the present invention or rice stem stem elongation albumen coded sequence, and the method that produces polypeptide of the present invention through recombinant technology.
Recombinant DNA technology (Science, 1984 by routine; 224:1431), can utilize polymerized nucleoside acid sequence of the present invention to can be used to express or produce the rice stem stem elongation albumen of reorganization.In general following steps are arranged:
(1). with the proteic polynucleotide of coding rice stem stem elongation of the present invention (or varient), or transform or the transduction proper host cell with the recombinant expression vector that contains these polynucleotide;
(2). the host cell of in suitable medium, cultivating;
(3). separation, protein purification from substratum or cell.
Among the present invention, rice stem stem elongation albumen polynucleotide sequence can be inserted in the recombinant expression vector.Term " recombinant expression vector " refers to bacterial plasmid well known in the art, phage, yeast plasmid, vegetable cell virus, mammalian cell virus or other carriers.In a word, as long as can duplicate in host and stablize, any plasmid and carrier can be used.A key character of expression vector is to contain replication orgin, promotor, marker gene and translation controlling elements usually.
Method well-known to those having ordinary skill in the art can be used to make up and contains paddy rice stem elongation encoding histone dna sequence dna and suitable transcribing/the translate expression vector of control signal.These methods comprise extracorporeal recombinant DNA technology, DNA synthetic technology, the interior recombinant technology of body etc.Described dna sequence dna can effectively be connected on the suitable promotor in the expression vector, and is synthetic to instruct mRNA.Expression vector also comprises ribosome bind site and the transcription terminator that translation initiation is used.
In addition, expression vector preferably comprises one or more selected markers, to be provided for selecting the phenotypic character of transformed host cells, cultivate Tetrahydrofolate dehydrogenase, neomycin resistance and the green fluorescent protein (GFP) of usefulness as eukaryotic cell, or be used for colibacillary tsiklomitsin or amicillin resistance.
Comprise the carrier of above-mentioned suitable dna sequence dna and suitable promotor or control sequence, can be used to transform appropriate host cell, so that it can marking protein.
Host cell can be a prokaryotic cell prokaryocyte, as bacterial cell; Or eukaryotic cell such as low, as yeast cell; Or higher eucaryotic cells, as vegetable cell.Representative example has: intestinal bacteria, streptomyces, Agrobacterium; Fungal cell such as yeast; Vegetable cell etc.
When polynucleotide of the present invention are expressed in higher eucaryotic cells, be enhanced if will make to transcribe when in carrier, inserting enhancer sequence.Enhanser is the cis acting factor of DNA, and nearly 10 to 300 base pairs act on promotor transcribing with enhancing gene usually.
Persons skilled in the art all know how to select appropriate carriers, promotor, enhanser and host cell.
Can carry out with routine techniques well known to those skilled in the art with the recombinant DNA transformed host cell.When the host was prokaryotic organism such as intestinal bacteria, the competent cell that can absorb DNA can be handled with the CaCl2 method in exponential growth after date results, and used step is well-known in this area.Another kind method is to use MgCl2.If desired, transforming also the method for available electroporation carries out.When the host is an eukaryote, can select following DNA transfection method for use: coprecipitation of calcium phosphate method, conventional mechanical method such as microinjection, electroporation, liposome packing etc.Transform plant and also can use methods such as Agrobacterium-mediated Transformation or particle gun conversion, for example leaf dish method.Can use ordinary method regeneration plant for plant transformed cell, tissue or organ, thereby obtain growth property fast/anti-ageing plant that sexually revises.
The transformant that obtains can be cultivated with ordinary method, expresses the polypeptide of coded by said gene of the present invention.According to used host cell, used substratum can be selected from various conventional substratum in the cultivation.Under the condition that is suitable for the host cell growth, cultivate.After host cell grows into suitable cell density, induce the promotor of selection with suitable method (as temperature transition or chemical induction), cell is cultivated for some time again.
The extracellular can be expressed or be secreted into to recombinant polypeptide in the above methods in cell or on cytolemma.If desired, can utilize its physics, the separating by various separation methods with other characteristic and the albumen of purification of Recombinant of chemistry.These methods are well-known to those skilled in the art.The example of these methods includes, but are not limited to: conventional renaturation handles, with protein precipitant handle (salt analysis method), centrifugal, the broken bacterium of infiltration, superly handle, the combination of super centrifugal, sieve chromatography (gel-filtration), adsorption chromatography, ion exchange chromatography, high performance liquid chromatography (HPLC) and other various liquid chromatography (LC) technology and these methods.
The rice stem stem elongation albumen or the polypeptide of reorganization are of use in many ways.For example be used to screen antibody, polypeptide or other part that promotes or resist rice stem stem elongation protein function.Can be used for seeking the valuable peptide molecule that can suppress or stimulate rice stem stem elongation protein function with the reorganization rice stem stem elongation protein screening peptide library of expressing.
On the other hand, the present invention also comprises rice stem stem elongation protein D NA or the polypeptide of its fragment coding has specific polyclonal antibody and monoclonal antibody, especially monoclonal antibody.
The present invention not only comprises complete mono-clonal or polyclonal antibody, but also comprises having immunocompetent antibody fragment or chimeric antibody.
Antibody of the present invention can be prepared by the known various technology of those skilled in that art.For example, the rice stem stem elongation protein gene product of purifying or its have antigenic fragment, can be applied to animal to induce the generation of polyclonal antibody.Similarly, expressing rice stem stem elongation albumen or its has antigenic segmental cell and can be used to immune animal and produce antibody.This type of monoclonal antibody can utilize hybridoma technology to prepare.Each antibody-like of the present invention can utilize the fragment or the functional zone of rice stem stem elongation protein gene product, obtains by the routine immunization technology.These fragments or functional zone can utilize recombinant methods or utilize Peptide synthesizer synthetic.Can come immune animal and produce with the gene product of producing in the prokaryotic cell prokaryocyte (for example E.Coli) with the unmodified form bonded antibody of rice stem stem elongation protein gene product; With posttranslational modification form bonded antibody (as the albumen or the polypeptide of glycosylation or phosphorylation), can come immune animal and obtain with the gene product that produces in the eukaryotic cell (for example yeast or insect cell).The proteic antibody of anti-rice stem stem elongation can be used for the rice stem stem elongation albumen in the test sample.
The invention still further relates to the testing method of quantitative and detection and localization rice stem stem elongation protein level.These tests are known in the art, and comprise that FISH measures and radioimmunoassay.The rice stem stem elongation protein level that is detected in the test can be used for explaining rice stem stem elongation albumen importance aspect growth property fast and inhibition elongation.
