KR102075477B1 - A gene that regulate the axillary shoot development of plant and uses thereof - Google Patents

Abstract

The present invention relates to: a transgenic plant having an increased expression of a PtrTMAX1 protein or a gene encoding the same, with accelerated axillary shoot development, and a manufacturing thereof; and an expression vector for promoting plant axillary shoot development, comprising the PtrTMAX1 gene, and a method for promoting plant axillary shoot development, comprising a step of introducing and expressing the expression vector. The present invention promotes the plant axillary shoot development by increasing the expression of the PtrTMAX1 protein or the gene encoding the same, thereby enabling the manufacture of plants with more developed axillary shoots compared to wild-type plants. Therefore, the present invention can be utilized for the development of functional crops having various sizes and shapes in the field of flowers and horticulture.

Description

A gene that regulate the axillary shoot development of plant and uses approximately}

The present invention provides a transgenic plant that promotes geodesic development, and a method of producing the same, wherein the expression of the PtrTMAX1 protein or the gene encoding the same is increased; And it relates to a plant geodevelopmental expression vector comprising a PtrTMAX1 gene and a method for promoting geodetic development of a plant comprising the step of introducing and expressing it.

According to the 2013 survey, the global flower industry market is about 50 trillion won per year, and has grown steadily with economic development. The Netherlands, which leads the flower industry, produces about 5 trillion won annually, accounting for a large part of the global flower industry. The Alsmere Flower Auction Site near Amsterdam, the capital of the Netherlands, is the world's largest, with 20 million flowers and 2 million pots a day, which accounts for 80% of the world's flowers. The Netherlands is also famous for developing new varieties and earning high royalties by improving flowers imported from all over the world.

The domestic flower market grew rapidly by 19 ~ 58% per year until the mid-1990s, following the Seoul Olympics in 1988. After a brief delay due to the IMF in 1997, the market grew steadily and reached about KRW1.1 trillion in 2008. The number of flower farmers, which was counted as only 2733 in 1980, was counted as 9450 as of 2012, an increase of about five times in 32 years. In 1991, the 21,000 pyeong aT flower market opened in Yangjae-dong. In addition, flower complexes are located in various places such as Seoul Express Bus Terminal flower wholesalers, Gupabal Flower Complex, and Jongmyo Shopping Street. As such, the domestic flower industry has grown significantly in appearance, but profitability is low due to damage caused by climate change, increased production costs due to rising oil prices, and royalties for varieties paid overseas.

Therefore, the development of highly competitive new varieties is urgent, and the solution to this situation is the development of new varieties of flowers, horticulture, crops and wood plants through the control of geodetic development. For example, it is possible to develop horticultural plants with more flowers by facilitating the geodetic development of flowers, as well as to develop horticulture / crops with significantly increased productivity by increasing fruit / seed production. In addition, by utilizing geodetic development in woody plants, it is possible to develop various types of garden trees, especially shrub trees. Furthermore, the formation of a large amount of geodetic, it is possible to produce bioenergy raw material wood with increased biomass.

Until now, the method of removing the apical growth point of the stem or using a dwarfing agent has been mainly used to promote the geodetic development of the plant through soaking. Among them, the method using a dwarfing agent is the most easily applied to various crops. Korean Patent No. 10-0126545 has developed a dwarfing agent including a fluorine-substituted absic acid derivative, but this has the disadvantage of causing environmental pollution as a chemical. Therefore, it is necessary to develop a method of plant geodetic development and morphological change without the risk of environmental pollution or physiological disorder.

Under this background, the present inventors have found TMAX1, a transcriptional regulator that regulates geodetic development in poplar trees, and by confirming that plants can promote geodetic development of plants by overexpressing the gene and its homologous genes. The invention was completed.

One object of the present invention is to provide a transgenic plant with enhanced geodetic development, with increased expression of the PtrTMAX1 protein or gene encoding the same.

It is another object of the present invention to provide a method for producing a transgenic plant that promotes geodesic development, comprising increasing the expression of a PtrTMAX1 protein or a gene encoding the same.

Another object of the present invention is to provide an expression vector for promoting geodetic development, comprising the PtrTMAX1 gene.

Still another object of the present invention is to provide a method for promoting geodetic development, comprising the step of introducing the expression vector for promoting geodetic development into a plant.

Each description and embodiment disclosed in the present invention may be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in the present invention fall within the scope of the present invention. In addition, the scope of the present invention is not to be limited by the specific description described below.

As one aspect for achieving the object of the present invention, the present invention provides a transgenic plant that promotes geodesic development with increased expression of the PtrTMAX1 protein or a gene encoding the same.

