CN112899292B

CN112899292B - Upland cotton plant height regulating gene GhGA20ox6 and its use

Info

Publication number: CN112899292B
Application number: CN202110162359.9A
Authority: CN
Inventors: 喻树迅; 冯震; 李黎贝; 杨炳磊; 刘林
Original assignee: Zhejiang A&F University ZAFU
Current assignee: Zhejiang A&F University ZAFU
Priority date: 2021-02-05
Filing date: 2021-02-05
Publication date: 2022-10-25
Anticipated expiration: 2041-02-05
Also published as: CN112899292A

Abstract

The invention discloses a upland cotton plant height regulating gene GhGA20ox6 and application thereof, wherein the gene has the nucleotide sequence shown in SEQ ID NO: 1. By regulating GhGA20ox6, the plant height of upland cotton can be regulated and controlled on a molecular level, so that upland cotton can be improved, plants can be dwarfed, upland cotton has strong lodging resistance, the population light energy utilization rate of crops is improved, and the yield of the crops is increased.

Description

Upland cotton plant height regulating gene GhGA20ox6 and its use

Technical Field

The invention relates to the technical field of genetic engineering, in particular to a gossypium hirsutum plant height regulating gene GhGA20ox6 and application thereof.

Background

Gibberellin (GA), one of six plant hormones, is a tetracyclic diterpene phytohormone, plays an important role in each growth and development stage of plants, and plays an important role in seed germination, root cap elongation, flowering and fruiting, stem elongation, leaf growth and development and the like. In higher plants, various enzymes are involved in the biosynthesis and metabolism of gibberellin through a multi-step enzymatic reaction, and the expression of these enzyme genes is effective in regulating the anabolic rate of gibberellin. There are many enzymes involved in the anabolism of gibberellin, and GA2-oxidase (GA 2-oxidase) is a key enzyme in the degradation of GA in plants, and GA2ox decomposes and inactivates GAs active in plants and its precursors and intermediates, thereby maintaining the homeostasis between the active GAs in plants. GA3-oxidase (GA 3-oxidase) and GA20-oxidase (GA 20-oxidase) as key enzymes in the last step of plant gibberellin anabolic pathway to catalyze GA ₁₂ And GA ₅₃ Conversion to active GA ₁ And GA ₄ In addition, GA3ox and GA20ox in turn regulate the negative feedback of active GAs within the receptor. Plant gibberellin oxidase gene family genusIn the 2OG-Fe (II) oxidative subfamily, it can be encoded by a multigene family. To date, the gibberellin synthesis and metabolic pathways of the model plant Arabidopsis thaliana have been well studied, and gibberellin-related genes have also been cloned and identified. Researches show that the 'green revolution' of the current main crops has close relationship with gibberellin. In plants, gibberellin participates in regulating the growth and development of plants to have the advantages of promoting stem elongation and plant height, for example, in rice, through cloning and gene silencing of GA20ox related genes, the genes are found to influence the stem length of the rice, and further the rice is caused to have a dwarf phenotype, and researches of Mauriat and the like find that the GA20ox related genes influence the stem elongation of populus tremula, GA20ox related gene expression is enhanced on populus tremula seedlings, and the gibberellin is shown to promote synthesis of active gibberellin in seedlings.

Cotton, the most important natural fiber source in the world, is one of the most widely cultivated cotton species in the world and remains an important commercial crop in many countries. The plant height is an important excellent agronomic character in crop breeding, and the lodging phenomenon can be generated when the plant is too high, so that the yield of the crops is reduced, the dwarfing of the plant can have stronger lodging resistance, the group light energy utilization rate of the crops can be improved, and the yield of the crops can be improved. Upland cotton (g.hirsutum l., AADD) has two subgenomic groups of a and D, and at present, the plant height is basically controlled by castration and topping in agriculture, which increases labor force and working time. The improvement of the upland cotton is of great significance from the molecular level.

Disclosure of Invention

The invention aims to provide an upland cotton plant height regulating gene GhGA20ox6 and application thereof, wherein the upland cotton plant height can be regulated and controlled from a molecular level by regulating the GhGA20ox6, so that upland cotton is improved and dwarfed, the upland cotton has stronger lodging resistance, the group light energy utilization rate of crops is improved, and the yield of the crops is increased.

The technical scheme adopted by the invention for solving the technical problem is as follows:

a upland cotton plant height regulating gene GhGA20ox6, which has the nucleotide sequence shown in SEQ ID NO: 1.

The inventor discovers and determines a key gene GhGA20ox6 for regulating the height of the gossypium hirsutum plant through research and research, screens 13 candidate genes for the first time on the whole gossypium hirsutum genome, and then further searches and analyzes, and verifies the function of the key gene GhGA20ox6. The discovered functional gene is utilized to regulate the plant height of upland cotton from molecular level, so as to improve upland cotton germplasm resources.

A plasmid comprising the upland cotton plant height-regulating gene GhGA20ox6 according to claim 1.

A plant expression vector comprising the upland cotton plant height regulatory gene GhGA20ox6 of claim 1.

A host cell comprising the Gossypium hirsutum plant height regulator GhGA20ox6 of claim 1.

A protein coded by a gossypium hirsutum plant height regulatory gene GhGA20ox6, which has the amino acid sequence shown in SEQ ID NO:14, or a pharmaceutically acceptable salt thereof.

An application of a Gossypium hirsutum plant height regulating gene GhGA20ox6 in regulating the Gossypium hirsutum plant height.

A method for dwarfing upland cotton is characterized by that the upland cotton plant height regulating gene GhGA20ox6 is gene-silenced so as to reduce upland cotton plant height and implement dwarfing.

The invention has the beneficial effects that: by regulating GhGA20ox6, the plant height of upland cotton can be regulated and controlled on a molecular level, so that upland cotton can be improved, plants can be dwarfed, upland cotton has strong lodging resistance, the population light energy utilization rate of crops is improved, and the yield of the crops is increased.

Drawings

FIG. 1 shows the expression levels of the GhGA20ox6 gene in different tissues of Gossypium hirsutum.

FIG. 2 is the subcellular localization of GhGA20ox6 in Nicotiana benthamiana; 35S, ghGA20ox6, GFP (A-C), 35S, GFP (D-F) subcellular localization in Nicotiana benthamiana leaf cells. A. D: epidermal cells under 488nm laser; B. e: epidermal cells under bright field; C. f: laser and bright field combined epidermal cells, 20 μm scale.

FIG. 3 is a positive control (CLCrVA:: PDS), VIGS plant (Line 1, line2, and Line 3), and empty vector control (CLCrVA) phenotype.

FIG. 4 shows plant height and internode length of VIGS positive plants and control group; * P <0.05,. P <0.01.

FIG. 5 shows the relative expression level of GhGA20ox6 in different internodes between VIGS-positive strain and control group; * P <0.05, P <0.01.

