LU102331B1

LU102331B1 - Gene for Regulating Plant Height of Upland Cotton and Application Thereof

Info

Publication number: LU102331B1
Application number: LU102331A
Authority: LU
Inventors: Caixiang Wang; Juncheng Wang; Yonglin Yang; Junji Su
Original assignee: Univ Gansu Agricultural
Priority date: 2020-12-24
Filing date: 2020-12-24
Publication date: 2021-06-25

Abstract

The invention relates to the field of plant height genes, in particular to a gene for regulating a plant height of upland cotton and an application thereof. The gene for regulating the plant height of upland cotton has a base sequence as shown in SEQ ID NO. 1 and a CDS sequence thereof has a base sequence as shown in SEQ ID NO. 2. The present invention combines natural populations of 355 upland cotton and a SLAF-seq simplified genome sequencing technology to develop a large number of SNPs covering the whole genome of cotton, wherein a gene GhAP1-D3 for regulating the height of upland cotton is screened out from XSK 18 and other varieties by qRT-PCR through phenotypic identification of two plant height trait loci in three years, and an analysis of genes for regulating plant height according to a GWAS method.

Description

| LU102331 Gene for Regulating Plant Height of Upland Cotton and Application Thereof

TECHNICAL FIELD The present invention relates to the field of plant height gene, in particular to a gene for regulating a plant height of upland cotton and an application thereof.

BACKGROUND Cotton (Gossypium spp.) is the most important fiber crop in the world and one of the most important economic crops in China. It plays a very important role in our national economy as China is a major cotton-producing country and a major exporter of cotton textiles. Featured by high yield, strong adaptability, medium and even prominent fiber quality, upland cotton (Gossypium hirsutum L.) has been planted in more than 70 countries and regions in the world as a cultivated variety with the widest scope of coverage, and its planting area accounts for more than 95% of the total cotton area. The plant height of upland cotton is related to its yield and quality as one of the important agronomic traits. More importantly, tall plants are prone to lodging if cultivated at high density, resulting in a significant reduction in yield. Therefore, a reasonable population structure should be established on the basis of a reasonable thick planting to obtain high yield of cotton. The plant height is one of the most important traits for achieving the purpose.

The plant height QTL of upland cotton has been studied more intensively with the development of molecular technology and the construction of genetic linkage map. Shappley et al. mapped two plant height QTLs by using F2 population of intraspecific hybridization in upland cotton, which were located on the 6 and 23" linkage groups respectively. Song Xianliang et al. constructed BC1S1 population by using TM-1 and HDM 7124, and mapped three QTLs for controlling plant height, which explained 9.56%-22.05% of phenotypic variation. Yu Jiwen detected two plant height QTLs according to the F 2:3 family population constructed by CRI 36 and H 7124, which were located on chromosomes A2 and A11 respectively, and explained 9.18% and 9.48% of phenotypic variations, respectively. The populations for these studies show complex genetic background, and QTLs interact with each other, so QTLs are not accurately mapped.

The genetic background of CSSLs is similar to that of recurrent parents, and the genome only contains one or several small chromosome fragments of donor parents. QTL test on this kind of population can effectively reduce the interference of genetic background and improve the accuracy of QTL mapping. Lan Mengjiao et al. obtained 7 plant height QTLs through land-sea chromosome segment substitution lines (BC 4 F 2, BC 4 F 3) constructed by using CRI 45 and HDMH 1. Liang Yan detected a plant height QTL through three substitution line populations (BC5 F 3, BC5 F3:4 and BCS F 3:5) constructed by CRI 36 and HDMH 1, and qPH-15-19 closely linked with MUSS563 explained 7.90% of phenotypic variation.

However, the existing marker genes about plant height are worse than expected.

The present invention has been proposed as a response to this.