The term "PtrTMAX1" of the present invention refers to a protein which is one of a family of three-Amino-acid-Loop-Extension (TALE) transcriptional regulators of poplar or a gene encoding the same. In the present invention, the expression of the gene is increased. In the case of the transgenic plants, the gene was found to promote geodetic development compared to the wild type plants, and the gene was named PtrTMAX1 (TOO MANY AXILLARY SHOOTS1). In addition, the gene encoding the PtrTMAX1 protein is Potri.005G232000 gene, and in the present invention, PtrTMAX1 is mixed with Potri.005G232000 .

In the present invention, amino acid sequence or nucleotide sequence information of a specific protein of PtrTMAX1 is known from NCBI. Specifically, PtrTMAX1 protein may be composed of the amino acid sequence of SEQ ID NO: 1, but is not limited thereto. In addition, the PtrTMAX1 protein of the present invention may include a polypeptide having at least 50%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% homology with SEQ ID NO: 1. It doesn't work. In addition, as long as it is an amino acid sequence having such homology and exhibiting efficacy corresponding to the protein, it is obvious that an auxiliary protein having an amino acid sequence in which some sequences are deleted, modified, substituted or added can also be used as the PtrTMAX1 protein of the present invention.

In addition, it is obvious that a polynucleotide that can be translated into a protein having an amino acid sequence consisting of SEQ ID NO: 1 or a homology thereto by codon degeneracy may also be included. Or a probe that can be prepared from a known gene sequence, such as a protein comprising an amino acid sequence consisting of SEQ ID NO: 1 by hydration under stringent conditions with complementary sequences for all or part of the nucleotide sequence. Any sequence encoding a protein having similar activity can be included without limitation.

The term “homology” refers to the percent identity between two polynucleotide or polypeptide moieties. It means the degree of coincidence with a given amino acid sequence or nucleotide sequence and can be expressed as a percentage. In this specification, homologous sequences thereof having the same or similar activity as a given amino acid sequence or base sequence are designated as "% homology". Homology between sequences from one moiety to another can be determined by known art. For example, using standard software that calculates parameters such as score, identity and similarity, in particular BLAST 2.0, or by hybridization experiments used under defined stringent conditions appropriate hybridization conditions is to check, by comparing the definition is within the skill of a method well known to those skilled in the art (e.g., J. Sambrook et al., Molecular Cloning, a Laboratory Manual, 2 ^nd Edition, Cold Spring Harbor Laboratory press Cold Spring Harbor, New York, 1989; FM Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, Inc., New York.

In addition, the PtrTMAX1 gene may comprise a polynucleotide having at least 50%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% homology with the nucleotide sequence of SEQ ID NO: 2 or SEQ ID NO: 2 But it is not limited thereto.

In addition, the transgenic plants promoted geodetic development of the present invention may be an increased expression of the homologous protein of the PtrTMAX1 protein or a gene encoding the same.

The term "homologous protein" of the present invention refers to a number of proteins derived from the same protein, it can be seen whether the homologous protein through the similarity of the amino acid sequence and the three-dimensional structure. For the purposes of the present invention, proteins having at least 55% identity, or at least 75% similarity may fall into the category of the homologous protein, but are not limited thereto.

The homologous protein may be derived from Arabidopsis thaliana , but is not limited thereto. In addition, the homologous protein may be AT1G75430, but is not limited thereto. The AtTMAX1 may be represented by the amino acid sequence of SEQ ID NO: 3, or may be encoded from a gene having a polynucleotide sequence of SEQ ID NO: 4, but is limited to a specific sequence if it has an activity that can induce plant geodetic development In the present invention, the same phenotype that promotes geodetic development, even in the transgenic plants overexpressing the AtTMAX1 gene (AT1G75430) derived from Arabidopsis as the homologous protein of PtrTMAX1, specifically, is the homologous protein of the present invention. It confirmed (test example 5). Through this, it was confirmed that PtrTMAX1 and its homologous protein can induce geodetic development of plants. AtTMAX1 of the present invention was confirmed to have 58% identity, 76% similarity (similarity), compared to the protein amino acid sequence of PtrTMAX1, homologous protein having a homology range corresponding to the scope of the present invention Can be.

As used herein, the term "increased expression" means that the PtrTMAX1 protein or the gene encoding the same is introduced and expressed, or the expression is increased compared to the intrinsic or pre-modification expression of the plant. "Introduction" of the protein means that the activity of a specific protein that the plant did not originally have appears naturally or artificially. For example, the increase in expression may include introducing a foreign PtrTMAX1 protein or a gene encoding the same to increase expression; Or increase the expression of the endogenous PtrTMAX1 protein or gene encoding the same. Specifically, the expression increasing method herein,

1) increasing the copy number of the polynucleotide encoding the protein,

2) modification of expression control sequences to increase expression of the polynucleotide,

3) modification of the polynucleotide sequence on the chromosome to increase expression of the protein, or

4) may be performed by a method of modifying the expression to increase expression by a combination thereof, but is not limited thereto.