FIG. 6 is the phenotype of wild-type Arabidopsis and GhGA20ox6 transgenic Arabidopsis. Note: a is Col-0 and positive strain rosette leaf phenotype; B. c is Col-0 and the height phenotype of the positive strain.

Fig. 7 is the expression level of GhGA20ox6 in transgenic arabidopsis thaliana, { P } 0.01; the reference gene is AtUBQ10.

Detailed Description

The technical solution of the present invention will be further specifically described below by way of specific examples.

In the present invention, the raw materials and equipment used are commercially available or commonly used in the art, unless otherwise specified. The methods in the following examples are conventional in the art unless otherwise specified.

The embodiment is as follows:

1 materials and methods

1.1 Experimental materials and treatment

Upland cotton (G.hirsutum L., AADD) TM-1 material is selected for analyzing tissue expression quantity, planted in a greenhouse of Pingshan base of Zhejiang agriculture and forestry university (Hangzhou city Lingan area in Zhejiang province), and managed by adopting a conventional field mode.

The Nicotiana benthamiana is selected for gene transient transformation, planted in 16 climate rooms of Zhejiang agriculture and forestry university, and under the environmental conditions: the illumination is 16h, and the darkness is 8h; the temperature is 23 ℃; the relative humidity is 70-75%.

Selecting upland cotton TM-1 material to perform VIGS phenotype statistics and gene expression analysis, planting in 16 climate rooms of Zhejiang agriculture and forestry university, and performing temperature conditions: the light is irradiated for 16 hours and the dark is irradiated for 8 hours; the temperature is 24 ℃; the relative humidity is 65-70%.

A transgenic overexpression experiment is carried out by utilizing Arabidopsis thaliana (Arabidopsis thaliana) col wild type material, and the wild type material is planted in a 16-climate chamber of Zhejiang university of agriculture and forestry under the temperature conditions: the temperature is 25 ℃; the illumination is 16h, and the darkness is 8h; the relative humidity is 65-70%.

The materials used are all from cotton breeding laboratories of agriculture and forestry university in Zhejiang.

1.2 vectors and reagents used

The transgenic expression vectors pBI121, pBINGFP4, and the cotton leaf-winkle virus vectors pCLCrVA (empty vector), pCLCrVB (auxiliary vector), and pCLCrVA-PDS (positive control vector) were stored in this laboratory. Escherichia coli DH 5. Alpha. And Agrobacterium tumefaciens strain GV3101 were purchased from Shanghai Diego Limited, inc. Restriction enzymes XbaI, speI and SacI were purchased from TaKaRa (China, dalian) and AscI was purchased from NEB. The RNA extraction kit and the plasmid extraction kit are purchased from Tiangen Biotechnology (Beijing) Co., ltd. Gene cloning vector pMD18-T, mightyAmp Hi-Fi enzyme, primeScript ^TM RT reagent Kit with gDNA Eraser reverse transcription Kit, DNA fragment purification Kit and fluorescent reagent TB

Premix Ex Taq ^TM II purchased from TaKaRa, inc. (Dalian, china). 2-morpholinoethanesulfonic acid, acetosyringone and 1/2MS solid medium were purchased from Yuanye Biotech, inc., and the synthesis of primers and gene sequencing were performed by Hangzhou Youkang Biotechnology, inc. The sequences of the primers used in this experiment are shown in Table 1.

Primer sequences used in Table 1

Note: the restriction sites are underlined.

1.3 Extraction of RNA and Synthesis of cDNA

According to the instruction of RNAprep Pure polysaccharide polyphenol plant total RNA extraction kit (DP 441), 9 tissue RNAs of TM-1, namely 15DPA fibers, ovules, flowers, sepals, terminal buds, stems, leaves, pistils and stamens, are extracted, and the tissue RNAs are subjected to reverse transcription by using a reverse transcription kit to synthesize single-stranded cDNA.

1.4 Cloning of GhGA20ox6 Gene

Using the published protein sequence of Arabidopsis gibberellin 20 oxidase as a reference, the upland cotton genome database released by Huazhong university of agriculture is downloaded from (A), (B)https://cottonfgd.org/) And constructing a local Blast database, and screening to obtain 13 candidate genes, wherein the 13 candidate genes are respectively as follows: ghGA20ox1 (SEQ ID NO: 2), ghGA20ox3 (SEQ ID NO: 3), ghGA20ox4 (SEQ ID NO: 4), ghGA20ox5 (SEQ ID NO: 5), ghGA20ox6 (SEQ ID NO: 1), ghGA20ox7 (SEQ ID NO: 7), ghGA20ox8 (SEQ ID NO: 8), ghGA20ox9 (SEQ ID NO: 9), ghGA20ox10 (SEQ ID NO: 10), ghGA20ox11 (SEQ ID NO: 11), ghGA20ox12 (SEQ ID NO: 12), ghGA20ox13 (SEQ ID NO: 13).

Through further exploration tests, a GhGA20ox6 gene with high expression level in a stem is finally obtained, specific primers GhGA20ox6-F and GhGA20ox6-R are designed, cDNA of a stem tissue of a TM-1 material is used as a template for amplification of the GhGA20ox6, the obtained fragment is connected to a pMD18-T vector, the recombinant plasmid is transformed into escherichia coli DH5 alpha, and a monoclonal bacterial liquid with correct sequencing is stored for subsequent laboratories.

Bioinformatics analysis of 1.5GhGA20Ox6

The obtained GhGA20ox6 protein sequence was processed with ExPASy (C: (A))https://web.expasy.org/ protparam/) The online analysis website predicts the physicochemical properties of the protein, including the length, isoelectric point and molecular mass of the protein. The amino acid sequence of the target gene was analyzed for conservation using DNAMAN. The conserved domain of GhGA20ox6 was obtained using SMART online website.

1.6 GhGA20ox6 spatial pattern expression

According to TB

Premix Ex Taq ^TM Reaction system of II kit and corresponding program for real-time fluorescence quantificationPCR (Real-time quantitative PCR, RT-PCR) with the instrument model of Roche LightCycler 96 and the operation method according to the instrument instruction. The quantitative primers used are GhGA20ox6-qF and GhGA20ox6-qR, and an Actin7 gene is selected as a reference gene of cotton. By use of 2 ^-△△Ct The method analyzes the relative expression quantity of the gene, ensures 3 times of biological and technical repetition, and analyzes and draws a gene relative expression graph by using Excel and Graphpad software.

1.7 Subcellular localization of GhGA20ox6

The target gene fragment is connected to a GFP fluorescent protein vector by using a pair of primers GhGA20ox6-SF and GhGA20ox6-SR, and the constructed over-expression vector 35S, ghGA20ox6, GFP is transformed into an agrobacterium strain GV3101 by using an agrobacterium transformation method. According to the Nicotiana benthamiana transient transformation method, the Agrobacterium liquid is injected into tobacco leaves cultured for 20 days. And (3) carrying out dark treatment on the infected tobacco leaves for 10 hours, and observing GFP cell position signals in the tobacco leaves under an LSM880 confocal microscope after culturing for 28 hours.

