CN113150089A

CN113150089A - Application of GhMKK6 gene and encoding protein thereof in cotton dwarf breeding

Info

Publication number: CN113150089A
Application number: CN202110159252.9A
Authority: CN
Inventors: 王琛; 孟慧; 郭兴启; 王丽君; 赵光栋; 牛晓静
Original assignee: Shandong Agricultural University
Current assignee: Shandong Agricultural University
Priority date: 2021-02-05
Filing date: 2021-02-05
Publication date: 2021-07-23

Abstract

The invention discloses application of a GhMKK6 gene and a protein coded by the gene in cotton dwarfing breeding. The invention provides a protein, named GhMKK6 protein, which is a protein shown as a sequence 1 in a sequence table. The gene coding the GhMKK6 protein also belongs to the protection scope of the invention. The invention also protects the application of the GhMKK6 protein in regulating and controlling the plant height. In the application, the expression of GhMKK6 protein is increased, and the plant height is reduced. The invention also provides a method for cultivating the transgenic plant, which comprises the following steps: the GhMKK6 gene is introduced into a receptor plant to obtain a transgenic plant with reduced plant height. The invention has good application prospect in cotton genetic breeding.

Description

Application of GhMKK6 gene and encoding protein thereof in cotton dwarf breeding

Technical Field

The invention belongs to the technical field of biology, and relates to a GhMKK6 gene and application of a protein coded by the gene in cotton dwarfing breeding.

Background

Cotton is one of the most important economic crops, and products produced by utilizing the cotton relate to the aspects of people's life. In addition, as the population of the world increases, the demand for cotton is further increased. How to improve the cotton yield and promote the development of the cotton industry becomes the key point of scientific research in the field of cotton planting and processing at present.

Cotton has the tendency of unlimited growth, cotton farmers often need to control the height of the main stem of a cotton plant and adjust the transportation direction of substances such as water, nutrients and the like in vivo by a manual topping and castration method in the traditional cotton planting process, so that more nutrients are supplied for reproductive organs to grow, the excessive consumption of water and fertilizer by ineffective fruit branches is reduced, the early boll-forming, multi-boll-forming and less dropping of the cotton plant are promoted, and the quality of the cotton is improved. Research shows that the cotton with short main branches has the characteristics of lodging resistance, high light energy utilization rate, suitability for high-density planting and the like. The manual topping consumes more labor force and has low efficiency, and cotton farmers are heavily burdened. Therefore, if a gene related to cotton growth regulation can be found and used for improving the growth traits of cotton and controlling the growth height of plants, the cost and labor force can be effectively saved, and meanwhile, the mechanized management of cotton can be realized, and the healthy development of the cotton industry can be promoted.

Disclosure of Invention

The invention aims to provide application of a GhMKK6 gene and a protein coded by the gene in cotton dwarf breeding.

The invention provides a protein, named GhMKK6 protein, obtained from Gossypium hirsutum Linn, which is (a1) or (a2) or (a3) or (a4) as follows:

(a1) protein shown as a sequence 1 in a sequence table;

(a2) the protein shown in the sequence 1 in the sequence table is subjected to substitution and/or deletion and/or addition of one or more amino acid residues, and is related to plant height and derived from the protein;

(a3) a fusion protein obtained by attaching a tag to the N-terminus or/and the C-terminus of the protein of (a 1);

(a4) and (b) a protein derived from cotton, having 98% or more identity to (a1) and associated with plant height.

The labels are specifically shown in table 1.

TABLE 1 sequences of tags

The gene coding the GhMKK6 protein also belongs to the protection scope of the invention.

The gene coding the GhMKK6 protein is named as GhMKK6 gene.

The GhMKK6 gene is (b1) or (b2) or (b3) as follows:

(b1) the coding region is a DNA molecule shown as a sequence 2 in a sequence table;

(b2) a DNA molecule that hybridizes under stringent conditions to the DNA molecule defined in (b1) and encodes said protein;

(b3) a DNA molecule derived from cotton and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% homology to the DNA molecule defined in (b1) or (b2) or (b3) and encoding said protein.