Whether having the proteic method of rice stem stem elongation in a kind of detection test sample is to utilize the proteic specific antibody of rice stem stem elongation to detect, and it comprises: sample is contacted with rice stem stem elongation protein specific antibody; Observe whether form antibody complex, formed antibody complex and just represented to exist in the sample rice stem stem elongation albumen.
Part or all of polynucleotide of the present invention can be used as probe stationary on microarray (microarray) or DNA chip (being called " gene chip " again), is used for analyzing the differential expression analysis of tissue gene.Carry out RNA-polymerase chain reaction (RT-PCR) amplification in vitro with the special primer of paddy rice stem elongation albumen and also can detect the proteic transcription product of rice stem stem elongation.
In an example of the present invention, a kind of isolating polynucleotide are provided, its coding has the polypeptide of aminoacid sequence shown in the SEQ ID NO:3.Polynucleotide of the present invention are isolated from rice genome and cDNA library, its sequence is shown in SEQ ID NO:2, the polynucleotide sequence total length that it comprises is 2172 bases, its open reading frame is positioned at the 1-2172 position, and the coding total length is 723 amino acid whose rice stem stem elongation albumen (SEQ ID NO:3).Also separate the genome sequence of cane elongation gene in another embodiment, shown in SEQID NO:1, (comprised intron).
Rice stem stem elongation albumen has the function of clear and definite controlling plant stem elongation, is the new gene of transgenic plant such as a transgenic crop that can be used to make up justice and antisense, flowers, fruits and vegetables.
The quick growth of plant for a change of rice stem stem elongation albumen comprises and suppresses to extend, anti-ageing property provides new approach, thereby had great application prospect.Can import justice and antisense stem elongation gene, can change the quick growth of existing good variety of crops, obtain stem elongation or suppress the farm crop such as wheat, paddy rice of elongation or other kinds such as woods grass, fruit tree, flowers and trees etc. fast, solve the practical problems that agroforestry exist in producing.
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment only to be used to the present invention is described and be not used in and limit the scope of the invention.The experimental technique of unreceipted actual conditions in the following example, usually according to people such as normal condition such as Sambrook, molecular cloning: laboratory manual (NewYork:ColdSpring Harbor Laboratory Press, 1989) or molecular biology of plants-laboratory manual (PlantMolecular Biology-A Laboratory Mannual, Melody S.Clark compiles, Springer-verlag Berlin Heidelberg, 1997) condition described in, or the condition of advising according to manufacturer.
Embodiment 1
The clone of rice stem stem elongation protein gene
Comprising of utilizing that the contriver makes up is that stem elongation protein gene Eui and mutator gene eui thereof form, those skilled in that art paddy gene positional cloning (map-based cloning or positioncloning) colony clearly, press molecule marker location Eui/eui in a little genomic fragment, in 30kb.On this basis, comprise this segmental genomic dna cloning with the ordinary method separation.Through enzyme cut and further hybridization identify and determine one of them stem that contains complete paddy rice elongation albumen Eui gene.
Show through full nucleotide sequence analysis result: rice stem stem elongation full length gene is 5900bp (SEQ ID NO:1 comprises control region and intron).Through software analysis and cDNA clone, its ORF shown in SEQ ID NO:2, the position, the coding total length is 723 amino acid whose rice stem stem elongation albumen, its sequence is shown in SEQ ID NO:3.
Through sequential analysis, rice stem stem elongation albumen shown in SEQ ID NO:3, may be the P450 monooxygenase albumen (P450 monooxganase) of a hormone metabolism process.
Further experiment shows, this albumen or degraded hormone GA or Auxin or brassinosteroids, or suppress synthetic GA or Auxin or brassinosteroids.And mutator gene eui can not be formed with the albumen of above-mentioned functions, and make the high-load hormone of plant cylinder accumulation, and the function (susceptibility) (Fig. 4-6) of increase external source GA, impel vegetable cell to divide, extend (Fig. 3) fast, make especially top internode highly elongation fast of plant stem, plant height increases, and the sterile line of eui is highly extended because of quick, no longer wrap neck (seeing Fig. 1,2).Because the eui plant contains the hormone of a large amount, so the vitality of plant is strengthened anti-ageing (Fig. 7).
For verifying the function of the Eui gene of cloning, the inventor is familiar with rice stem stem elongation gene Eui (promotor that contains itself) by above-mentioned those skilled in that art paddy rice transgenic method, forward in the plant that contains mutator gene eui, verified the function that Eui can complementary eui, transfer-gen plant restore normal growth (Fig. 8).
Embodiment 2
The RT-PCR method obtains the proteic encoding sequence of rice stem stem elongation
In order to verify the exactness of sequence,, design and synthesized following primer according to the sequence of SEQ ID NO:1:
Forward primer 5 '-3 ' ACGGAGGCCCATGAGCAAATTTCAA (SEQ ID NO:4)
Reverse primer 5 '-3 ' ACGTATCAGAGCCAGTCACAGCTTGG (SEQ ID NO:5).
Correspondingly, be to obtain the cDNA probe of stem elongation protein gene, design and synthesized following primer:
Forward primer 5 '-3 ' ACCAGCGTCATCTTCAGCA (SEQ ID NO:6)
Reverse primer 5 '-3 ' AACGTCTCCCGCACCAC (SEQ ID NO:7)
The cDNA probe of RT-PCR amplification 500bp.
To use the extractive paddy rice Poly of ordinary method A
+RNA is a template, carries out RT-PCR amplification, amplification condition be 94 ℃ 5 minutes, 94 ℃ 30 seconds, 60 ℃ 30 seconds, 72 ℃ were carried out 35 circulations in 1 minute subsequently, extended 5 minutes with 72 ℃ at last.The electrophoresis detection pcr amplification product reclaims, be connected to adopt on the T-Easy carrier stop the thing fluorescent mark (Big-Dye, Perkin-Elmer, method USA), (Perkin-Elmer checks order on USA) at ABI 377 sequenators.The sequencing result result verification exactness of the proteic complete encoding sequence of rice stem stem elongation (SEQ ID NO:2).
Embodiment 3
The proteic structural analysis of rice stem stem elongation
According to the sequence of the Nucleotide of cDNA (SEQ ID NO:2), the stem that obtains coding extends proteic sequence (SEQ ID NO:3).Through the albumen database online query, stem elongation albumen contains the Cytochrome P450 structural domain of high conservative.So conclude that stem elongation albumen is a P450 monooxygenase.
Embodiment 4
The proteic point mutation of rice stem stem elongation
Through the sequence of several Eui mutator gene eui relatively, the stem point mutation of extending proteic disappearance, frameshit and P450 conserved regions all can be eliminated wild stem and extends proteic function.2 disappearances (promptly lacking 2927-3169 position and 3153-3157 position among the SEQ ID NO:1) that present embodiment is illustrated in the coding region cause the forfeiture of rice stem stem elongation protein function respectively.