1.8 construction and infection of Virus-induced Gene silencing Vector (VIGS)

And (3) putting full seeds of the TM-1 material on the seedling raising blocks until two cotyledons are completely opened. The enzyme cutting primer GhGA20ox6-VF and GhGA20ox6-VR is used for recombining partial gene fragment of GhGA20ox6 with pCLCrVA virus vector, the constructed vectors pCLCrVA-GhGA20ox6, pCLCrVA-pCLCrVB and PDS-pCLCrVB are transferred into agrobacterium CV3101, and the agrobacterium liquid is injected into the cotyledons with consistent growth vigor, dark treatment is carried out for 24 hours, and then the cotyledons are placed into a climate chamber for normal culture. Observing and recording the change between six leaf stages and the total length, taking stem tissues to detect the gene expression amount, and verifying the silencing effect.

1.9 transgenic Arabidopsis overexpression

Transferring the pBI121-GhGA20ox6 overexpression vector with correct sequencing into the GV3101 agrobacterium strain for amplification, converting colarabidopsis thaliana with pod-bearing siliques cut off by using an agrobacterium-floral dip method, and performing dark treatment for 20 hours in a climatic chamber to obtain seeds T ₀ And (5) seed generation. Will T ₀ Subculturing the seed untilPure and generational. Observing and recording the flowering period and height difference of the transgenic arabidopsis thaliana of the homozygous generation, and detecting the gene expression quantity of the GhGA20ox6 in the transgenic arabidopsis thaliana.

2 results and analysis

2.1 Sequence analysis of GhGA20ox6 Gene

GhGA20ox6 is located on chromosome 9 of subgroup D, and has an open reading frame of 1155bp, an amino acid sequence length of 384 (SEQ ID NO: 14), 2 introns and 3 exons. The relative molecular mass of the GhGA20ox6 protein sequence is predicted to be 42.32kDa by using ExPASy online software, and the isoelectric point is 6.37. The DIOX _ N domain is located in the amino acid sequence from 63 th to 172 th, and the 2OG-Fe (II) oxidase domain is located in the amino acid sequence from 228 th to 323 th. The amino acid sequence analysis of the proteins corresponding to the 13 candidate genes shows that the proteins corresponding to the 13 candidate genes including the GhGA20ox6 have a common conserved protein sequence: the highly conserved substrate binding motif LPWKET, the conserved sequence NYPXCQKP associated with binding 2-oxoglutarate, all have Fe ²⁺ A binding site.

2.2 Analysis of spatial expression Pattern of GhGA20ox6 Gene

In order to research the tissue expression characteristics of GhGA20ox6, the expression level of the GhGA20ox6 in 9 tissues of TM-1 upland cotton material 15DPA fibers, ovules, flowers, sepals, terminal buds, stems, leaves, pistils and stamens is analyzed by utilizing a qRT-PCR method. The results show that: in these 9 tissues, ghGA20ox6 was predominantly expressed in the stem, while the expression level was lower in both ovule and flower tissues (FIG. 1). It is further speculated that GhGA20ox6 functions during the growth and development of the stem.

2.3GhGA20Ox6 protein subcellular localization analysis

The protein sequence of GhGA20ox6 is in WoLF PSORT (https://wolfpsort.hgc.jp/) Subcellular localization prediction was performed on online software and results showed localization at the cell membrane. To verify whether the GhGA20ox6 protein was localized on the cell membrane, leaf cells of Nicotiana benthamiana were transiently expressed with GhGA20ox6 (FIG. 2). The results showed a bright green distribution in the cell membrane, indicating a large amount of green fluorescence signal in the cell membrane, consistent with the predicted results, indicating that GhGA20The ox6 protein is a membrane-localized protein.

2.4 phenotypic Observation of GhGA20ox6 Gene after silencing and expression level analysis

The cotton plants of the positive control pCLCrVA-PDS showed albino phenotype, and the cotton plants of pCLCrVA-GhGA20ox6 showed different shortening of internodes of stems compared with the control group (FIG. 3), and the plant height data were plotted as a bar graph (FIG. 4). The results show that the height of the positive strain subjected to virus-induced gene silencing is obviously reduced, the length of the first internode and the length of the third internode are slightly shortened, and the length of the second internode, the length of the fourth internode and the length of the sixth internode are obviously shortened, which indicates that GhGA20ox6 silencing can cause dwarfing of the strain type. The expression level of the GhGA20ox6 between the six sections was measured, and the real-time fluorescence quantitative results showed that the expression levels of the GhGA20ox6 between the first section and the third section were not significantly different from those of the inoculated no-load control group, and the expression levels of the GhGA20ox6 between the second section, the fourth section, the fifth section and the sixth section were significantly different from those of the control group (FIG. 5). Silencing of the GhGA20ox6 gene is presumed to result in shortening between different nodes of the stem.

2.5 Functional verification of GhGA20ox6 gene in arabidopsis thaliana

Under long-day conditions, wild-type and transgenic Arabidopsis thaliana were grown simultaneously and growth was observed (FIG. 6). The results show that the wild type arabidopsis started bolting 22 days after seedling transplantation, the average bolting time was 25 days, the transgenic arabidopsis started bolting 18 days after seedling transplantation, the average bolting time was 20 days, and the bolting time was significantly earlier than that of the control wild type. The average plant height of the transgenic arabidopsis line was about 31cm, which was significantly higher than the control group arabidopsis, 24cm, and the number of rosette leaves of the transgenic arabidopsis was also significantly increased (table 2). The stem of transgenic arabidopsis is sampled, RNA is extracted, the expression condition of the GhGA20ox6 gene in the stem of arabidopsis is detected (figure 7), the GhGA20ox6 in the transgenic arabidopsis is highly expressed, the condition that a 35S promoter drives the exogenous gene to be transferred is met, the GhGA20ox6 is integrated on the chromosome of arabidopsis, and the GhGA20ox6 gene promotes the growth of the stem.

TABLE 2 bolting time, plant height and rosette number statistics of wild type and transgenic Arabidopsis thaliana

Material	Plant height	Number of rosette leaves	Bolting time
				Materials	Plant height(cm)	Number of rosette leaves	Bolting time(d)
WT	24.00+0.26	9.90+0.26	25.44+0.71
				Line1	31.46+0.47**	11.80+0.21**	20.22+0.70**
Line2	30.21+0.29**	11.55+0.17**	20.56+0.56**
				Line3	29.67+0.35**	11.40+0.18**	20.67+0.60**

。

3. Zxfoom

The gibberellin 20 oxidase family plays an important role in the growth and development of plants and environmental adaptation, and is an important key enzyme for synthesizing active gibberellin in plants.