SUMMARY

The fest purpose of the present invention Is fo provide a gene for regidating the plant height of upland colton. The present invention comöines 355 natures populations of upland cotion a nel the SLAF-sag simplified genomes sequencing technology fo develop à large number of SNPs covering the whole genomes of cation, wherein à gene for regulating the height of upland colton is screensd out through phenotypic identification of two plant height irait lool i hres years, and an analysis of genes for regulating plant haight soconting fo à GWAR method. The hdhengih gene sequence of CES of the gene Is shown in BEQ ID NOT, and the specie sequence Is shown In SEQ NO 2 The second purpose of the present invention is io provide an appitation of the gene for regulating the plant height of upland coton, that is, the plant halght of upland colton can be regulated by promoting or inhibiting the expression of the gene.

in order to achieve the above purpose of the present invention, the technical solution used herein i as follows: The gane for regulating the plant height of upland colton has à base sequence as shows in SEQ IT NOL The present vantion also provides & specific gang fragment fer regulating the plant height of upland cotfon, of which the base sequence Is shown as SEQ I NOZ.

The present invention combines 318 natural popuiations of upland colton and the SLAF-seq Simplified genomes sequencing lechnology to develop à large number of SNP covering the whole genome of cotton, when À GATE GhaPt D3 for regulating the plant height of upland colton, with seguanoe shown as

SEQ ID NO.1 and the specific sequence shown as SEQ ID NO.2, is screened out from excellent haplotypes and candidate genes of target traits through phenotypic identification of two plant height trait loci in three years and an analysis of the genetic basis of plant height according to a GWAS method.

The present invention further provides a primer pair for amplifying the sequence shown in SEQ ID NO.1, and the upstream and downstream primers thereof are shown in SEQ ID NO.3 and SEQ ID NO 4.

The present invention further provides a primer pair for detecting the expression of the sequence shown in SEQ ID NO.1, and the upstream and downstream primers thereof are shown in SEQ ID NO.5 and SEQ ID NO.6.

The present invention further provides a primer for amplifying a specific gene fragment shown in SEQ ID NO.2, and the upstream and downstream primers thereof are shown in SEQ ID NO.7 and SEQ ID NO 8.

The primer is highly specific and amplified with a few spurious bands.

Further, the present invention also provides an application of the gene for regulating the plant height of upland cotton.

Further, the regulation is intended to increase or decrease plant height.

The gene GhAP1-D3 for regulating the plant height of upland cotton screened by the present invention is a gene for inhibiting plant height, and thus can control the plant height by regulating the activity and expression of the gene GhAP1-D3. Particularly, the plant height is increased by reducing the activity and content of GhAP1-D3 in target plants, inhibiting the expression of or knocking out the coding genes of GhAP1-D3 in target plants to obtain transgenic plants, where the height of transgenic plants obtained is higher than that of untranslated plants.

The plant height is lowered by increasing the activity or content of GhAP1- D3 in target plants or promoting the expression of coding genes of GhAP1-D3 to obtain transgenic plants, where the height of transgenic plants is lower than that of untranslated plants.

Therefore, the plant height can be regulated by overexpression or silencing of this gene.

Further, the regulation refers to increasing plant height, which is realized by inhibiting the expression of the gene for regulating the plant height of upland cotton.

Further, the inhibition is intended to regulate gene silencing of the plant height of upland cotton.

Further, the gene silencing for regulating the plant height of upland cotton is realized by the virus-induced gene silencing (VIGS) technology.

Further, the technical steps of virus-induced gene silencing are as follows: inserting the gene for regulating the plant height of upland cotton into a virus vector and transferring into agrobacterium; and infecting cotyledons of upland cotton seedlings with agrobacterium carrying virus vector, and cultivating to obtain the upland cotton capable of regulating gene silencing of the plant height.

Further, the virus vector is pCLCrVA, and the pCLCrVA and the auxiliary vector pCLCrVB are respectively transferred into agrobacterium; The gene for regulating the plant height of upland cotton is inserted into a virus vector by the following steps: realizing double-enzyme digestion of the genes for regulating the plant height of upland cotton and pCLCrVA by Spe land Asc |, respectively, and then connecting them.

Further, the regulation is intended to reduce plant height, which is realized by promoting the expression of the gene for regulating the plant height of upland cotton.