Embodiment 5
Wild-type and the proteic functional experiment of mutant rice stem stem elongation
For checking wild-type stem extends proteic function, the promotor of wild-type rice stem stem elongation gene Eui with itself, the paddy rice transgenic method of being familiar with by those skilled in that art, forward in the plant that contains mutator gene eui, verified that wild-type stem elongation albumen can the proteic function of complementary mutant rice stem stem elongation, transfer-gen plant restore normal growth, and mutant rice stem stem elongation albumen can not produce said function (Fig. 8) among the present invention.
All quote in this application as a reference at all documents that the present invention mentions, just quoted as a reference separately as each piece document.Should be understood that in addition those skilled in the art can make various changes or modifications the present invention after having read above-mentioned teachings of the present invention, these equivalent form of values fall within the application's appended claims institute restricted portion equally.
Sequence table
＜110〉Shanghai Inst. of Plant Physiology, Chinese Academy of Sciences
＜120〉paddy rice cane elongation gene and proteins encoded and purposes
<130>027344
<160>7
<170>PatentIn?version?3.1
<210>1
<211>5900
<212>DNA
＜213〉paddy rice (Oryza sativa)
<400>1
taatcaagta?tgtgagttat?tcaatccttg?gcaatttcca?atacaaatac?agacacacta 60
tgtatggaac?ttttttatat?atagtctatt?aaacaatata?tctttgagcc?gtctgaaacc 120
aaattatatg?atattttttt?agttaatcat?accaagatca?ttaattgagt?aggaatataa 180
gacttcaagg?gggaatcaac?aatattaatt?gtttctgaga?acaattattc?aaagcaattc 240
ataataaaag?catatttatt?ttaggagtgg?gtacacacac?tatctagctt?gtaccatata 300
taagtcaatg?accacgatat?atggctagcc?acctccctcc?atcatgaaat?aaattaacct 360
catacgaatt?tagatataga?atatgtccag?atttattgat?taaatcctag?attgatttat 420
tttaggacga?agagattacc?tgcctgcctg?cactaaccat?ctgcgacgcc?gacgtgtcat 480
ccatcggtcg?acaaaacgta?cgcaggccaa?cacatgcata?cgtacacaca?attgcccatt 540
tgcaccccgc?tccccatgtc?cacgtcagac?ggcgacacgt?acacattaac?acatgtacac 600
gtcgttttca?cgtacgagct?aagcaaatcc?aacgtacaga?aaatagttaa?tttacccggt 660
tgccaactgg?gacacgtacg?ccggccacgt?cacaggtcta?cgtggcgcgg?tgtgggccgg 720
gctaggaggg?ttaaaagccc?acgcttgtgt?gctttggtca?actcgtgcac?ttattactct 780
gaagagttat?tagccggttc?gatgtgtcgc?gattactgtt?gtaattaagt?ctgtctctgt 840
ggcgatatgt?acgttgacga?tgattaaggt?ggtgtttgga?tctagagact?taactttagt 900
ctctgtattt?agacactaat?ttaaagtatt?aaatatagac?taattacaaa?actaattaca 960
taaataaaaa?cttatttgcg?agataatttt?ttttaagcct?aattaatcca?taattagaaa 1020
atatttaatg?tagcatcaca?taggataatc?atgggttaat?taggctcaat?agaatcgtct 1080
cgcgaattag?tctaagatta?tggatagttt?ttattaatag?tctacgttta?atatatataa 1140
ttagtgtcca?aacatccgat?aagatagaga?cttaaaagtt?ttaggtggtg?tttagatcta 1200
gggacttaac?tttagtccct?atatttagac?actaatttag?agtattaaat?atagactact 1260
tacaaaacta?attacataaa?taaaagctaa?ttcgcgagat?aattttttta?agcctaatta 1320
atatataatt?agagaatgtt?tactgtagca?tcacatatgc?taatcatgga?ttaattaggc 1380
tcaatagatt?cgtctcgcga?attagttcaa?gattatggat?gggttttatt?aataatctac 1440
gtttaatatt?tataattagt?gtccaaacat?ccgatatgat?agggacttaa?aagttttaat 1500
ctcatctaaa?tatggtttta?gtctcatcta?aacagggtct?aatagcagaa?agttacgtga 1560
gattcgtata?ctcgtacgta?cgtatttgca?tcggaatttt?caaaatccta?gactctttct 1620
agtactggtg?gtacagtttg?gcaaatctgg?ttttttggat?ggtttcagca?tgtttaattt 1680
gcagtgtgga?attgttgaga?tgctagagat?tttttgcaga?acttatgtga?aatttaatta 1740
ggcgtttcaa?ctgtggctaa?acgtgaagat?atagcgcgct?agagacgtgt?gtgttggcag 1800
tgttgctttg?caatttgcat?aggctcgtgg?tgtttggttc?agaaatttat?ttaaggatcc 1860
ggttgttttt?tttttcattt?tccttggtca?atgttgtaaa?gaatccgtag?aatcttggca 1920
agaagattct?cccaaacctc?caggaaagag?gagaagattc?tccggaatca?gtggaagtag 1980
tgcagtagct?attgactggg?atttttgtaa?gcccggtcaa?taggttcccc?tgtacccttg 2040