The invention clones 1 gibberellin oxidase gene GhGA20ox6 from TM-1 land cotton material, and through amino acid multiple sequence comparison, finds that the GhGA20ox6 protein sequence has highly conserved motif LPWKET combined with substrate, conserved sequence NYPXCQKP related to 2-ketoglutarate combination, and the GhGA20ox6 gene has obvious difference of expression in different TM-1 tissues and higher expression in stem tissues. The GhGA20ox6 gene is considered to be involved in the growth and development of cotton stems, and the plant height character is influenced.

At present, the plant height is basically controlled by castration and topping in agriculture, so that on one hand, labor force is increased, and the working time is prolonged. By using VIGS experiments and transgenic overexpression experiments, the research on the expression level of the GhGA20ox6 gene in a plant body on a molecular level proves that the GhGA20ox6 gene plays a role in regulating and controlling the plant height trait.

4. Conclusion

The invention clones GhGA20ox6 gene from stem of upland cotton material TM-1 by T-A cloning method, and the amino acid sequence analysis shows that the GhGA20ox6 gene has 2OG-Fe (II) oxydenase and DIOX _ N structural domain. The cDNA of the GhGA20ox6 gene is 1155bp, and codes 384 amino acids. Subcellular localization prediction and experimental results showed localization to the cell membrane. The expression level of TM-1 shows that the GhGA20ox6 gene has obvious tissue specificity and the highest expression level in the stem. VIGS experiment results show that different stem nodes of cotton have dwarfing phenotypes in different degrees compared with a control group due to silencing of the GhGA20ox6 gene in the cotton, and the expression level of the GhGA20ox6 gene among different nodes is reduced in different degrees. Results of heterologous overexpression of the transgenic arabidopsis show that compared with wild type, the transgenic arabidopsis has the advantages of increased rosette leaf number, advanced bolting time and obvious increase of plant height, and the GhGA20ox6 gene plays an important role in plant height and shape.

The above-described embodiments are only preferred embodiments of the present invention, and are not intended to limit the present invention in any way, and other variations and modifications may be made without departing from the spirit of the invention as set forth in the claims.

Sequence listing

<110> Zhejiang agriculture and forestry university

<120> upland cotton plant height regulatory gene GhGA20ox6 and application thereof