The increase in the number of copies of the polynucleotide 1) is not particularly limited, but may be performed in a form operably linked to a vector or by inserting into a chromosome of a plant of interest. In addition, as an aspect of increasing the copy number, it may be carried out by introducing a polynucleotide capable of exhibiting the activity of the protein into the plant of interest. The introduction may be performed by a person skilled in the art appropriately selected known transformation methods, the expression of the protein can be increased by the expression of the introduced polynucleotide in the plant of interest.

Next, 2) expression control sequences can be modified to increase the expression of the polynucleotide. The expression control sequence may include, but is not limited to, a promoter, an operator sequence, a sequence encoding a ribosomal binding site, a sequence that controls the termination of transcription and translation, and the like.

Specifically, a strong heterologous promoter may be linked to the top of the polynucleotide expression unit instead of the original promoter. In the present invention, the 35S promoter may be used as the promoter, but is not limited thereto.

In addition, 3) modification of the polynucleotide sequence on the chromosome may be carried out by inducing a variation on the expression control sequence, or by replacing with a polynucleotide sequence that is improved to have stronger activity.

Finally, 4) the method of modifying the expression to increase by the combination of 1) to 3), the increase in the number of copies of the polynucleotide encoding the protein, the modification of the expression control sequence to increase its expression, the above on the chromosome One or more of the modifications to the polynucleotide sequence can be performed together.

The polynucleotide may be described as a gene when the polynucleotide aggregate can function. Polynucleotides and genes may be used interchangeably herein, and polynucleotide sequences and nucleotide sequences may be used interchangeably.

The present invention prepared transgenic plants in which plant growth was inhibited by introducing PtrTMAX1 gene to increase expression of PtrTMAX1 protein or gene encoding the same. In addition, the PtrTMAX1 gene may be introduced into a operably linked state, but is not limited thereto.

The term "promoter" refers to a DNA sequence to which transcriptional regulators can bind, and the promoter binds to RNA polymerase via a transcriptional regulator, thereby transcription of an open translation frame (ORF) located downstream thereof. Can be derived. Specifically, the 35S promoter may be used, but is not limited thereto.

In addition, the term “operably linked” of the present invention refers to a state in which a nucleic acid expression control sequence and a nucleic acid sequence encoding a desired protein or RNA are functionally linked to perform a general function. It means. For example, a promoter and a nucleic acid sequence encoding a protein or RNA may be operably linked to affect expression of the coding sequence. Operative linkage with expression vectors can be prepared using genetic recombination techniques well known in the art, and site-specific DNA cleavage and ligation can employ enzymes commonly known in the art.

In the present invention, the term "promoting geodetic development" means that the geodetic development of the plant is promoted compared to the wild type (standard type). Therefore, the plant that has been promoted geodetic development may exhibit a phenotype with an increase in the stalk of the plant, the growth of length is suppressed, specifically, the geodetic development may be promoted in the present invention, but is not limited thereto.

The term "transformation" of the present invention means that DNA is introduced into a host so that the DNA can be reproduced as a factor of the chromosome or by chromosome integration. Specifically, in the present invention, it may mean introducing the PtrTMAX1 gene described above into the host.

As used herein, the term "transgenic plant" means a plant produced by the transformation using a plant as a host.

Transformation into a host cell in the present invention includes any method for introducing a nucleic acid into an organism, cell, tissue or organ, and can be carried out by selecting a suitable standard technique according to plants as is known in the art. These methods include electroshock nucleic acid molecule electroporation, plasma fusion, calcium phosphate precipitation, calcium chloride precipitation, agitation with silicon carbide fibers, agrobacterial mediated transformation, PEG, dextran sulfate lipomethamine and drying / inhibiting Mediated transformation methods and the like, but are not limited thereto, Agrobacteria mediated transformation methods are preferred.

For the purposes of the present invention, the transgenic plants can be promoted geodetic development, both herbaceous and woody plants can include without limitation, so long as they can have a phenotype promoted geodetic development by overexpression of the PtrTMAX1 gene.