gagagaccct?gcaagtgcac?acagatcacg?tatatatcca?catgcctata?tgtgcataga 2100
tcacataggc?acgcatcaaa?agctttttaa?tcctagctag?cttactttaa?tctttgatta 2160
ttttcaatag?aaatttaatg?acgtttaact?tgattcatca?ttcggcgacg?tctcttgaac 2220
atgttaaagg?ctgggccgag?gcccaccaaa?aacggaggcc?catgagcaaa?tttcaaattc 2280
tgactcgcgc?gcggtggcgg?caaaattttg?caggcgagac?gtacctgtac?tggctgcggc 2340
ggcggccggc?gctgtacgtg?acggacccgg?agctcatcgg?cgagatcggg?cggtgcgtgt 2400
cgctcgacat?gggcaagccc?aagtacctcc?agaaaggcca?ggagccactc?ttcggcggcg 2460
gcgtcctcaa?ggccaacggc?gcgtgctggg?cgcgccagcg?caaggtcatc?gcgccggagt 2520
tctacatggc?ccgtgtcagg?gccatggtcc?agctcatggt?cgacgccgcg?cagccgctga 2580
tcgcctcctg?ggaatccagg?atcgacgccg?ctggaggcgc?ggcggcggcg?gaggtcgtcg 2640
tcgacggcga?cctccggagc?ttctccttcg?atgtgatatc?gcgggcttgc?tttgggagtg 2700
attactcgag?ggggagggag?atcttcctcc?gcctccgtga?gctgtccggg?ctcatgtcgg 2760
agaccagcgt?catcttcagc?atcccttcgc?tgaggcacct?gccgacgggg?aagaaccgga 2820
ggatctggag?gctcacgggg?gagatccggt?cgctgatcat?ggagctcgtc?agggagcgga 2880
ggtgcgcggc?gagggcggcg?agggagcacg?gcgggaaggc?ggcgccgccg?tcgccgccgg 2940
agcgcgactt?cctcggctcc?atcatcgaga?acagcggcgg?gcagccgcgg?ccggacgact 3000
tcgtggtgga?caactgcaag?aacatctact?tcgccgggca?cgagacgagc?gcggtcaccg 3060
cgacgtggtg?cctcatgctg?ctcgccgcgc?acccggagtg?gcaggaccgc?gcccgcgccg 3120
aggtgctcga?ggtctgcggc?ggcgacggcg?ccgccgcccc?cgccgcgccg?gacttcgaca 3180
tggtgtcccg?gatgcggacg?gtggggatgg?tggtgcagga?aacgctgcgg?ctgttcccgc 3240
cgtcgtcgtt?cgtggtgcgg?gagacgttcc?gggacatgca?gttgggtagg?ctgctggcgc 3300
ccaagggcac?ctacctgttc?gtcccggtgt?ccaccatgca?ccacgacgtc?gccgcctggg 3360
gcccgacggc?gaggctgttc?gacccgtccc?gcttccgcga?cggcgtggcg?gcggcgtgca 3420
agcacccgca?ggcgtcgttc?atgccgttcg?gcctcggcgc?ccgcacctgc?ctcggccaga 3480
acctcgcgct?cgtcgaggtc?aagacgctcg?tcgccgtcgt?cctcgcccgg?ttcgagttca 3540
cgctctcgcc?ggagtacagg?cactcgccgg?cgttccggct?catcatcgag?ccggagttcg 3600
gcctccgcct?ccgcatccgc?cgcgccggcg?gtcaggacgc?cacgtcacaa?gttgacacat 3660
ctactgcacc?cgtgcatagt?tctcataatt?aattggcttc?tgtaaacagc?tagtaagtcg 3720
tgaactgaaa?aaaatacatc?acacatacag?agacaacaga?ggattaccat?tttaccagca 3780
tacatccatg?gcataccaaa?acaatttact?atacattata?attaacatga?tgctatacac 3840
agtataaaag?gacatgcatt?tttctgcgat?taatggatta?agccgattaa?ctgtttgtca 3900
agtgattaca?gtgtttaatt?cagaatctag?taattaattt?gttgaattta?gatagtctgt 3960
agcctgttcc?tctctgaaag?caacagagca?gagcaaaagg?caaaatcaag?tttcagaagc 4020
acatggaaaa?gtatgatttc?tttggcaaaa?ttgttgtgcc?attctcttat?gctataggcc 4080
tgtggttgag?tgagcggccg?tttgattggg?acgccgacga?acaggtgtct?tttcgtttct 4140
gctttctaat?ttctggcact?aataaaaaca?tgttggttgc?tgagttaaaa?tcaccaaagt 4200
ggagtggaga?tcgcgatgtt?atcctgtaga?cttgttgtat?ctggtctaat?ggtctctgta 4260
agcaaaacga?tcagaagtac?tgaattagtt?caattttttg?atagacctgg?tatggtcttt 4320
taaattcaac?ttttgaaagt?acagtgaaat?acaaatctag?tgatgccata?ttgtgggctt 4380
gtgtagtata?aattgtgaat?caagcctacg?gaattttttg?ctgaaataca?gtacgtgcct 4440
gtcttataag?aaaaacctga?accgagaagc?cattagcgtt?gtcacatacg?acgccacgct 4500
cggtgactcg?gtgagccagc?gtgtgcacca?tcagctcgcc?agcagcgcgc?gccaagaaag 4560
accagcagcc?atgtcggggt?ccggcgacgc?cgcgccggcg?cggcacaacg?cgggtcacgg 4620
tcggcgacga?cggcgcgtcc?tcgtctgggc?cagcttcgcc?gcgctggtcc?tgctgctcgt 4680
cgccgcggcg?gccgccatcg?ccgcgctcgc?cgtgctccgg?ccgcgggacc?cgaccacgga 4740
gctcctgtcc?gtgaacgcca?cgggcgccac?cccgcgcgtc?gccgcgctgc?cggccgtctc 4800
cgtccagctc?aacgtcacct?tcctcctcgt?cgtccgcgtg?cgcaacccga?accgggcgga 4860
gttccgccac?ggcgccgcca?cgacggcgct?cctgtaccgc?ggcgccgagg?tgggcgccgc 4920
cggcgtgccc?gcggggacgg?tgccgagccg?cggcgcggcg?acgctgaggc?tgaacatgac 4980
ggtgcgggcg?gacagggtgg?tggcggcggc?cggggtgggc?ggcctcctcg?cggacgtgct 5040
cgccggcgag?atggagttcg?aggcgaggac?ggaggtgcgc?gggcgggtga?agctgctcgg 5100
gctcgtccgg?cggagcgccg?tggcgaggtc?gctgtgccgc?gtcgtgatcg?gcgtcgccga 5160
cgtgaaggtc?cggcgccagg?agtgccacaa?cgagtccaag?ctgtgactgg?ctctgatacg 5220
tattatacgt?atgtacgctg?tctttgtatt?tgtacatgta?tacgtgtgct?gaagacagtc 5280