<130> 2021.02

<160> 28

<170> SIPOSequenceListing 1.0

<210> 1

<211> 1155

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 1

atgtctctct ctgtcttaat ggattcaggt tctccaacca ttcttttaca tccatccata 60

gaaccaagag atgagacggg tggtgtcctt tttgacccat ccaagttgca aaaccaatca 120

agtttaccct cagagttcat atggccatgt ggggacttgg ttcacaccca agaagagctt 180

aacgaaccat tgatagactt gggcgggttc atcaaaggag acgaagaagc tactgcacat 240

gcagtgggcc ttgttaagac tgcctgttct aaacatggtt tcttccaagt tacaaaccat 300

ggtgttgatt caaaccttat ccaagctgct taccaagaga ttgatgctgt gtttaagtta 360

ccccttaaca agaagctaag tttccaaaga aagcctggtg gtttttcagg ctattctgct 420

gcccatgctg accggttttc cgctaagttg ccatggaagg aaacattttc ttttggctac 480

caaggactta actccgatcc ttctgtggtt cattatttca gctctgcctt gggggaagat 540

tttgagcaaa ccgggtggat ttaccaaaag tattgtgaaa agatgaggga gctgtcgcta 600

ctgatattcg aactgttggc aatcagcttg gggatagatc gtttacacta cagaaaattt 660

ttcgaagatg ggaattcaat aatgaggtgt aattactatc ccccatgcaa taattctggc 720

ctcacccttg gcactggccc tcactctgat cctacttcct taacaattct tcaccaagat 780

caagtgggag ggctcgaagt tttcaacaat aacaaatggt atgctgttcg acctcgacaa 840

gatgcctttg tcattaacat tggtgatact ttcatggcat tatgtaacgg aagatacaag 900

agttgcctgc atagagcagt ggtgaacaag gagagggaga ggagatcatt ggtatacttt 960

gtgtgcccaa aagaagacaa aatagtgaga cccccacaag atctaatgtg cagatcagga 1020

gggccaagag tgtatcctga tttcacatgg tctgatttgt tggaattcac tcaaaatcac 1080

tatagagctg acgttgctac actccaaagc ttcttccctt ggctcctttc ttctaaccct 1140

acttccaact tctag 1155

<210> 2

<211> 1140

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 2

atggcaattg attgcgtttc caagataccc tccatgcctc atcatcctaa agatgaaaaa 60

agagaggagc aaaaacaact ggtttttgat gcctcggtgc ttaaatacca acccgacata 120

ccaaaacaat ttatatggcc cgaccacgaa aagcctaatg ttaatgcacc agaactccaa 180

gtgccattca ttgacctagg agggttcctt tccggtgacc ctgtttctgc aatggaagca 240

tcgaggctgg tcggcgaggc atgtcggcag catggtttct tccttgtggt taatcatgga 300

gtggatgcaa cactggtggc tgatgctcac agttatatgg gtaacttctt tgaattgcca 360

ctcaatgata agcaaagggc tcagaggaaa cttggtgagc actgtggata tgctagtagc 420

ttcactggta gattctcctc caagctgcca tggaaggaaa cgctttcctt ccggtattca 480

gctgacaaaa agtcatccaa gattgttgaa gactaccttg atggcaaatt gggagatgaa 540

ttcaagcatt tcgggagggt ttaccaagat tactgcgagg caatgagcaa gctatctcta 600

gggataatgg agctattagc cattagtctt ggcgtaggaa gatcacattt cagggaattt 660

ttcgaggaaa atgaatcaat aatgaggctg aattactacc caccatgcca aaaaccagac 720

ctcactttag gaacagggcc tcattgcgat ccaacctcat taaccatcct tcaccaagac 780

cgagttggtg gtcttcaagt gtttgtagac aatgaatggc gttcaattag cccaaatgtc 840

gaagcatttg tcgttaacat tggcgacacc ttcatggcac tttcaaatgg gcgatacaag 900

agttgcttgc accgggcagt ggtgaaccgc cacatcccaa gaaaatctct ggctttcttc 960

ctatgtccca agggtgataa agtggtagcc ccaccaacag agttggtcga cgcctataat 1020

cccagagtat atccagattt tacttggcct atgctgcttg aattcacaca aaagcattat 1080

agagctgata tgaacacact tgaagtcttc tcaaactggg ttcaacagag aaacagctga 1140

<210> 3

<211> 1257

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 3

atgataccaa ataatttgat aatagcttgt aatgggagtc tattacctat atttaatgca 60

actctctcac cttcttgccc tacgtccaca tacactgtct tcctaatgga ttcaacgcat 120

ctcctttcct ctccgcttga gattcaagat caaacactag ttgaccatca tagttcttct 180

atcggctcat cttttctcca aaaccaaacc aacgtcccca aagaatttct ttggcccaaa 240

gttgatttag ttaatgctca tcaagagctg ttggagccac ttgtagatct tgaacgcttc 300

tttagaggtg atgaattggc gattcaacag gctgccaagg tcattagggc tgcttgtcta 360

acccacggtt gctttcaagt catcaatcat ggggttgatt cccacctcat caatgctgca 420

tattatcacc tcaatcgttt cttccatttg ccacttagcc acaaattaag ggctcgaagg 480

gccactactg ctggattaaa caccttgagc tattcaggtg cacattcgga tcgtttctct 540

tcgaatttgc catggaagga aactttaact ttccggttgc atgagaatcc caaggaatcc 600

agtgtcgtag atttgttcaa atccagttta ggagatgatt ttgaagaaat gggcataaca 660

tatcaaaagt actgtgaagg aatgaagagc ttagctctag cagtgatgga aatactagca 720

atcagcttgg gagttgatcg attgcactat aaaaattatt ttcaggacgg tgggtctata 780

atgcgatgta actattatcc gccatgcccc gagccaggac ttaccttcgg cactggtcct 840

cattgtgacg ccacatcttt aacaattctc caccaagacg aagttggagg cttggaaatc 900

tttgcaaaca acaaatggca gattgttcga cctcgtcagg atgccctagt aatcaacatc 960

ggtgagacct tcacggcatt aacaaatggg agatacaaga gttgcctgca tagggcagta 1020

gtaaacagcg agagggcgag aaaatcattg gtatactttg tttgcccacg agaagacaag 1080

gtggtgagac ccccagagga tcttgtacaa gttgatcaac tcccaagagc ttaccctgat 1140

ttcacatggt ccgatttcct ccatttcacc caaaactact acagagctga cgctcatact 1200

ctccatagct tcatcaaatg gctctcatct tccagcccca ttcatcacaa ccgttaa 1257

<210> 4

<211> 1140

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 4

atggcaattg actgcgtctc cagcatagcc tccatgcctc accatctcaa agacgacagt 60

aaagatgagc aaaaacaact agtttttgat gcttctgtgc ttaaatacca atcctgcata 120

ccacaacaat ttatatggcc tgacgacgaa aagccttgcg ctagtgcacc agaactccaa 180

gtaccactca ttgacattaa aggtttcctc tctggtgacc ctgttgctgc gatggaagct 240

tcaaggcttg tcggcgaggc atgtcaacag catggtttct ttcttgtagt taatcatgga 300

gtcgatgcca cactcgtggc cgatgctcat aggtacatgg ataacttctt tgaattgcca 360

ctctgcgaaa agcaatgggc tcagagaaaa gttggtgagc actgtggata tgccagcagc 420

ttcaccggca ggttctcctc caagttgcct tggaaagaaa cactttcttt tcgctactca 480

gctcagagaa acgcatccac gatggttgaa gactaccttg ttaataaaat gggagatgaa 540

ttcaggcatt tcggaagggt ataccagagc tactgcgagg ctatgagcaa gctttctcta 600

gggattatgg agcttttagc catcagtctg ggcgtaggca gagcgcattt cagggaattt 660

tttgaggaaa atgattcaat aatgaggctg aattactatc cgctttgcca aaaaccagac 720

ctcactttag gaacggggcc tcattgcgat ccaacgtctt taaccatcct tcaccaagac 780

cgagttggtg gtcttcaagt gtttgtagac aacgaatggc gttcaatcag cccaaatttc 840

gaagcatttg ttgttaacat tggcgacacc ttcatggctt tatcaaatgg aagatacaaa 900

agttgcttgc accgggcggt tgtgaacagt cacaccccaa gaaaatccct tgctttcttt 960

ttgtgcccaa agggtgataa agtggttacc ccaccaaaag agttggtgga cgcaaatagc 1020