The stringent conditions are hybridization and washing of the membrane 2 times 5min at 68 ℃ in a solution of 2 XSSC, 0.1% SDS and 2 times 15min at 68 ℃ in a solution of 0.5 XSSC, 0.1% SDS.

The recombinant vector, expression cassette or recombinant bacterium containing the GhMKK6 gene belong to the protection scope of the invention.

The existing plant expression vector can be used for constructing a recombinant vector containing the GhMKK6 gene.

When constructing a recombinant vector, any one of an enhanced, constitutive, tissue-specific or inducible promoter may be added in front of its transcription initiation nucleotide, and they may be used alone or in combination with other plant promoters. In addition, in constructing recombinant vectors, enhancers, including translational or transcriptional enhancers, may be used, and these enhancer regions may be ATG initiation codons or initiation codons in adjacent regions, etc., but must be in the same reading frame as the coding sequence to ensure proper translation of the entire sequence. The translational control signals and initiation codons are widely derived, either naturally or synthetically. The translation initiation region may be derived from a transcription initiation region or a structural gene. In order to facilitate the identification and screening of transgenic plants, the recombinant vector used may be processed, for example, by adding a gene expressing an enzyme or a luminescent compound which produces a color change in a plant, a gene for a resistant antibiotic marker or a gene for a chemical-resistant marker, etc. From the viewpoint of safety of transgenes, transformed plants can be directly screened for phenotypes without adding any selectable marker gene.

The plant expression vector can be specifically a vector pRI 201-AN.

The recombinant vector may specifically be: a recombinant plasmid obtained by replacing small fragments between multiple cloning sites (such as NdeI and KpnI enzyme cutting sites) of the vector pRI 201-AN with double-stranded DNA molecules shown by 4 th-1065 th nucleotides in a sequence 2 of a sequence table.

The invention also protects the application of the GhMKK6 protein in regulating and controlling the plant height. In the application, the expression of GhMKK6 protein is increased, and the plant height is reduced.

The invention also protects the application of the GhMKK6 gene or a recombinant vector containing the GhMKK6 gene or an expression cassette containing the GhMKK6 gene, and is used for cultivating transgenic plants with reduced plant height.

The invention also provides a method for cultivating the transgenic plant, which comprises the following steps: the GhMKK6 gene is introduced into a receptor plant to obtain a transgenic plant with reduced plant height. The GhMKK6 gene is specifically introduced into a recipient plant through a recombinant vector containing the GhMKK6 gene.

The invention also provides a plant breeding method, which comprises the following steps: the content and/or activity of GhMKK6 protein in the target plant is increased, so that the plant height of the plant is reduced.

Any of the above plants is a monocot or a dicot.

Any of the above plants is a Malvaceae plant.

Any of the above plants is a cotton plant.

Any one of the above plants is upland cotton.

Any of the above plants is cotton.

Any of the above plants is cotton variety (line) R15.

In the invention, an over-expression vector containing the CDS of the GhMKK6 gene is constructed by an inventor, and a stably inherited GhMKK6 gene over-expression cotton strain is obtained by an agrobacterium-mediated hypocotyl transformation method. Transgenic cotton plants overexpressing the GhMKK6 gene have a significantly dwarf phenotype in greenhouse and field planting. Cell morphology analysis shows that the number of cells in the leaves of transgenic plants over expressing the GhMKK6 gene is obviously reduced. The observation of an optical microscope and a transmission electron microscope shows that the stem tip growing point sub-microstructure in the transgenic plant over-expressing the GhMKK6 gene is not changed, but the cell number of the meristematic region is obviously reduced. The results show that the GhMKK6 protein can regulate the growth and development of cotton by influencing the cell number of meristematic regions. The over-expression GhMKK6 gene can shorten the main branch of cotton and has excellent application foreground in cotton genetic breeding.

Drawings

FIG. 1 is a photograph of a plant after 3 weeks of cultivation in a greenhouse.

FIG. 2 shows the statistical results of plant heights after 3 weeks of cultivation in a greenhouse.