gatttgtacg?tactctgata?tatgtgcacg?aatttgtcgt?ggcaataaat?gtttaggcta 5340
ttcataagct?gtttctttgg?tagactgtat?atgtatattg?ttaaaagtta?ctgcagcctc 5400
attgtcattg?ttgtgtttcc?aaaattttgg?agaaatcaac?aatatggtta?ttcaatttac 5460
atttgcaacc?cctaaatata?ttattcaagt?agataaagtg?gtcgcactgt?gtgtttttct 5520
tccatttttt?caaagagata?attaaacttg?gtctgtggtg?cacattgcag?aaagctactg 5580
tgaaacggat?ggactattga?actactctcg?ttgtcattgg?ttggcctaat?gttagattac 5640
atttttctac?ctgaatatat?gttttgagta?attcatgtgt?gttttatttc?ccgtggcatt 5700
acatatatgc?tgtaacattc?cgagttgtat?ttttcttttt?ttatttttat?aatattccta 5760
cgtttcaaag?gagagtctgt?gaaacaaaaa?ggtagattag?aacaacatat?agctctcaaa 5820
ctagtgagaa?atttttgctc?ttaatgcaac?agaggcacaa?caaggagaaa?atattcaaca 5880
aacacgtcta?ggttgctcct 5900
<210>2
<211>2172
<212>DNA
＜213〉paddy rice (Oryza sativa)
<220>
<221>CDS
<222>(1)..(2172)
<223>
<400>2
atg?agc?aaa?ttt?caa?att?ctg?act?cgc?gcg?cgg?tgg?cgg?caa?aat?ttt 48
Met?Ser?Lys?Phe?Gln?Ile?Leu?Thr?Arg?Ala?Arg?Trp?Arg?Gln?Asn?Phe
1 5 10 15
gca?ggc?gag?acg?tac?ctg?tac?tgg?ctg?cgg?cgg?cgg?ccg?gcg?ctg?tac 96
Ala?Gly?Glu?Thr?Tyr?Leu?Tyr?Trp?Leu?Arg?Arg?Arg?Pro?Ala?Leu?Tyr
20 25 30
gtg?acg?gac?ccg?gag?ctc?atc?ggc?gag?atc?ggg?cgg?tgc?gtg?tcg?ctc 144
Val?Thr?Asp?Pro?Glu?Leu?Ile?Gly?Glu?Ile?Gly?Arg?Cys?Val?Ser?Leu
35 40 45
gac?atg?ggc?aag?ccc?aag?tac?ctc?cag?aaa?ggc?cag?gag?cca?ctc?ttc 192
Asp?Met?Gly?Lys?Pro?Lys?Tyr?Leu?Gln?Lys?Gly?Gln?Glu?Pro?Leu?Phe
50 55 60
ggc?ggc?ggc?gtc?ctc?aag?gcc?aac?ggc?gcg?tgc?tgg?gcg?cgc?cag?cgc 240
Gly?Gly?Gly?Val?Leu?Lys?Ala?Asn?Gly?Ala?Cys?Trp?Ala?Arg?Gln?Arg
65 70 75 80
aag?gtc?atc?gcg?ccg?gag?ttc?tac?atg?gcc?cgt?gtc?agg?gcc?atg?gtc 288
Lys?Val?Ile?Ala?Pro?Glu?Phe?Tyr?Met?Ala?Arg?Val?Arg?Ala?Met?Val
85 90 95
cag?ctc?atg?gtc?gac?gcc?gcg?cag?ccg?ctg?atc?gcc?tcc?tgg?gaa?tcc 336
Gln?Leu?Met?Val?Asp?Ala?Ala?Gln?Pro?Leu?Ile?Ala?Ser?Trp?Glu?Ser
100 105 110
agg?atc?gac?gcc?gct?gga?ggc?gcg?gcg?gcg?gcg?gag?gtc?gtc?gtc?gac 384
Arg?Ile?Asp?Ala?Ala?Gly?Gly?Ala?Ala?Ala?Ala?Glu?Val?Val?Val?Asp
115 120 125
ggc?gac?ctc?cgg?agc?ttc?tcc?ttc?gat?gtg?ata?tcg?cgg?gct?tgc?ttt 432
Gly?Asp?Leu?Arg?Ser?Phe?Ser?Phe?Asp?Val?Ile?Ser?Arg?Ala?Cys?Phe
130 135 140
ggg?agt?gat?tac?tcg?agg?ggg?agg?gag?atc?ttc?ctc?cgc?ctc?cgt?gag 480
Gly?Ser?Asp?Tyr?Ser?Arg?Gly?Arg?Glu?Ile?Phe?Leu?Arg?Leu?Arg?Glu
145 150 155 160
ctg?tcc?ggg?ctc?atg?tcg?gag?acc?agc?gtc?atc?ttc?agc?atc?cct?tcg 528
Leu?Ser?Gly?Leu?Met?Ser?Glu?Thr?Ser?Val?Ile?Phe?Ser?Ile?Pro?Ser
165 170 175
ctg?agg?cac?ctg?ccg?acg?ggg?aag?aac?cgg?agg?atc?tgg?agg?ctc?acg 576
Leu?Arg?His?Leu?Pro?Thr?Gly?Lys?Asn?Arg?Arg?Ile?Trp?Arg?Leu?Thr
180 185 190
ggg?gag?atc?cgg?tcg?ctg?atc?atg?gag?ctc?gtc?agg?gag?cgg?agg?tgc 624
Gly?Glu?Ile?Arg?Ser?Leu?Ile?Met?Glu?Leu?Val?Arg?Glu?Arg?Arg?Cys
195 200 205
gcg?gcg?agg?gcg?gcg?agg?gag?cac?ggc?ggg?aag?gcg?gcg?ccg?ccg?tcg 672
Ala?Ala?Arg?Ala?Ala?Arg?Glu?His?Gly?Gly?Lys?Ala?Ala?Pro?Pro?Ser
210 215 220
ccg?ccg?gag?cgc?gac?ttc?ctc?ggc?tcc?atc?atc?gag?aac?agc?ggc?ggg 720
Pro?Pro?Glu?Arg?Asp?Phe?Leu?Gly?Ser?Ile?Ile?Glu?Asn?Ser?Gly?Gly
225 230 235 240
cag?ccg?cgg?ccg?gac?gac?ttc?gtg?gtg?gac?aac?tgc?aag?aac?atc?tac 768
Gln?Pro?Arg?Pro?Asp?Asp?Phe?Val?Val?Asp?Asn?Cys?Lys?Asn?Ile?Tyr
245 250 255
ttc?gcc?ggg?cac?gag?acg?agc?gcg?gtc?acc?gcg?acg?tgg?tgc?ctc?atg 816
Phe?Ala?Gly?His?Glu?Thr?Ser?Ala?Val?Thr?Ala?Thr?Trp?Cys?Leu?Met
260 265 270
ctg?ctc?gcc?gcg?cac?ccg?gag?tgg?cag?gac?cgc?gcc?cgc?gcc?gag?gtg 864
Leu?Leu?Ala?Ala?His?Pro?Glu?Trp?Gln?Asp?Arg?Ala?Arg?Ala?Glu?Val
275 280 285
ctc?gag?gtc?tgc?ggc?ggc?gac?ggc?gcc?gcc?gcc?ccc?gcc?gcg?ccg?gac 912
Leu?Glu?Val?Cys?Gly?Gly?Asp?Gly?Ala?Ala?Ala?Pro?Ala?Ala?Pro?Asp
290 295 300
ttc?gac?atg?gtg?tcc?cgg?atg?cgg?acg?gtg?ggg?atg?gtg?gtg?cag?gaa 960
Phe?Asp?Met?Val?Ser?Arg?Met?Arg?Thr?Val?Gly?Met?Val?Val?Gln?Glu
305 310 315 320
acg?ctg?cgg?ctg?ttc?ccg?ccg?tcg?tcg?ttc?gtg?gtg?cgg?gag?acg?ttc 1008