cccagagtat atccagattt tacatggcct atgctgcttg aattcacaca aaagcactac 1080

agagctgaca tgaatacgct tgaagtattc tcaaactggg ttcaacagag aagccgctga 1140

<210> 5

<211> 1158

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 5

atggtacatt gtctgccaaa ggtttctgtc attcaagaca agccgatgct tccaccaact 60

gctgctgtcg caaagaataa gcatcgccct ttagctttcg atgcatcaat ccttggatct 120

gagatatcct ctcagttcat atggcctgac gacgacaagc catgcctgga tgccccggaa 180

cttgtaattc caaccattga cttgggagct ttcttgcttg gggactctct tgctgtatca 240

aaagcggcag aggcggtgaa tgaggcatgc aagaagcatg ggttttttct ggttgtaaac 300

catggggtgg attcagggct tatagacaaa gctcatcagt acatggaccg cttcttcagc 360

ctgcaactct ctgagaagca aaaggctaag agaaaagtag gagaaagcta tgggtatgca 420

agcagtttcg ttgggaggtt ttcctccaag ttgccatgga aagaaacact gtcttttcgg 480

tactgtcctc acactcaaaa catcgtgcaa cactatatgg tgaattggat gggtgaagat 540

tttagagatt tcgggaggct ttaccaggaa tattgtgaag ccatgaacaa ggtttcccaa 600

gagataatgg ggctgctagg gataagtcta ggacttgacc aagcctactt caaagacttt 660

ttcgacgaaa atgattcaat cctgagactg aatcactacc ctccatgcca aaagcctgag 720

ttgactctgg ggactggtcc ccacaccgat cccacttcct tgacaatcct tcatcaggat 780

caagttggag gccttcaagt gtttgcagat gaaaagtggc actccgttgc tcccatccca 840

cgagctttcg ttgtcaacat tggcgacaca ttcatggctt tgacaaacgg tatttacaag 900

agctgcttgc acagagcagt ggtgaatact gaaacggtga gaaaatcact tgtattcttt 960

ctatgtccca agctggagag accggtgaca ccagcagctg gcctagttaa tgccgcaaat 1020

tcgaggaagt acccagactt tacttgggca gcactgcttg aatttacaca gaaccattac 1080

cgtgctgaca tgaaaaccct tgttgcattc tccaaatggg ttcaagaaca agaatcgaac 1140

aacaaattaa taccttag 1158

<210> 6

<211> 1128

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 6

atggttcatt gtctgccaaa ggtttctgtc attcaagaca agccgatgct tccatcaact 60

tctgctgtcg caaaggatga gtattggccg cctttagctt tcgatgcatc gatccttgga 120

tctgaatcca acataccctc tcagttcata tggcctgacg acgagaagcc atgcctggat 180

gccccagaac ttgtaattcc aaccattgac ttgggagctt tcttgcttag gtactctctt 240

gctgtatcaa aagcggcaga ggtggtgaat gaggcatgca agaagcatgg gttttttctg 300

gttgtaaacc atggggtgga ttcagggctt atagacaaag ctcatcagta catggaccgc 360

ttcttcagcc tgcaactttc tgagaagcaa aaggctaaga gaaaagtagg agaaagctat 420

gggtatgcaa gcagtttcgt tgggaggttt tcctccaagt tgccatggaa agaaacactg 480

tcttttcggt actgtcctca cactcaaaat atcgtgcaac actatatggt gaatttgatg 540

ggtgaagatt ttagagattt cgggttccaa gagataatgg ggctgctagg gataagtcta 600

ggacttgacc aagcctactt caaagacttc ttcgaagaaa atgattcaat cctgagactg 660

aatcactatc ccccatgcca aaagcctgag ttgactctgg gaaccggtcc ccacaccgat 720

cccacttcct tgacaatcct tcatcaggat caagtgggag gcctttcagg ttttgcagat 780

gaaaagtggc actccgttgc tcccatccca ggagctttcg ttgtcaacat tggcgacaca 840

ttcatggctt tgacaaacgg tatttacaag agctgcttgc acagagcagt ggtgaatact 900

gaaacggtga gaaaatcact tgcattcttt ctatgtccca agctggagag accggtgaca 960

ccagcagctg gcttagtaaa tgccgcaaat ccgaggaaat acccagactt cacttgggca 1020

gcactgctga aatttacaca gaaccattac cgtgctgaca tgaaaaccct tgttgcattc 1080

tccaaatggg ttcaagaaca agaatcgaac aacaaattaa taccttag 1128

<210> 7

<211> 1104

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 7

atggcaattg attgcgtttc caagatatcc atgcctcatc atcctaaaga tgaaaaaaga 60

gaggagcaaa aacaactggt ttttgatgcc tcggtgctta aataccaacc cgacatacca 120

aaacaattta tatggcccga caaggaaaag cctagtgtta atgcaccaga actccaagtg 180

ccattcattg acctaggagg gttcctttcc ggtgaccctg tttctgcaat ggaagcatcg 240

aggctggtgg gcgaggcatg tcggcagcat ggtttcttcc ttgtggttaa tcatggagtg 300

gatgcaacac tggtggctga tgctcacagt tatatggcaa aggctcagag gaaacttggt 360

gagcactgtg gatatgctag tagcttcact ggtagattct cctccaagct gccatggaag 420

gaaacgcttt cctttcggta ttcagctgac aaaaactcat ccaagattgt tgaagactac 480

cttgttggca aattgggaga tgaattcaag catttcggga gggtttacca agattactgc 540

gaggcaatga gcaagctatc tctagggata atggagctat tagccattag tcttggcgta 600

ggaagatcac atttcaggga attttttgag gaaaatgaat caataatgag gctcaattac 660

tacccaccat gccaaaaacc agacctcacc ttaggaacag ggcctcattg cgatccaacg 720

tcattaacca tccttcacca agaccgagtt ggtggtcttc aagtgtttgt agacaatgaa 780

tggcgttcaa ttagcccaaa tctcgaagca tttgtcgtta acattggcga caccttcatg 840

gcactttcaa atgggcgata caagagttgc ttgcaccggg cagtggtgaa ccgccacatc 900

ccaagaaaat ctctggcttt cttcctatgt cccaagggtg ataaagtggt agccccacca 960

acagagttgg tcgacgccta taatcccaga gtatatccag attttacttg gcctatgctg 1020

cttgaattca cacaaaagca ttatagagct gatatgaaca cactcgaagt cttctcaaac 1080

tgggttcaac agagaaacag ctga 1104

<210> 8

<211> 1140

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 8

atggcaattg attgcgtttc caagataccc tccatgcctc atcatcctaa agatgaaaaa 60

agagaggagc aaaaacaact ggtttttgat gcctcggtgc ttaaatacca acccgacata 120

ccaaaacaat ttatatggcc cgaccacgaa aagcctaatg ttaatgcacc agaactccaa 180

gtgccattca ttgacctagg agggttcctt tccggtgacc ctgtttctgc aatggaagca 240

tcgaggctgg tcggcgaggc atgtcggcag catggtttct tccttgtggt taatcatgga 300

gtggatgcaa cactggtggc tgatgctcac agttatatgg gtaacttctt tgaattgcca 360

ctcaatgata agcaaagggc tcagaggaaa cttggtgagc actgtggata tgctagtagc 420

ttcactggta gattctcctc caagctgcca tggaaggaaa cgctttcctt ccggtattca 480

gctgacaaaa agtcatccaa gattgttgaa gactaccttg atggcaaatt gggagatgaa 540

ttcaagcatt tcgggagggt ttaccaagat tactgcgagg caatgagcaa gctatctcta 600

gggataatgg agctattagc cattagtctt ggcgtaggaa gatcacattt cagggaattt 660

ttcgaggaaa atgaatcaat aatgaggctg aattactacc caccatgcca aaaaccagac 720

ctcactttag gaacagggcc tcattgcgat ccaacctcat taaccatcct tcaccaagac 780

cgagttggtg gtcttcaagt gtttgtagac aatgaatggc gttcaattag cccaaatgtc 840

gaagcatttg tcgttaacat tggcgacacc ttcatggcac tttcaaatgg gcgatacaag 900

agttgcttgc accgggcagt ggtgaaccgc cacatcccaa gaaaatctct ggctttcttc 960

ctatgtccca agggtgataa agtggtagcc ccaccaacag agttggtcga cgcctataat 1020

cccagagtat atccagattt tacttggcct atgctgcttg aattcacaca aaagcattat 1080

agagctgata tgaacacact tgaagtcttc tcaaactggg ttcaacagag aaacagctga 1140