The expression of the gene for regulating the plant height of upland cotton can be promoted by adopting the existing overexpression method.

Further, the present invention also provides a vector containing an enzyme digestion product of the gene for regulating the plant height of upland cotton as shown in SEQ ID NO 2.

If the vector is a virus vector pCLCrVA, the gene for regulating the plant height of upland cotton as shown in SEQ ID NO.2 is digested with Spe | and Asc |, and the virus vector pCLCrVA is digested with the same enzyme, and then the digested products of the two are connected to obtain a vector containing the digested products of the target gene.

Compared with the prior art, the advantageous effect of the present invention is as follows: (1) The present invention combines 355 natural populations of upland cotton and the SLAF-seq simplified genome sequencing technology to develop a large number of SNPs covering the whole genome of cotton, wherein a gene for regulating the height of upland cotton is screened out through phenotypic identification of two plant height trait loci in three years, and an analysis of genes for regulating plant height according to a GWAS method. The full-length gene sequence of CDS of the gene is shown in SEQ ID NO.1, and the specific sequence is shown in SEQ ID NO 2.

(2) The present invention also provides a primer pair for amplifying a CDS sequence for regulating the plant height of upland cotton, and a primer pair for amplifying a specific sequence in the CDS sequence, which are highly specific and amplified with a few spurious bands.

(3) The present invention also provides an application of the gene for regulating the plant height of upland cotton; that is, the plant height of upland cotton can be regulated by promoting or inhibiting the expression of the gene.

BRIEF DESCRIPTION OF THE FIGURES In order to explain more clearly the embodiments in the present invention or the technical solutions in the prior art, the following will briefly introduce the figures needed in the description of the embodiments or the prior art.

Fig. 1 provides an association analysis of SNP markers and plant height in Example 1 of the present invention; Fig. 2 provides a haplotype analysis of related loci in Example 1 of the present invention; Fig. 3 provides a regional analysis of the plant height candidate genes in Example 1 of the present invention; Fig. 4 is a bar chart showing the expression of GhAP1-D3 genes of different cotton varieties in different young leaf stages in Example 1 of the present invention; Fig. 5 is an electrophoretogram of a specific sequence for silencing in CDS amplified by primers in Example 2 of the present invention: Fig. 6 is a graph of the plant height growth of different cotton plants at different days after transfection in Example 2 of the present invention; Fig. 7 shows the plant height growth of different cotton plants at different days after transfection in Example 2 of the present invention; Fig. 8 is an electrophoresis of the primers amplifying the full length of the CDS of the GhAP1-D3 gene in Example 2 of the present invention.

DESCRIPTION OF THE INVENTION Embodiments of the present invention are described in detail below with reference to examples. However, it should be understood by those skilled in the art that the following examples are for only illustration of the present invention but not a limit thereto. If any specific conditions are not indicated in the embodiment, the conventional conditions or the conditions suggested by the manufacturer shall prevail. The reagents or instruments used herein without specifying a manufacturer are conventional products that can be purchased commercially.

Example 1

1.1 Test materials The natural population constructed in this experiment included 355 representative upland cotton genetic resources, of which 185 were obtained from the variety line resources collected by the Early Maturity Group of Institute of Cotton Research (ICR) of CAAS for years, and 170 were provided by the low temperature cotton genetic resource bank in China (Anyang, Henan Province). Among the 355 genetic resources, 331 variety lines were from various ecological cotton planting regions in China, and 24 variety lines were introduced from abroad (mainly from America). Depending on the sources of different ecological regions, 355 genetic resources were divided into five groups: (1) Yellow River Basin Variety Group (YR, 162 varieties); (2) Yangtze River Basin Variety Group (YZR, 51 varieties); (3) Northwest Inland Variety Group (NW, 98 varieties); (4) Liaoning Variety Group or Northern Super Early-Maturing Variety Group (LN, 20 varieties) and (5) American Variety Group (USA, 20 varieties). All genetic materials were selected from typical plants and purified by selfing for more than 3 generations. For detailed information of 355 upland cotton varieties (lines), refer to references (a doctoral dissertation written by Su Junji, ie. Genome-Wide Association Study and Candidate Gene Screening of Early Maturity, Yield and Fiber Quality Traits of Upland Cotton).