FIG. 3 is a photograph of a plant cultured in a field for 2 months.

FIG. 4 is a photograph showing leaf cells observed under a microscope.

FIG. 5 is a statistical image of the number of microscopic cells per unit area of leaf.

FIG. 6 is a photograph under an optical microscope of a growing point of a stem tip.

FIG. 7 is a photograph of the growing point of the stem tip under a transmission electron microscope.

Detailed Description

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

The experimental procedures in the following examples, unless otherwise indicated, are conventional and are carried out according to the techniques or conditions described in the literature in the field or according to the instructions of the products. Materials, reagents and the like used in the following examples are commercially available unless otherwise specified. The wild-type cotton used in the examples refers to cotton variety (line) R15, also known as upland cotton R15, denoted by WT. The cotton GhMKK6 protein is shown as a sequence 1 in a sequence table. The gene encoding the GhMKK6 protein was designated as GhMKK6 gene. In the cotton cDNA, the open reading frame of the GhMKK6 gene is shown as a sequence 2 in a sequence table.

Examples 1,

Construction of recombinant plasmid

1. Taking a double-stranded DNA molecule shown in a sequence 2 of the sequence table as a template, adopting a primer pair consisting of 2K6-PRI-5 and 2K6-PRI-3 to carry out PCR amplification, and recovering a PCR amplification product.

2K6-PRI-5：5’-CATATGAAGAGCAAGAAGCCATTGAAGC-3’；

2K6-PRI-3：5’-GGTACCTTATCTTGGGTAATTCACAGG-3’。

2. Taking the PCR amplification product obtained in the step 1, carrying out double enzyme digestion by using restriction enzymes NdeI and KpnI, and recovering the enzyme digestion product.

3. Taking the vector pRI 201-AN, carrying out double enzyme digestion by using restriction enzymes NdeI and KpnI, and recovering a vector skeleton of about 10000 bp.

4. And (3) connecting the enzyme digestion product obtained in the step (2) with the vector skeleton obtained in the step (3) to obtain the recombinant plasmid pRI 201-GhMKK 6. According to the sequencing result, the structure of the recombinant plasmid pRI 201-GhMKK6 is described as follows: the small fragment between NdeI and KpnI cleavage sites of vector pRI 201-AN was substituted for the double-stranded DNA molecule shown as nucleotides 4 to 1065 in sequence 2 of the sequence listing.

Secondly, preparing transgenic plants

1. Introducing the recombinant plasmid pRI 201-GhMKK6 into agrobacterium LBA4404 to obtain recombinant agrobacterium; inoculating the recombinant Agrobacterium single colony to LB liquid medium containing 100.0mg/L kanamycin and 50.0mg/L rifampicin, performing shake culture at 28.0 deg.C and 180-₆₀₀After centrifugation at 3600rpm at 28.0 ℃ for 10min, the pellet was collected, suspended in an equal volume of 1/2MS liquid medium, and cultured with shaking at 160rpm at 28.0 ℃ for 30 min.

2. Taking mature seed of wild type cotton, stripping intact kernel with scissors, and aseptically applying 0.1% HgCl onto kernel₂Soaking in water solution for 4-5min, washing with sterilized double distilled water for 3-5 times, and placing the kernel in a container filled with MS₀In tubes of medium (2 kernels per tube), culture was performed for one week. The culture conditions are as follows: 27. + -. 1 ℃ with 16h light/8 h dark.

MS₀Culture medium: containing 2.2g/L MS, 30g/L sucrose, 5.5g/L agar, by ddH₂O, fixing the volume; pH 6.0.

3. And (3) after the step 2 is finished, taking off the hypocotyls by using forceps under an aseptic condition, and cutting into 0.5cm small sections by using a scalpel, namely, the hypocotyl cutting sections.

4. And (3) under an aseptic condition, immersing the hypocotyl cut section obtained in the step (3) into the bacterial liquid obtained in the step (1), slightly shaking for 3min, filtering out the bacterial liquid, then, sucking the redundant bacterial liquid on the hypocotyl cut section by using aseptic filter paper, then, flatly paving the hypocotyl cut section on a co-culture medium, and culturing for 2 days. The culture conditions are as follows: 28.0 ℃ in the dark.