<210> 9

<211> 1152

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 9

atggattcaa cgcatctcct ttcctctccg cttgagattc aagatcagac actagtcgac 60

catcataatt cttctatcgg ctcatctttt ctccaaaacc aaaccaacgt ccccaaagaa 120

tttctttggc ccaaagttga tttagttaat gctcatcaag agctgttgga gccacttgta 180

gatcttgaac gcttctttag aggtgatgaa ttggcgattc agcagtctgc caaggtcatt 240

agggctgctt gtctaaccca cggttgcttt caagtcatca atcatggggt tgattcccac 300

ctcatcaatg ctgcatatta tcacctcaat cgtttcttcc atttgccact tagccacaaa 360

ttaagggctc gaagggccac tactgctgga ttaaacacct tgaactattc aggtgcacat 420

tcggatcgtt tctcttcgaa tttgccatgg aaggaaactt taactttccg ggtccatgag 480

aatctcaagg aatccagtgt cgtagacttg ttcaaatcca gtttaggaga tgattttgaa 540

gaaatgggca taacatatca aaagtactgt gaaggaatga agagcttagc tctagcagtg 600

atggaaatac tagcaatcag cttgggagtt gatcgattgc actataaaaa ttattttcag 660

gacggtgggt ctataatgcg atgtaactat tatccgccat gccccgagcc aggacttacc 720

ttcggcactg gtcctcattg tgacaccaca tctttaacaa ttctccacca agacgaagtt 780

gggggcctgg aaatctttgc aaacaacaaa tggcagattg ttcgacctcg tcaggatgcc 840

ctagtaatca acatcggtga gaccttcacg gcattaacaa atgggagata caagagttgc 900

ctgcataggg cagtagtaaa cagcgagagg gcgagaaaat cattggtata ctttgtttgc 960

ccacgagaag acaaggtggt gagaccccca gaggatcttg tacaaggtga tcaactccca 1020

agagcttacc ctgatttcac atggtccgat ttcctccatt tcacccaaaa ctactacaga 1080

gctgacgctc atactctcca tagcttcatc aaatggctct catcttccaa ccccattcat 1140

cacaaccgtt aa 1152

<210> 10

<211> 1164

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 10

atggtacatt gtctgccaaa ggtttttgtc attcaagaca agccgatgct tccatcaact 60

gctgctgtcg caaaggatga gcatcggcct ttagctttcg atgcatcgat ccttggatct 120

gaatccaaca taccctctca gttcatatgg cctgacgacg agaagccatg cctggatgcc 180

ccggcacttg taattccaac cattgacttg ggagctttct tgcttgggga ctctcttgct 240

gtatcaaaag cggcagaggt ggtgaatgag gcatgcaaga agcatgggtt ttttctggtt 300

gtaaaccatg gggtggattc agggcttata gacaaagccc atcagtacat ggaccgcttc 360

ttcagcctac aactctctga gaagcaaaag gctaagagaa aagtaggaga aagctatggg 420

tatgcaagca gtttcgttgg caggttttcc tccaagttgc catggaaaga gacactgtct 480

tttcggtact gtcctcacac tcaaaatatc gtgcaacact atatggtgaa ttggatgggt 540

gaagatttta gagatttcgg gaggctttac caggaatatt gtgaagccat gaacaaggtt 600

tcccaagaga taatggggct gctagggata agtctaggac ttgaccaagc ctacttcaaa 660

gactttttcg aacaaaatga ttcaatcctg agactgaatc actatccccc atgccaaaag 720

cctgagttga ctctgggaac cggtccccac accgatccca cttccttgac aatccttcat 780

caggatcaag ttggaggcct tcaggtgttt gcagatgaaa agtggcactc cgttgctccc 840

atcccaggag cttttgttgt caacgttggc gacacattca tggctttgac aaacggtttt 900

tacaagagct gcttgcacag agcagtggtg aatactgaaa cggtgagaaa atcacttgca 960

ttctttctat gtcccaagct ggagagaccg gtgacaccag cagctggctt agttactgcc 1020

gaaaatccga ggaaataccc agacttcact tgggcagcac tgctgaagtt cacacagaac 1080

cattaccgtg ctgacatgaa aacccttgtt gcattctcca aatgggttca agaacaagaa 1140

tcgaactaca aattaatacc ttag 1164

<210> 11

<211> 1155

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 11

atgtctctct ctgtcttaat ggattcaagt tctccaacca ttcttttaca tccatccata 60

gaaccaagag atgagacggg tagtgtcctt tttgacccat ccaagttgca aaaacaatca 120

agtttaccct cagagttcat atggccatgt ggggacttgg ttcacaccca agaagagctt 180

aacgagccat tgatagactt gggtgggttc atcaaaggag acgaagaagc tactgcacat 240

gcagtcgacc ttgttaagac tgcctgttct aaacatggtt tcttccaagt tacaaaccat 300

ggtgttgatt caaatcttat ccaagctgct taccaagaga ttgatgctgt gtttaagtta 360

ccccttaaca agaagctcgg tttccaaaga aagcctggtg gtttttcagg ctattctgct 420

gcccatgctg accggttttc cgctaagttg ccatggaagg aaacattttc ttttggctac 480

caaggactta actccgatcc ttctgtggtt cattatttca gctctgcttt gggggaagat 540

tttgagcaca ccgggaggat ttaccaaaag tattgtgaaa agatgaggga gctgtcgcta 600

gtgatcttcg aactgttggc aatcagcttg gggatagatc gtttacacta cagaaaattt 660

ttcgaagatg ggaattcaat aatgaggtgt aattactatc ccccatgcaa taattctggc 720

ctcacccttg gcactggccc tcactctgat cctacttcct taacaattct tcaccaagat 780

caagtgggag ggctcgaagt tttcatcaat aacaaatggt atgctgttcg acctcgacaa 840

gatgcctttg tcattaacat tggtgatact ttcatggcat tatgtaacgg aagatacaag 900

agttgcctgc atagagcagt ggtgaacaag gagagagaga ggagatcatt ggtatacttt 960

gtgtgcccaa aagaagacaa aatagtgaga cccccacaag atctaatgtg cagatcagga 1020

gggccaagag tgtatcctga tttcacatgg tctgatttgt tggacttcac tcaaaatcac 1080

tatagagctg acgttgctac actccaaagc ttctttcctt ggctcctttc ttctaaccct 1140

acttccaact tctag 1155

<210> 12

<211> 1083

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 12

atggcaattg attgcgtttc caagataccc tccatgcctc atcatcctaa agatgaaaaa 60

agagaggagc aaaaacaact ggtttttgat gcctcgccta atgttaatgc accagaactc 120

caagtgccat tcattgacct aggagggttc ctttccggtg accctgtttc tgcaatggaa 180

gcatcgaggc tggtcggcga ggcatgtcgg cagcatggtt tcttccttgt ggttaatcat 240

ggagtggatg caacactggt ggctgatgct cacagttata tgggtaactt ctttgaattg 300

ccactcaatg ataagcaaag ggctcagagg aaacttggtg agcactgtgg atatgctagt 360

agcttcactg gtagattctc ctccaagctg ccatggaagg aaacgctttc cttccggtat 420

tcagctgaca aaaagtcatc caagattgtt gaagactacc ttgatggcaa attgggagat 480

gaattcaagc atttcgggag ggtttaccaa gattactgcg aggcaatgag caagctatct 540

ctagggataa tggagctatt agccattagt cttggcgtag gaagatcaca tttcagggaa 600

tttttcgagg aaaatgaatc aataatgagg ctgaattact acccaccatg ccaaaaacca 660

gacctcactt taggaacagg gcctcattgc gatccaacct cattaaccat ccttcaccaa 720

gaccgagttg gtggtcttca agtgtttgta gacaatgaat ggcgttcaat tagcccaaat 780

gtcgaagcat ttgtcgttaa cattggcgac accttcatgg cactttcaaa tgggcgatac 840

aagagttgct tgcaccgggc agtggtgaac cgccacatcc caagaaaatc tctggctttc 900

ttcctatgtc ccaagggtga taaagtggta gccccaccaa cagagttggt cgacgcctat 960

aatcccagag tatatccaga ttttacttgg cctatgctgc ttgaattcac acaaaagcat 1020

tatagagctg atatgaacac acttgaagtc ttctcaaact gggttcaaca gagaaacagc 1080

tga 1083

<210> 13

<211> 1140

<212> DNA

<213> upland cotton (G.hirsutum L.)