1.2 Extraction of genomic DNA DNA was extracted by CTAB method (Paterson et al. 1993).

1.3 Investigation of plant height traits 355 upland cotton genetic resources were sown in the experimental field of Institute of Cotton Research (ICR) of CAAS in Anyang, Henan Province (36° 08' N, 114° 48' E) and the experimental field of Shihezi Experimental Station of Cotton Research Institute of Xinjiang Academy of Agricultural Sciences in Shihezi, Xinjiang (44° 31'N, 86° 01'E' e) from 2014 to 2016. The field experiments were repeated for three times through a randomized block design. In Anyang Experimental Site, Henan Province, each material was planted in a single row, with a length of 5.00 m, a row spacing of 0.80 m and a plant spacing of 0.20 m, and 18 to 23 individual plants. In Shihezi Experimental Site, Xinjiang, each material was planted in two rows, with a length of 2.00 m, a row spacing of 0.45 m and a plant spacing of 0.10 m, and 30 to 40 individual plants. Each experimental site was managed in the field by local routine methods. The cotton plant height was investigated, and the plant height of natural population was investigated in the middle ten days and the last ten days of August every year from 2014 to 2016. Ten plants were investigated for each material.

1.4 Genome-wide association study The mixed linear model (MLM) method of GAPIT software was used for genome-wide association study of 93,250 SNPs, plant height and BLUPs in each environment, taking the principal component analysis population structure (PCs) and genetic relationship {K) as covariates. The selection threshold of significance p is -logio (p) > 4.96 (p = 1/n, where n is the number of markers used; -log10 (1/93250) = 4.96). Manhattan map was drawn by R language software package "Cmplot", and the significant difference of phenotypic traits corresponding to alleles of significant association markers was verified by t test in R language program.

Fig. 1 provides the association study of SNP markers and plant height. As seen from Fig. 1, five SNPs on D03 chromosome were significantly correlated to the target traits, among which D03 31584163, D03_31972871, D03_ 32037225 and D03_32132408 were significantly correlated (-log1o (P) >

9.00, ranging from 9.29 to 13.23), and greatly contributed to the traits (10.68-

12.86%).

1.5 Haplotype analysis of related loci

Four associated SNP loci on D03 chromosome, i.e. A/G, C/T, A/G and A/G, were analyzed to identify excellent haplotypes of major QTL for controlling PH. The linkage disequilibrium analysis (LD) of the four SNP loci (D03 31584163, D03_31972871, D03 32037225 and D03_32132408) revealed a close linkage relationship between them. The natural population included five haplotypes; namely, Hap1 (ACAA), Hap2 (ATAA), Hap3 (GCGG), Hap4 (GTAA) and Hap5 (GTGG). See Fig. 2 for details.

Based on significant association loci obtained by GWAS, 355 upland cotton samples were classified depending on three different allelic variation types, and the tested materials were divided into three different groups. Further analysis showed that the average plant height (54.36cm) of 58 Hap2 varieties was significantly lower than that of Hap3 (68.20 cm), Hap5 (61.25 cm) and HAP 1 (61.90 cm). So, Hap2 was considered as an excellent haplotype for selecting lower plants.

The genomic region of SNP linked to a gene was 200 kb in the previous cotton research. Referring to this method, we intercepted 200 kb of genomic sequences at each of the four SNP loci on both sides of the LD block and identified the candidate gene region of plant height as D03: 31.38-32.33 Mb. A total of 21 genes were annotated in the 0.95Mb genome, as shown in Fig. 3.

1.6 Screening of candidate genes All SNPs within the 0.95 Mb gene region were selected by intercepting 200 kb at each of the left and right sides of the significantly associated SNP loci. LD block was mapped by the R language package to select a LD block region containing a significant association marker. The annotated genes in the segment were identified with reference to the upland cotton reference genome,

and the candidate genes for the target traits were identified by RT-PCR, transcriptome data, and homology comparison.