Thr?Leu?Arg?Leu?Phe?Pro?Pro?Ser?Ser?Phe?Val?Val?Arg?Glu?Thr?Phe
325 330 335
cgg?gac?atg?cag?ttg?ggt?agg?ctg?ctg?gcg?ccc?aag?ggc?acc?tac?ctg 1056
Arg?Asp?Met?Gln?Leu?Gly?Arg?Leu?Leu?Ala?Pro?Lys?Gly?Thr?Tyr?Leu
340 345 350
ttc?gtc?ccg?gtg?tcc?acc?atg?cac?cac?gac?gtc?gcc?gcc?tgg?ggc?ccg 1104
Phe?Val?Pro?Val?Ser?Thr?Met?His?His?Asp?Val?Ala?Ala?Trp?Gly?Pro
355 360 365
acg?gcg?agg?ctg?ttc?gac?ccg?tcc?cgc?ttc?cgc?gac?ggc?gtg?gcg?gcg 1152
Thr?Ala?Arg?Leu?Phe?Asp?Pro?Ser?Arg?Phe?Arg?Asp?Gly?Val?Ala?Ala
370 375 380
gcg?tgc?aag?cac?ccg?cag?gcg?tcg?ttc?atg?ccg?ttc?ggc?ctc?ggc?gcc 1200
Ala?Cys?Lys?His?Pro?Gln?Ala?Ser?Phe?Met?Pro?Phe?Gly?Leu?Gly?Ala
385 390 395 400
cgc?acc?tgc?ctc?ggc?cag?aac?ctc?gcg?ctc?gtc?gag?gtc?aag?acg?ctc 1248
Arg?Thr?Cys?Leu?Gly?Gln?Asn?Leu?Ala?Leu?Val?Glu?Val?Lys?Thr?Leu
405 410 415
gtc?gcc?gtc?gtc?ctc?gcc?cgg?ttc?gag?ttc?acg?ctc?tcg?ccg?gag?tac 1296
Val?Ala?Val?Val?Leu?Ala?Arg?Phe?Glu?Phe?Thr?Leu?Ser?Pro?Glu?Tyr
420 425 430
agg?cac?tcg?ccg?gcg?ttc?cgg?ctc?atc?atc?gag?ccg?gag?ttc?ggc?ctc 1344
Arg?His?Ser?Pro?Ala?Phe?Arg?Leu?Ile?Ile?Glu?Pro?Glu?Phe?Gly?Leu
435 440 445
cgc?ctc?cgc?atc?cgc?cgc?gcc?ggc?ggc?ctg?tgg?ttg?agt?gag?cgg?ccg 1392
Arg?Leu?Arg?Ile?Arg?Arg?Ala?Gly?Gly?Leu?Trp?Leu?Ser?Glu?Arg?Pro
450 455 460
ttt?gat?tgg?gac?gcc?gac?gaa?cag?aaa?aac?ctg?aac?cga?gaa?gcc?att 1440
Phe?Asp?Trp?Asp?Ala?Asp?Glu?Gln?Lys?Asn?Leu?Asn?Arg?Glu?Ala?Ile
465 470 475 480
agc?gtt?gtc?aca?tac?gac?gcc?acg?ctc?ggt?gac?tcg?gtg?agc?cag?cgt 1488
Ser?Val?Val?Thr?Tyr?Asp?Ala?Thr?Leu?Gly?Asp?Ser?Val?Ser?Gln?Arg
485 490 495
gtg?cac?cat?cag?ctc?gcc?agc?agc?gcg?cgc?caa?gaa?aga?cca?gca?gcc 1536
Val?His?His?Gln?Leu?Ala?Ser?Ser?Ala?Arg?Gln?Glu?Arg?Pro?Ala?Ala
500 505 510
atg?tcg?ggg?tcc?ggc?gac?gcc?gcg?ccg?gcg?cgg?cac?aac?gcg?ggt?cac 1584
Met?Ser?Gly?Ser?Gly?Asp?Ala?Ala?Pro?Ala?Arg?His?Asn?Ala?Gly?His
515 520 525
ggt?cgg?cga?cga?cgg?cgc?gtc?ctc?gtc?tgg?gcc?agc?ttc?gcc?gcg?ctg 1632
Gly?Arg?Arg?Arg?Arg?Arg?Val?Leu?Val?Trp?Ala?Ser?Phe?Ala?Ala?Leu
530 535 540
gtc?ctg?ctg?ctc?gtc?gcc?gcg?gcg?gcc?gcc?atc?gcc?gcg?ctc?gcc?gtg 1680
Val?Leu?Leu?Leu?Val?Ala?Ala?Ala?Ala?Ala?Ile?Ala?Ala?Leu?Ala?Val
545 550 555 560
ctc?cgg?ccg?cgg?gac?ccg?acc?acg?gag?ctc?ctg?tcc?gtg?aac?gcc?acg 1728
Leu?Arg?Pro?Arg?Asp?Pro?Thr?Thr?Glu?Leu?Leu?Ser?Val?Asn?Ala?Thr
565 570 575
ggc?gcc?acc?ccg?cgc?gtc?gcc?gcg?ctg?ccg?gcc?gtc?tcc?gtc?cag?ctc 1776
Gly?Ala?Thr?Pro?Arg?Val?Ala?Ala?Leu?Pro?Ala?Val?Ser?Val?Gln?Leu
580 585 590
aac?gtc?acc?ttc?ctc?ctc?gtc?gtc?cgc?gtg?cgc?aac?ccg?aac?cgg?gcg 1824
Asn?Val?Thr?Phe?Leu?Leu?Val?Val?Arg?Val?Arg?Asn?Pro?Asn?Arg?Ala
595 600 605
gag?ttc?cgc?cac?ggc?gcc?gcc?acg?acg?gcg?ctc?ctg?tac?cgc?ggc?gcc 1872
Glu?Phe?Arg?His?Gly?Ala?Ala?Thr?Thr?Ala?Leu?Leu?Tyr?Arg?Gly?Ala
610 615 620
gag?gtg?ggc?gcc?gcc?ggc?gtg?ccc?gcg?ggg?acg?gtg?ccg?agc?cgc?ggc 1920
Glu?Val?Gly?Ala?Ala?Gly?Val?Pro?Ala?Gly?Thr?Val?Pro?Ser?Arg?Gly
625 630 635 640
gcg?gcg?acg?ctg?agg?ctg?aac?atg?acg?gtg?cgg?gcg?gac?agg?gtg?gtg 1968
Ala?Ala?Thr?Leu?Arg?Leu?Asn?Met?Thr?Val?Arg?Ala?Asp?Arg?Val?Val
645 650 655
gcg?gcg?gcc?ggg?gtg?ggc?ggc?ctc?ctc?gcg?gac?gtg?ctc?gcc?ggc?gag 2016
Ala?Ala?Ala?Gly?Val?Gly?Gly?Leu?Leu?Ala?Asp?Val?Leu?Ala?Gly?Glu
660 665 670
atg?gag?ttc?gag?gcg?agg?acg?gag?gtg?cgc?ggg?cgg?gtg?aag?ctg?ctc 2064
Met?Glu?Phe?Glu?Ala?Arg?Thr?Glu?Val?Arg?Gly?Arg?Val?Lys?Leu?Leu
675 680 685
ggg?ctc?gtc?cgg?cgg?agc?gcc?gtg?gcg?agg?tcg?ctg?tgc?cgc?gtc?gtg 2112
Gly?Leu?Val?Arg?Arg?Ser?Ala?Val?Ala?Arg?Ser?Leu?Cys?Arg?Val?Val
690 695 700
atc?ggc?gtc?gcc?gac?gtg?aag?gtc?cgg?cgc?cag?gag?tgc?cac?aac?gag 2160
Ile?Gly?Val?Ala?Asp?Val?Lys?Val?Arg?Arg?Gln?Glu?Cys?His?Asn?Glu
705 710 715 720
tcc?aag?ctg?tga 2172
Ser?Lys?Leu
<210>3
<21l>723
<212>PRT
＜213〉paddy rice (Oryza sativa)
<400>3
Met?Ser?Lys?Phe?Gln?Ile?Leu?Thr?Arg?Ala?Arg?Trp?Arg?Gln?Asn?Phe
1 5 10 15