<400> 13

atggccatcg actgcatctc caacatagcc tccatgactc accatccgaa agatgaaaaa 60

aaagatgagc agaaaaagct ggtttttgat gcctcggtgc tcaagttcga atcccagata 120

ccaaaagaat ttatatggcc tgatgacgaa aagccttccg ctaatgcacc agaactccaa 180

gtaccactca ttgacctagg aggcttcctt tctggtgacc ctgttgctac aatggaagct 240

tcaaggttta tcagcgaggc atgtcagcag catggtttct tccttgtagt taatcatgga 300

gtcgatgcaa aactcttggc tgatgcccac aagtacatgg ataacttctt tctattgcca 360

cttagacaaa agcaaagggc tcaaagaaaa cttggtgagc actgtggata tgccagtagc 420

ttcactggca ggttctcgac caagctccca tggaaggaaa cactttcttt tcgctattca 480

gccgagaaca actcatccaa gatggtggaa gactaccttg ttaataaaat gggaaatgaa 540

ttgaggcaac tcgggagggt ttaccaggac tactgcgagg cgatgagcaa gctttctctg 600

gggataatgg agcttttagc cattagtctg ggcgtaggca gagcacattt ccgggagttt 660

tttgataaaa atgattcaat aatgagacta aactactatc ccccttgtca aaaaccagac 720

ctcactttag gaacagggcc tcattgcgat ccaacgtctt taaccatcct tcaccaagac 780

agagttggtg gccttcaagt gtttgtagac aacgaatggc attcaattag cccaaatttc 840

gaagcatttg tcgttaacat tggcgacacc tttatggcac tgtcaaatgg gagatataaa 900

agttgcttgc accaagcagt ggtgaacagc cataagccaa gaaaatctct ggctttcttt 960

ttgtgtccag agggggataa agtggtaacc ccaccagcag agttggtgag ccagaacagt 1020

cccagagtat atccagattt tacatggcct atgctgcttg aattcacaca aaagcattac 1080

agagctgaca tgaacactct tcaagaattc tcaaactggg ttcaacagag aaacagctga 1140

<210> 14

<211> 384

<212> PRT

<213> upland cotton (G.hirsutum L.)

<400> 14

Met Ser Leu Ser Val Leu Met Asp Ser Gly Ser Pro Thr Ile Leu Leu

1 5 10 15

His Pro Ser Ile Glu Pro Arg Asp Glu Thr Gly Gly Val Leu Phe Asp

20 25 30

Pro Ser Lys Leu Gln Asn Gln Ser Ser Leu Pro Ser Glu Phe Ile Trp

35 40 45

Pro Cys Gly Asp Leu Val His Thr Gln Glu Glu Leu Asn Glu Pro Leu

50 55 60

Ile Asp Leu Gly Gly Phe Ile Lys Gly Asp Glu Glu Ala Thr Ala His

65 70 75 80

Ala Val Gly Leu Val Lys Thr Ala Cys Ser Lys His Gly Phe Phe Gln

85 90 95

Val Thr Asn His Gly Val Asp Ser Asn Leu Ile Gln Ala Ala Tyr Gln

100 105 110

Glu Ile Asp Ala Val Phe Lys Leu Pro Leu Asn Lys Lys Leu Ser Phe

115 120 125

Gln Arg Lys Pro Gly Gly Phe Ser Gly Tyr Ser Ala Ala His Ala Asp

130 135 140

Arg Phe Ser Ala Lys Leu Pro Trp Lys Glu Thr Phe Ser Phe Gly Tyr

145 150 155 160

Gln Gly Leu Asn Ser Asp Pro Ser Val Val His Tyr Phe Ser Ser Ala

165 170 175

Leu Gly Glu Asp Phe Glu Gln Thr Gly Trp Ile Tyr Gln Lys Tyr Cys

180 185 190

Glu Lys Met Arg Glu Leu Ser Leu Leu Ile Phe Glu Leu Leu Ala Ile

195 200 205

Ser Leu Gly Ile Asp Arg Leu His Tyr Arg Lys Phe Phe Glu Asp Gly

210 215 220

Asn Ser Ile Met Arg Cys Asn Tyr Tyr Pro Pro Cys Asn Asn Ser Gly

225 230 235 240

Leu Thr Leu Gly Thr Gly Pro His Ser Asp Pro Thr Ser Leu Thr Ile

245 250 255

Leu His Gln Asp Gln Val Gly Gly Leu Glu Val Phe Asn Asn Asn Lys

260 265 270

Trp Tyr Ala Val Arg Pro Arg Gln Asp Ala Phe Val Ile Asn Ile Gly

275 280 285

Asp Thr Phe Met Ala Leu Cys Asn Gly Arg Tyr Lys Ser Cys Leu His

290 295 300

Arg Ala Val Val Asn Lys Glu Arg Glu Arg Arg Ser Leu Val Tyr Phe

305 310 315 320

Val Cys Pro Lys Glu Asp Lys Ile Val Arg Pro Pro Gln Asp Leu Met

325 330 335

Cys Arg Ser Gly Gly Pro Arg Val Tyr Pro Asp Phe Thr Trp Ser Asp

340 345 350

Leu Leu Glu Phe Thr Gln Asn His Tyr Arg Ala Asp Val Ala Thr Leu

355 360 365

Gln Ser Phe Phe Pro Trp Leu Leu Ser Ser Asn Pro Thr Ser Asn Phe

370 375 380

<210> 15

<211> 21

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 15

atgtctctct ctgtcttaat g 21

<210> 16

<211> 20

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 16

ctagaagttg gaagtagggt 20

<210> 17

<211> 27

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 17

tgctctagag caatgtctct ctctgtc 27

<210> 18

<211> 23

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 18

cgagctcgct agaagttgga agt 23

<210> 19

<211> 30

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 19

cgagctcgat gtctctctct gtcttaatgg 30

<210> 20

<211> 23

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 20

ggactagtga agttggaagt agg 23

<210> 21

<211> 27

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 21

ggactagtgt gaacaaggag agagaga 27

<210> 22

<211> 28

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 22

ttggcgcgcc aagtagggtt agaagaaa 28

<210> 23

<211> 22

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 23

tgcacatgca gtcgaccttg tt 22

<210> 24

<211> 22

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 24

ggtcagcatg ggcagcagaa ta 22

<210> 25

<211> 19

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 25

atcctccgtc ttgaccttg 19

<210> 26

<211> 19

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 26

tgtccgtcag gcaactcat 19

<210> 27

<211> 24

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 27

agatccagga caaggaaggt attc 24

<210> 28

<211> 22

<212> DNA

<213> Artificial sequence (Artificial sequence)

<400> 28

cgcaggacca agtgaagagt ag 22

Claims

1. A method for dwarfing upland cotton is characterized by using upland cotton plant height regulation geneGhGA20ox6Gene silencing is carried out, so that the plant height of upland cotton is reduced, and dwarfing is realized;GhGA20ox6has the sequence shown in SEQ ID NO: 1.