Co-culture medium: contains 4.4g/L MS, 30g/L glucose, 0.1mg/L KT, 0.1 mg/L2, 4-D, 2.0g/L MgCl₂·6H₂O, 2.0g/L plant gel with ddH₂O, fixing the volume; pH 6.0.

5. After completion of step 4, hypocotyl sections were transferred to resistant callus induction medium and cultured for about 3 weeks. The culture conditions are as follows: 27. + -. 1 ℃ with 16h light/8 h dark. In the culture process, callus is generated at two ends of the hypocotyl, and when the culture is finished, the callus grows to be about 2-3cm in diameter.

Resistant callus induction medium: contains 4.4g/L MS, 30g/L glucose, 0.1mg/L KT, 0.1 mg/L2, 4-D, 2.0g/L MgCl₂·6H₂O, 500mg/L cefuroxime, 50mg/L kanamycin, 2.0g/L plant gel, and ddH₂O, fixing the volume; pH 6.0.

6. And (5) after the step 5 is completed, cutting the callus, inoculating the callus to a resistant callus induction culture medium, and culturing until the diameter of the callus block reaches 7-8 cm. The culture conditions are as follows: 27. + -. 1 ℃ with 16h light/8 h dark.

7. After step 6, transferring the callus blocks to an embryogenic callus induction culture medium, and culturing until yellow green or gray green rice-shaped embryogenic callus appears. In the culture process, the same culture medium is adopted for subculture once every month, and subculture is carried out for about 2-6 times. The culture conditions are as follows: 27. + -. 1 ℃ with 16h light/8 h dark.

Embryogenic callus induction medium: contains 4.4g/L MS, 30g/L glucose, 1.9g/L KNO₃、2.0g/L MgCl₂·6H₂O, 2.5g/L plant gel with ddH₂O, fixing the volume; pH 6.0.

8. After step 7, the embryogenic callus was transferred to differentiation medium and cultured until plants were formed. In the culture process, subculture is carried out every 3-4 weeks. The culture conditions are as follows: 27. + -. 1 ℃ with 16h light/8 h dark. During the culture process, it can be observed that the embryogenic callus forms embryoid bodies first and then plants are formed.

Differentiation medium: contains 30g/L glucose and 1.9g/L KNO₃0.5g/L asparagine, 1.0g/L glutamine, 2.0g/L MgCl₂·6H₂O, 2.5g/L MSB-NH of plant gel₂NO₃Medium, pH 6.0.

MSB-NH₂NO₃The medium formulation is shown in Table 2.

TABLE 2

9. Plants which develop completely and have developed root systems in the step 8 (namely T)₀Regenerated plant) is transferred to a flowerpot filled with culture medium, and is transplanted to a field after the plant grows well.

10. From T₀And (4) screening transgenic plants from the generation plants.

11、T₀Selfing the transgenic plant and harvesting the seeds to obtain T₁Seed generation, T₁The plant grown by the seed generation is T₁And (5) plant generation.

12. From T₁And (4) screening transgenic plants from the generation plants.

13、T₁Selfing the transgenic plant and harvesting the seeds to obtain T₂Seed generation, T₂The plant grown by the seed generation is T₂And (5) plant generation.

14. From T₂And (4) screening transgenic plants from the generation plants.

15. Two plants T₂The transgenic plants were named OE2 plant and OE3 plant, respectively.

The method for screening transgenic plants in the steps 10, 12 and 14 is as follows: extracting total RNA and carrying out reverse transcription to obtain cDNA, carrying out real-time quantitative PCR by adopting a primer pair consisting of 2K6-5 and 2K6-3, and judging the plants with the relative abundance of target genes more than 100 times that of wild plants as transgenic plants. Transgenic plants are also known as over-expressed plants.