Ala?Gly?Glu?Thr?Tyr?Leu?Tyr?Trp?Leu?Arg?Arg?Arg?Pro?Ala?Leu?Tyr
20 25 30
Val?Thr?Asp?Pro?Glu?Leu?Ile?Gly?Glu?Ile?Gly?Arg?Cys?Val?Ser?Leu
35 40 45
Asp?Met?Gly?Lys?Pro?Lys?Tyr?Leu?Gln?Lys?Gly?Gln?Glu?Pro?Leu?Phe
50 55 60
Gly?Gly?Gly?Val?Leu?Lys?Ala?Asn?Gly?Ala?Cys?Trp?Ala?Arg?Gln?Arg
65 70 75 80
Lys?Val?Ile?Ala?Pro?Glu?Phe?Tyr?Met?Ala?Arg?Val?Arg?Ala?Met?Val
85 90 95
Gln?Leu?Met?Val?Asp?Ala?Ala?Gln?Pro?Leu?Ile?Ala?Ser?Trp?Glu?Ser
100 105 110
Arg?Ile?Asp?Ala?Ala?Gly?Gly?Ala?Ala?Ala?Ala?Glu?Val?Val?Val?Asp
115 120 125
Gly?Asp?Leu?Arg?Ser?Phe?Ser?Phe?Asp?Val?Ile?Ser?Arg?Ala?Cys?Phe
130 135 140
Gly?Ser?Asp?Tyr?Ser?Arg?Gly?Arg?Glu?Ile?Phe?Leu?Arg?Leu?Arg?Glu
145 150 155 160
Leu?Ser?Gly?Leu?Met?Ser?Glu?Thr?Ser?Val?Ile?Phe?Ser?Ile?Pro?Ser
165 170 175
Leu?Arg?His?Leu?Pro?Thr?Gly?Lys?Asn?Arg?Arg?Ile?Trp?Arg?Leu?Thr
180 185 190
Gly?Glu?Ile?Arg?Ser?Leu?Ile?Met?Glu?Leu?Val?Arg?Glu?Arg?Arg?Cys
195 200 205
Ala?Ala?Arg?Ala?Ala?Arg?Glu?His?Gly?Gly?Lys?Ala?Ala?Pro?Pro?Ser
210 215 220
Pro?Pro?Glu?Arg?Asp?Phe?Leu?Gly?Ser?Ile?Ile?Glu?Asn?Ser?Gly?Gly
225 230 235 240
Gln?Pro?Arg?Pro?Asp?Asp?Phe?Val?Val?Asp?Asn?Cys?Lys?Asn?Ile?Tyr
245 250 255
Phe?Ala?Gly?His?Glu?Thr?Ser?Ala?Val?Thr?Ala?Thr?Trp?Cys?Leu?Met
260 265 270
Leu?Leu?Ala?Ala?His?Pro?Glu?Trp?Gln?Asp?Arg?Ala?Arg?Ala?Glu?Val
275 280 285
Leu?Glu?Val?Cys?Gly?Gly?Asp?Gly?Ala?Ala?Ala?Pro?Ala?Ala?Pro?Asp
290 295 300
Phe?Asp?Met?Val?Ser?Arg?Met?Arg?Thr?Val?Gly?Met?Val?Val?Gln?Glu
305 310 315 320
Thr?Leu?Arg?Leu?Phe?Pro?Pro?Ser?Ser?Phe?Val?Val?Arg?Glu?Thr?Phe
325 330 335
Arg?Asp?Met?Gln?Leu?Gly?Arg?Leu?Leu?Ala?Pro?Lys?Gly?Thr?Tyr?Leu
340 345 350
Phe?Val?Pro?Val?Ser?Thr?Met?His?His?Asp?Val?Ala?Ala?Trp?Gly?Pro
355 360 365
Thr?Ala?Arg?Leu?Phe?Asp?Pro?Ser?Arg?Phe?Arg?Asp?Gly?Val?Ala?Ala
370 375 380
Ala?Cys?Lys?His?Pro?Gln?Ala?Ser?Phe?Met?Pro?Phe?Gly?Leu?Gly?Ala
385 390 395 400
Arg?Thr?Cys?Leu?Gly?Gln?Asn?Leu?Ala?Leu?Val?Glu?Val?Lys?Thr?Leu
405 410 415
Val?Ala?Val?Val?Leu?Ala?Arg?Phe?Glu?Phe?Thr?Leu?Ser?Pro?Glu?Tyr
420 425 430
Arg?His?Ser?Pro?Ala?Phe?Arg?Leu?Ile?Ile?Glu?Pro?Glu?Phe?Gly?Leu
435 440 445
Arg?Leu?Arg?Ile?Arg?Arg?Ala?Gly?Gly?Leu?Trp?Leu?Ser?Glu?Arg?Pro
450 455 460
Phe?Asp?Trp?Asp?Ala?Asp?Glu?Gln?Lys?Asn?Leu?Asn?Arg?Glu?Ala?Ile
465 470 475 480
Ser?Val?Val?Thr?Tyr?Asp?Ala?Thr?Leu?Gly?Asp?Ser?Val?Ser?Gln?Arg
485 490 495
Val?His?His?Gln?Leu?Ala?Ser?Ser?Ala?Arg?Gln?Glu?Arg?Pro?Ala?Ala
500 505 510
Met?Ser?Gly?Ser?Gly?Asp?Ala?Ala?Pro?Ala?Arg?His?Asn?Ala?Gly?His
515 520 525
Gly?Arg?Arg?Arg?Arg?Arg?Val?Leu?Val?Trp?Ala?Ser?Phe?Ala?Ala?Leu
530 535 540
Val?Leu?Leu?Leu?Val?Ala?Ala?Ala?Ala?Ala?Ile?Ala?Ala?Leu?Ala?Val
545 550 555 560
Leu?Arg?Pro?Arg?Asp?Pro?Thr?Thr?Glu?Leu?Leu?Ser?Val?Asn?Ala?Thr
565 570 575
Gly?Ala?Thr?Pro?Arg?Val?Ala?Ala?Leu?Pro?Ala?Val?Ser?Val?Gln?Leu
580 585 590
Asn?Val?Thr?Phe?Leu?Leu?Val?Val?Arg?Val?Arg?Asn?Pro?Asn?Arg?Ala
595 600 605
Glu?Phe?Arg?His?Gly?Ala?Ala?Thr?Thr?Ala?Leu?Leu?Tyr?Arg?Gly?Ala
610 615 620
Glu?Val?Gly?Ala?Ala?Gly?Val?Pro?Ala?Gly?Thr?Val?Pro?Ser?Arg?Gly
625 630 635 640
Ala?Ala?Thr?Leu?Arg?Leu?Asn?Met?Thr?Val?Arg?Ala?Asp?Arg?Val?Val
645 650 655
Ala?Ala?Ala?Gly?Val?Gly?Gly?Leu?Leu?Ala?Asp?Val?Leu?Ala?Gly?Glu
660 665 670
Met?Glu?Phe?Glu?Ala?Arg?Thr?Glu?Val?Arg?Gly?Arg?Val?Lys?Leu?Leu
675 680 685
Gly?Leu?Val?Arg?Arg?Ser?Ala?Val?Ala?Arg?Ser?Leu?Cys?Arg?Val?Val
690 695 700
Ile?Gly?Val?Ala?Asp?Val?Lys?Val?Arg?Arg?Gln?Glu?Cys?His?Asn?Glu
705 710 715 720
Ser?Lys?Leu
<210>4
<211>25
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
<222>(1)..(25)
＜223〉primer
<400>4
acggaggccc?atgagcaaat?ttcaa 25
<210>5
<211>26
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
<222>(1)..(26)
＜223〉primer
acgtatcaga?gccagtcaca?gcttgg 26
<210>6
<211>19
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
<222>(1)..(19)
＜223〉primer
<400>6
accagcgtca?tcttcagca 19
<210>7
<211>17
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