RNA was extracted from the terminal buds (including young leaves) and reversely transcribed to cDNA taking short plant varieties (CRI 50, plant height: 49 90cm; CRI 74, plant height: 49.50cm) and the high plant varieties (CRI 41, plant height: 65.50cm; XSK18, plant height: 66.30cm) as materials. The specific primers of the candidate genes as shown in SEQ ID NO. 5 (GhAP1-D3-F: GGAGAAAACTAATGTGGAGCAGG) and SEQ ID NO. 6 (GhAP1-D3-R: TCAAGGTGGTGGCGAATCAT) were subjected to PCR in an Applied Biosystems 7500 fluorescence quantitative PCR instrument, and participated in PCR reactions in a two-step method. The PCR procedure was as follows: pre-denaturation: 95 °C for 2 min, PCR reaction procedure: 95 °C, 5 sec; 60 °C, 34 sec, 35 cycles. The fluorescence signal was collected during annealing (60°C, 34 sec). The data were exported after reaction, and the gene expression was calculated by 2-44€T, The expression of terminal buds in 2 to 4 leaf stages was analyzed by qRT-PCR in combination with 21 annotated genes. Some of the other 20 genes were not obviously different, and some expressed differently in different stages, which could not reflect the specific relationship with plant height. However, the expression of Gh_D03G0922 gene in two short varieties was significantly higher than that in high varieties, which showed that Gh_D03G0922 gene was closely related to the plant height of upland cotton. The specific results are shown in Fig. 4. Figs. 4(a), 4(b) and 4(c) show the plant heights of different cotton varieties in 2-leaf stage, 3-leaf stage and 4- leaf stage respectively, Figs. 4(d), 4(e) and 4(f) respectively indicate the expression of Gh_D03G0922 gene in different cotton varieties in 2-leaf stage, 3-leaf stage and 4-leaf stage.

Gh_D03G0922 gene is named as GhAP1-D3 as it belongs to MADS-box family gene and homologous to Arabidopsis AP1/AGLB.

Example 2 The VIGS function is verified on GhAP1-D3 screened in Example 1.

Construction of vector of PCLCrVA-pCLCrVB VIGS system: Sequences of designed specific primers are shown in SEQ ID NO.7 and SEQ ID NO.8, specifically, GRAP1-D3V-F:AGGACAAAGCACTGCAAGAACA; GhAP1-D3V-R:TCAAGGTGGTGGCGAATCAT.

The 217bp gene silencing specific fragment of GhAP1-D3 gene “AGGACAAAGCACTGCAAGAACAGAATAACATACTTGCAAAGAAGGAAA

AGGAGAAAACTAATGTGGAGCAGGCACATTGGCAGCTGAACAACAATT GCCAAGATTCATCCTCCATGCTTCTGCCCCTTAACATCAGCTCCAATGG

AAGGGAGAAGGAAGATAATGAAACCACCAACAGTGGCGTCTTGCTGCC ATGGATGATTCGCCACCACCTTGA"” was amplified with this primer, and the amplification product is shown in Fig. 5. As can be seen from Fig. 5, the primer is highly specific and amplified without other spurious bands.

Based on Spe | (ACTAGT) and Asc | (GGCGCGCC) digestion sites, the sequence between two digestion sites on pCLCrVA vector were replaced with the C-terminal sequence of GhAP1-D3-217 to obtain pCLCrVA- GhAP1-D3-

217. pCLCrVA- GhAP1-D3-217, matching vector pCLCrVA-PDS (indicator vector), PCLCrVA (virus empty vector) and pCLCrVB (auxiliary vector) were transferred into agrobacterium LBA4404 respectively. The normal growth of monoclonal antibody was obtained by LB solid screening medium (50mg/L streptomycin + 25mg/L rifampicin + 50mg/L kanamycin), and the positive monoclonal antibody was confirmed again by bacterial liquid PCR.