2K6-5：CCTATCTCTTCTTTCTTGACGGCG；

2K6-3：CTTCAAGGCAAACAATCTTCCAACC。

Third, plant height identification

Test seeds: wild type cotton seeds, seeds from selfing of OE2 plants, and seeds from selfing of OE3 plants.

1. The test seeds were sown in pots filled with moist soil and cultured in a greenhouse.

The culture conditions are as follows: 27 +/-1 ℃; the photoperiod is 16h of light/8 h of dark; the relative humidity is 60-75%.

After the seeds emerge, screening transgenic plants (the same method as the second step).

The photograph after 3 weeks of culture is shown in FIG. 1.

The statistical results of plant height are shown in FIG. 2 (ordinate unit is cm) (3 independent repeated experiments, each time, the average value of the plant height of 15 cotton plants is counted).

In FIGS. 1 and 2, WT represents a plant grown from wild-type cotton seed, OE2 represents a transgenic plant grown from seed obtained by selfing an OE2 plant, and OE3 represents a transgenic plant grown from seed obtained by selfing an OE3 plant.

2. And (4) sowing the test seeds in the field, and carrying out normal cultivation management.

The photograph of the plant 2 months after sowing is shown in FIG. 3.

In FIG. 3, WT represents a plant grown from wild-type cotton seed, OE2 represents a transgenic plant grown from seed obtained by selfing an OE2 plant, and OE3 represents a transgenic plant grown from seed obtained by selfing an OE3 plant.

The results of step 1 and step 2 indicate that the transgenic plants have a significantly dwarf phenotype compared to wild-type cotton plants.

Fourth, true leaf cell number identification

Test leaves: and step three, true leaves of wild cotton plants cultured for 3 weeks in step three 1, true leaves of transgenic plants grown from seeds obtained by selfing of OE2 plants cultured for 3 weeks in step three 1, and true leaves of transgenic plants grown from seeds obtained by selfing of OE3 plants cultured for 3 weeks in step three 1. Selecting true leaves with similar growth states.

Data are presented as mean standard deviation of 3 independent experiments, each of which contained 15 independent plants as biological replicates. Fixing liquid: ethanol, lactic acid, glycerol as 3: 1: mixing at a volume ratio of 1.

And (3) decoloring: (1) placing cotton true leaf in prepared stationary liquid heated to boiling, and treating for 10 min; (2) transferring the cotton true leaves completing the step (1) into a fresh fixing solution, and treating at room temperature overnight.

And (3) microscopic observation: and (4) taking decolorized cotton true leaves, flaking, and then placing under a microscope for observation.

The photograph is shown in FIG. 4 (scale bar 50 μm).

A statistical map of the number of microscopic cells per unit area is shown in FIG. 5.

In FIGS. 4 and 5, WT represents a plant grown from wild-type cotton seed, OE2 represents a transgenic plant grown from seed obtained by selfing an OE2 plant, and OE3 represents a transgenic plant grown from seed obtained by selfing an OE3 plant.

The results show that the number of cells in the leaves of transgenic cotton plants is significantly reduced compared to wild-type cotton plants.

Morphological observation of growing points of stem tips

Test stem tip: and step three, culturing the stem tip of the wild-type cotton plant for 3 weeks in step three 1, culturing the stem tip of the transgenic plant grown from the seed obtained by selfing the OE2 plant for 3 weeks in step three 1, and culturing the stem tip of the transgenic plant grown from the seed obtained by selfing the OE3 plant for 3 weeks in step three 1.

Fixing: firstly, immersing the stem tip into pre-cooled 4% glutaraldehyde, vacuumizing for 30min, and then putting the stem tip in a refrigerator at 4 ℃ overnight; the following day, washing with phosphate buffer 5 times (4 deg.C), 30min each time; the stem tips were then fixed in a 1% osmic acid fixative for 5h and then washed 5 times with phosphate buffer (4 ℃ C.) for 30min each.

And (3) dehydrating: after the fixation is finished, the stem tip is sequentially immersed into 45%, 55%, 70%, 85%, 95% and 100% ethanol for dehydration, and each stage of treatment lasts for 1 hour; then dehydrated for 4h by 100% ethanol, and repeated once.