<222>(1)..(17)
＜223〉primer
<400>7
aacgtctccc?gcaccac 17
Claims (10)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB031293298A CN100393744C (en) | 2003-06-18 | 2003-06-18 | Rice stem elongation gene and its encoded protein and application |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB031293298A CN100393744C (en) | 2003-06-18 | 2003-06-18 | Rice stem elongation gene and its encoded protein and application |
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| Publication Number | Publication Date |
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| CN1566146A true CN1566146A (en) | 2005-01-19 |
| CN100393744C CN100393744C (en) | 2008-06-11 |
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| CNB031293298A Expired - Fee Related CN100393744C (en) | 2003-06-18 | 2003-06-18 | Rice stem elongation gene and its encoded protein and application |
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| CN (1) | CN100393744C (en) |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2006340611A (en) * | 2005-06-07 | 2006-12-21 | Institute Of Physical & Chemical Research | Gibberellin metabolic enzyme |
| WO2009021448A1 (en) * | 2007-08-10 | 2009-02-19 | Shanghai Institutes For Biological Sciences, Chinese Academy Of Sciences | A plant height regulatory gene and uses thereof |
| CN101376674B (en) * | 2007-08-29 | 2012-01-04 | 中国科学院上海生命科学研究院 | Rice flavoprotein gene and use |
| CN102618572A (en) * | 2012-03-21 | 2012-08-01 | 中国农业科学院生物技术研究所 | Method for cultivating drought-enduring plants |
| CN103524608A (en) * | 2013-10-15 | 2014-01-22 | 中国水稻研究所 | Rice spike neck node regulation gene SUI1 (shorted uppermost Internode 1) and application thereof |
| CN105087633A (en) * | 2014-04-29 | 2015-11-25 | 中国科学院上海生命科学研究院 | Gene for regulating plant height, tiller number and leaf inclination and application thereof |
-
2003
- 2003-06-18 CN CNB031293298A patent/CN100393744C/en not_active Expired - Fee Related
Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2006340611A (en) * | 2005-06-07 | 2006-12-21 | Institute Of Physical & Chemical Research | Gibberellin metabolic enzyme |
| WO2009021448A1 (en) * | 2007-08-10 | 2009-02-19 | Shanghai Institutes For Biological Sciences, Chinese Academy Of Sciences | A plant height regulatory gene and uses thereof |
| AU2008286583B2 (en) * | 2007-08-10 | 2012-11-29 | Shanghai Institutes For Biological Sciences, Chinese Academy Of Sciences | A plant height regulatory gene and uses thereof |
| US8461419B2 (en) | 2007-08-10 | 2013-06-11 | Shanghai Institutes For Biological Sciences, Cas | Plant height regulatory gene and uses thereof |
| CN101932596B (en) * | 2007-08-10 | 2016-04-20 | 中国科学院上海生命科学研究院 | The gene of regulating plant plant height and application thereof |
| CN101376674B (en) * | 2007-08-29 | 2012-01-04 | 中国科学院上海生命科学研究院 | Rice flavoprotein gene and use |
| CN102618572A (en) * | 2012-03-21 | 2012-08-01 | 中国农业科学院生物技术研究所 | Method for cultivating drought-enduring plants |
| CN103524608A (en) * | 2013-10-15 | 2014-01-22 | 中国水稻研究所 | Rice spike neck node regulation gene SUI1 (shorted uppermost Internode 1) and application thereof |
| CN103524608B (en) * | 2013-10-15 | 2015-05-13 | 中国水稻研究所 | Rice spike neck node regulation gene SUI1 (shorted uppermost Internode 1) and application thereof |
| CN105087633A (en) * | 2014-04-29 | 2015-11-25 | 中国科学院上海生命科学研究院 | Gene for regulating plant height, tiller number and leaf inclination and application thereof |
| CN105087633B (en) * | 2014-04-29 | 2018-03-27 | 中国科学院上海生命科学研究院 | Adjust gene and its application of plant plant height, tillering number and Leaf inclination |
Also Published As
| Publication number | Publication date |
|---|---|
| CN100393744C (en) | 2008-06-11 |
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