Enlargement of various bacterial solutions: placing agrobacterium LBA4404-pCLCrVA-GhAP1-D3-217, LBA4404-pCLCIVA, LBA4404- pCLCrVB and LBA4404 pCLCrVA -pYL156-PDS into a 50mL of LB liquid medium containing three antibodies (50mg/L streptomycin +25ma/L rifampicin), cultivating at 28 °C, 180 rpm until the OD600 was between 1.5 and

2.0, centrifuging at 5000 rpm, 10 min to collect bacteria, and resuspending all kinds of bacterial precipitates with permeate to make OD600=1.5. (Formula of permeate: 10mM Magnesium Chloride (MgCl), 10mM 2-(4-morpholino) ethanesulfonic acid (MES) and 200pM Acetosyringone (AS)).

VIGS-based cotton seedling infection The osmotic bacteria solution was allowed to stand at room temperature in the dark for at least 3 hours, and then LBA4404-pCLCrVA-GhAP1-D3-217, LBA4404-pCLCrVA, LBA4404 pCLCrVA -pYL156-PDS and LBA4404- pCLCrVB were mixed evenly by volume. The needle of the syringe was removed to puncture the epidermis at different parts of the back of the cotyledon, the mixed bacteria solution was drawn with the syringe without a needle, and injected into the cotyledon gently, finally the back of cotyledon was fully filled with mixed bacteria solution. All cotton seedlings were placed in the dark for 12-16 hours, and then transferred to the cotton culture room (22 °C, 16 h in the light /8h in the dark, low humidity) for culture. After the leaves of pPCLCrVA-PDS cotton seedlings appeared chlorosis phenotype, RNA of all cotton plants was extracted, and the expression level of target genes was detected by fluorescence quantitative PCR, pCLCrVA-GhAP1-D3-217 cotton plants were identified, and the phenotype of all cotton plants was observed. The result is shown in Fig. 6.

As seen from Fig. 6, the height of pCLCrVA-GhAP1-D3-217(VG) positive plants in different growth stages is significantly higher than that of pCLCrvVA(CK) positive plants, which indicated that the silencing of GhAP1-D3-217 gene could increase the plant height of upland cotton, and GhAP1-D3-217 gene had an obvious regulatory effect on the plant height.

Fig. 7 shows a growth comparison of cotton plants in CK and VG groups in different growth stages. So, cotton plants in VG group were significantly higher than those in CK group.

In addition, the genome was amplified by PCR through the upstream and downstream primers shown in SEQ ID NO.3 and SEQ ID NO.4, and the resulting products were electrophoresed. The results are shown in Fig. 8.

Therefore, the amplified sequence shown in SEQ ID NO.1 provided by the present application is highly specific.

Although the present invention has been described with reference to specific embodiments, it should be understood that many other changes and modifications can be made without departing from the spirit and scope of the present invention. Therefore, it is intended that all such variations and modifications falling within the scope of the present invention are included in the appended claims.

SEQUENCE LISTING <110> Gansu Agricultural University <120> Gene for Regulating Plant Height of Upland Cotton and Application Thereof <138> PT@885 <16@> 8 <170> BiSSAP 1.3.6 <218> 1 <211> 698 <212> DNA <213> Artificial Sequence <228> <223> Gene 1 for regulating plant height <400> 1 atgggaagge gtaggettca actgaagaga atagagaata agatcaacag gcaagtgacg 60 ttttcgaaac gaaggtcggg cttgttgaag aaagcccatg aaatctetgt getttgtgat 120 gctcaagtcg ctttgatggt cttetectteg aaaggcaaac tctttgaata cgcgactgag 188 tettgcatgg aaaggatcct tgaacgatat gaaagaaact cgtatactga gatccaatgt 240 gctacagatg aaattcaaca aaatggaaac tggacctggg aacatgcaaa acttaaagct 300 agaatggaga ctttacaaag aaacctgagg cattacgaag gagaagatgt ccagaatttg 368 agtcttagag agcttcaaaa tttggagcaa caacttgatt ctgcccttaa acgcataaga 420 tccagaaaga atcaacttat gcttgaatcg atttctgagc ttcagaaaaa ggacaaagca 480 ctgcaagaac agaataacat acttgcaaag aaggaaaagg agaaaactaa tgtggagcag 540 gcacattggc agctgaacaa caattgecaa gattcatcct ccatgettet gccccttaac 600 atcagctcca atggaaggga gaaggaagat aatgaaacca ccaacagtgg cetettectg 660 ccatggatga ttcgccacca ccttgaataa 690 <210> 2 <211> 217 <212> DNA <213> Artificial Sequence <228>