And (3) transparency: after dehydration, the stem tips were transferred to propylene oxide solution for 2 treatments, each for 4-7 h.

Resin impregnation: after the transparency is finished, taking the stem tip, and firstly using resin: the propylene oxide (1:1) mixture was treated overnight, then with the resin for 12h, repeated once, and finally with the addition of the resin containing DMP-30(2, 4, 6-triaminophenol) and treated overnight.

Embedding: after resin impregnation is completed, embedding the material on an embedding plate, and sequentially placing the embedded material in a 36 ℃ oven for drying for 12h, a 45 ℃ oven for drying for 12h and a 60 ℃ oven for drying for 36 h.

Slicing: the sections were sliced with a LEICARM2265 microtome to a thickness of 2 μm, mounted on a mounting table at 42 ℃ and stained with toluidine blue for 3-4 minutes.

The development condition of the growing point of the cotton stem tip is observed by using an optical microscope and a transmission electron microscope (JEM-1200EX), respectively.

The photograph under the optical microscope is shown in FIG. 6. The number of cells in the meristematic region of the transgenic cotton plant is significantly reduced compared to wild-type cotton plants.

The transmission electron micrograph is shown in FIG. 7. The cellular sub-microstructure of the meristematic region of the growing point of the transgenic cotton plant is not altered compared to the wild type cotton plant.

In FIGS. 6 and 7, WT represents a plant grown from wild-type cotton seed, OE2 represents a transgenic plant grown from seed obtained by selfing an OE2 plant, and OE3 represents a transgenic plant grown from seed obtained by selfing an OE3 plant.

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

Sequence listing

<110> Shandong university of agriculture

<120> GhMKK6 gene and application of encoding protein thereof in cotton dwarf breeding

<130> GNCYX210541

<160> 2

<170> SIPOSequenceListing 1.0

<210> 1

<211> 354

<212> PRT

<213> Gossypium hirsutum Linn.

<400> 1

Met Lys Ser Lys Lys Pro Leu Lys Gln Leu Lys Leu Ala Val Pro Ala

1 5 10 15

Gln Glu Thr Pro Ile Ser Ser Phe Leu Thr Ala Ser Gly Thr Phe His

20 25 30

Asp Gly Asp Leu Leu Leu Asn Gln Lys Gly Leu Arg Leu Ile Ser Glu

35 40 45

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

50 55 60

Ser Leu Glu Asp Leu Glu Thr Ile Lys Val Ile Gly Lys Gly Ser Gly

65 70 75 80

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

85 90 95

Leu Lys Val Ile Gln Met Asn Ile Gln Glu Glu Ile Arg Lys Gln Ile

100 105 110

Val Gln Glu Leu Lys Ile Asn Gln Ala Ser Gln Cys Ser His Val Val

115 120 125

Val Cys Tyr His Ser Phe Tyr His Asn Gly Ala Ile Ser Leu Val Leu

130 135 140

Glu Tyr Met Asp Arg Gly Ser Leu Ala Asp Val Ile Arg Gln Val Asn

145 150 155 160

Thr Ile Leu Glu Pro Tyr Leu Ala Val Val Cys Lys Gln Val Leu Gln

165 170 175

Gly Leu Val Tyr Leu His His Glu Arg His Val Ile His Arg Asp Ile

180 185 190

Lys Pro Ser Asn Leu Leu Val Asn His Lys Gly Glu Val Lys Ile Thr

195 200 205

Asp Phe Gly Val Ser Ala Met Leu Ala Ser Ser Met Gly Gln Arg Asp

210 215 220

Thr Phe Val Gly Thr Tyr Asn Tyr Met Ser Pro Glu Arg Ile Ser Gly

225 230 235 240

Ser Thr Tyr Asp Tyr Ser Ser Asp Ile Trp Ser Leu Gly Met Val Val

245 250 255

Leu Glu Cys Ala Ile Gly Arg Phe Pro Tyr Met Gln Ser Glu Asp Gln

260 265 270

Gln Ser Trp Pro Ser Phe Tyr Glu Leu Leu Glu Ala Ile Val Glu Lys

275 280 285

Pro Pro Pro Thr Ala Pro Ser Asp Gln Phe Ser Pro Glu Phe Cys Ser

290 295 300

Phe Val Ser Ala Cys Ile Lys Lys Asn Pro Lys Glu Arg Ala Ser Ser

305 310 315 320

Leu Asp Leu Leu Ser His Pro Phe Ile Arg Lys Phe Glu Asp Lys Asp

325 330 335

Ile Asp Leu Gly Ile Leu Val Gly Ser Leu Glu Pro Pro Val Asn Tyr

340 345 350

Pro Arg

<210> 2

<211> 1065

<212> DNA

<213> Gossypium hirsutum Linn.

<400> 2

atgaagagca agaagccatt gaagcaactg aagctcgctg ttccagctca agaaacccct 60

atctcttctt tcttgacggc gagtgggaca tttcatgatg gcgatttgct tctaaatcag 120

aaaggattgc gtcttatttc cgaggaaaag gaatctcgac cttctgacgc caaggagctt 180

gattttgaat tctcattgga agacctagag acaatcaaag ttattgggaa gggcagtggt 240

ggtgtagtac aacttgttcg ccacaaatgg gttggaagat tgtttgcctt gaaggtcatc 300

caaatgaaca tacaagaaga aattcgcaag caaattgtgc aggagctaaa aataaaccaa 360

gcatcacaat gttcgcatgt tgtagtttgc taccattctt tctatcacaa tggggccata 420

tctctggtgc tagaatacat ggaccgtgga tctctggccg atgtgatcag acaagttaac 480

acaattcttg aaccatatct tgcagttgtg tgtaagcagg ttctacaggg acttgtgtat 540

ttgcaccatg aaaggcacgt aatacatagg gacataaaac catccaatct actggtaaac 600

cataaagggg aagtgaagat cactgatttt ggtgtcagtg caatgctagc tagctctatg 660

ggccagagag atacatttgt tgggacttac aactacatgt cgccggagag gattagtggg 720

agcacttatg actatagcag tgatatttgg agtttgggaa tggtagtgct tgaatgtgct 780

attggacgtt tcccatatat gcaatctgaa gatcaacaaa gctggcctag cttttatgag 840

cttttggagg caatagtgga aaagcctcca ccaactgctc catcagatca attctctcca 900

gagttctgtt catttgtatc agcctgcata aagaagaacc ctaaagaaag agcatcatct 960

ttggacctct tgagtcaccc tttcatcaga aagttcgaag acaaggacat agaccttggg 1020

attttggtag gtagcttgga acctcctgtg aattacccaa gataa 1065

Claims

1. A protein which is (a1) or (a2) or (a3) or (a4) as follows:

(a1) protein shown as a sequence 1 in a sequence table;

(a2) the protein shown in the sequence 1 in the sequence table is subjected to substitution and/or deletion and/or addition of one or more amino acid residues, and is related to plant height and derived from the plant height;

2. A gene encoding the protein of claim 1.

3. The gene of claim 2, wherein: the genes are (b1) or (b2) or (b3) as follows:

(b3) a DNA molecule derived from cotton and encoding said protein and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% homology to the DNA molecule defined in (b1) or (b2) or (b 3).

4. A recombinant vector, expression cassette or recombinant bacterium comprising the gene of claim 2 or 3.

5. Use of the protein of claim 1 for regulating plant height.

6. Use of the gene according to claim 2 or 3 or the recombinant vector according to claim 4 or the expression cassette according to claim 4 for the production of transgenic plants with reduced plant height.

7. A method of breeding a transgenic plant comprising the steps of: a transgenic plant having a reduced plant height is obtained by introducing the gene according to claim 2 or 3 into a recipient plant.

8. A method of plant breeding comprising the steps of: increasing the content and/or activity of the protein of claim 1 in the target plant, thereby reducing the plant height.

9. The use according to claim 5 or 6 or the method according to claim 7 or 8, characterized in that: the plant is a monocotyledon or a dicotyledon.