<223> Gene 2 for regulating plant height <4ee> 2 aggacaaagc actgcaagaa cagaataaca tacttgcaaa gaaggaaaag gagaaaacta 60 atgtggageca ggcacattgg cagctgaaca acaattgcca agattcatcc tecatgctte 120 tgccccttaa catcagctee aatggaaggg agaaggaaga taatgaaacc accaacagtg 188 gcetcttect gccatggatg attcgccacc accttga 217 <218> 3 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Upstream primer <460> 3 atgggaaggg gtagegttca 20 <218> 4 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Downstream primer <400> à ttattcaagg tggtggcgaa 20 <218> 5 <211> 23 <212> DNA <213> Artificial Sequence <228> <223> Upstream primer 2 <408> 5 ggagaaaact aatgtggagc agg 23 <218> 6 <211> 20 <212> DNA

<213> Artificial Sequence <228> <223> Downstream primer 2 <400> 6 tcaaggtggt ggcgaatcat 28 <218> 7 <211> 22 <212> DNA <213> Artificial Sequence <228> <223> Upstram primer 2 <488> 7 aggacaaagc actgcaagaa ca 22 <218> 8 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Dowstream primer 2 <400> 8 tcaagetget ggcgaatcat 20

Claims

1. The gene for regulating the plant height of upland cotton is characterized in having a base sequence as shown in SEQ ID NO.1.

2. The gene for regulating the plant height of upland cotton is characterized in having a base sequence as shown in SEQ 1D NO.2

3. The primer pair is characterized in that the upstream and downstream primers thereof are shown in SEQ ID NO.3 and SEQ ID NO 4.

4. The primer pair is characterized in that the upstream and downstream primers thereof are shown in SEQ 1D NO.5 and SEQ ID NO.6

5. The primer pair is characterized in that the upstream and downstream primers thereof are shown in SEQ ID NO.7and SEQ ID NO.8

6. The application of the gene for regulating the plant height of upland cotton according to any one of claims 1-2 in regulating the plant height of upland cotton; further, the regulation is regulation of increasing plant height or regulation of decreasing plant height; further, the regulation is the regulation of increasing plant height, which is realized by inhibiting the expression of the gene regulating the plant height of upland cotton; further, the inhibition is to silence the gene regulating the plant height of upland cotton.

7. The application, according to claim 6, is characterized in that the gene silencing for regulating the plant height of upland cotton is realized by adopting a virus-induced gene silencing technology; the virus-induced gene silencing technology comprises the following steps: the gene for regulating the plant height of upland cotton is inserted into a virus vector and then transferred into Agrobacterium tumefaciens; Agrobacterium carrying virus vector infects cotyledons of upland cotton seedlings, and cultivates to obtain the upland cotton with gene silencing for regulating the plant height of upland cotton.

8. The application according to claim 7 is characterized in that the virus vector is pCLCrVA, and the pCLCrVA and the auxiliary vector pCLCrVB are respectively transferred into agrobacterium; according to Claim 2, realizing double-enzyme digestion of the genes for regulating the plant height of upland cotton and pCLCrVA by Spe | and Asc |, respectively, and then connecting them.

9. The application according to claim 6, is characterized in that the regulation is the regulation of reducing plant height, which is realized by promoting the expression of the gene regulating the plant height of upland cotton.

10. A vector is characterized by containing the enzyme digestion product of the gene for regulating the plant height of upland cotton according to claim

2.