For example, the herbaceous plants include Arabidopsis thaliana , Oryza sativa (rice), Zea mays (corn), Miscanthus species or Pennisetum purpureum And woody plants, such as Eucalyptus species (e.g. E. alba, E. albens, E. amygdalina, E. aromaphloia, E. baileyana, E. balladoniensis, E. bicostata, E). botryoides, E. brachyandra, E. brassiana, E. brevistylis, E. brockwayi E. camaldulensis, E. ceracea, E. cloeziana, E. coccifera, E. cordata, E. cornuta, E. corticosa, E. crebra, E. croajingoleisis, E. curtisii, E. dalrympleana, E. deglupta, E. delegatensis, E. delicata, E. diversicolor, E. diversifolia, E. dives, E. dolichocarpa, E. dundasii, E. dunnii, E. elata, E. erythrocoiys, E. erythrophloia, E. eudesmoides, E. falcata, E. gamophylla, E. glaucina, E. globulus, E. globulus subsp.bicostata, E. globulus subsp.globulus, E. gongylocarpa, E. grandis , E. grandisХurophylla, E. guilfoylei, E. gunnii, E. hallii, E. houseana, E. jacksonii, E. lansdowneana, E. latisinensis, E. leucophloia, E. leucoxylon, E. lockyeri, E. lucasii, E maidenii, E. marginata, E. megacarpa, E. melliodora, E. michaeliana, E. microcorys, E. microtheca, E. muelleriana, E. nitens, E. nitida, E. obliqua, E. obtusiflora, E. occidentalis , E. optima, E. ovata, E. pachyphylla, E. pauciflora, E. pellita, E. perriniana, E. petiolaris, E. pilularis, E. piperita, E. platyphylla, E. polyanthemos, E. populnea, E preissiana, E. pseudoglobulus, E. pulchella, E. radiata, E. radiata subsp. radiata, E. regnans, E. risdoni, E. robertsonii E. rodwayi, E. rubida, E. rubiginosa, E. saligna, E. salmonophloia, E. scoparia, E. sieberi, E. spathulata, E. staeri E. stoatei, E. tenuipes, E. tenuiramis, E. tereticornis, E. tetragona, E. tetrodonta, E. tindaliae, E. torquata, E. umbra, E. urophylla, E. vernicosa, E. viminalis, E. wandoo, E. wetarensis, E. willisii, E. willisii subsp. falciformis, E. willisii subsp. willisii, E. woodwardii ), Populus species (e.g., P. alba, P. albaХP. grandidentata, P. albaХP. tremula, P. albaХ P. tremulavar. glandulosa, P. albaХ P. tremuloides, P. balsamifera, P. balsamifera subsp.trichocarpa, P. balsamifera subsp.trichocarpaХP.deltoides, P. ciliata, P. deltoides, P. euphratica, P. euramericana, P. kitakamiensis, P. lasiocarpa, P. laurifolia, P. maximowicz maximowicziiХP.balsamferasubsp.trichocarpa, P. nigra, P. sieboldiiХP.grandideiztata, P. suaveolens, P. szechuanica, P. tomentosa, P. tremula, P. tremulaХ P. tremuloides, P. tremuloides, P. canadensis , P. yunnanensis ), conifers (eg, loblolly pine ( Pinus taeda ), slash pine ( Pinus elliotii ), ponderosa pine ( Pinus ponderosa ), lodgepole pine ( Pinus contorta ), Monterey pine ( Pinus radiata ); Douglas-fir ( Pseudotsuga menziesii ); Western hemlock ( Tsuga canadensis ); Sitka spruce ( Picea glauca ); redwood ( Sequoia sempervirens ); silver fir ( Abiesamabilis balsam fir ( Abies balsamea )) and cedar (for example, Western redcedar ( Thuja plicata ), Alaska yellow-cedar ( Chamecyparis nootkatensis )) and the like, but are not particularly limited thereto.

As another example, the herbaceous plant may be a Arabidopsis, the woody plant may be a poplar, but is not particularly limited thereto.

The term " Arabidopsis thaliana " of the present invention is a flowering plant, belongs to the Brassicaceae (Brassicaceae) and has no agricultural significance, but is a very important plant for genetic and molecular biological research widely used as a model plant in modern botany. . In the present invention, the Arabidopsis can be used as a representative plant of the herbaceous plant, specifically may be Arabidopsis thaliana, ecotype Columbia (Col-0), but is not particularly limited thereto.

The term " Populus trichocarpa " of the present invention is a generic term for plants belonging to the dicotyledonous Willow tree Willowaceae. In the present invention, the poplar may be used as a representative plant of the woody plant, Poplus alba x P. tremula var. glandulosa, clone BH and the like, but is not particularly limited thereto.

As another aspect for achieving the object of the present invention, the present invention provides a method for producing a transgenic plant with improved geodetic development, comprising the step of increasing the expression of the PtrTMAX1 protein or gene encoding the same in the plant. . Promoters used in the production method, PtrTMAX1 gene, geodetic development, transgenic plants and the like are as described above, it is possible to produce a transgenic plant of the present invention by the above method.

In addition, the method may further include the step of culturing the transformed plant prepared by the production method in soil or medium. Cultivation of the transformed plant in the present invention can be carried out according to well-known methods, conditions such as the cultivation temperature, the cultivation time and the pH of the medium can be appropriately adjusted.

The medium used should suitably meet the requirements of the specific transgenic plant. The medium may be a solid medium, a liquid medium, or a double layer medium. The components of the medium may include water, colloidal materials, agar, agarose, and gellite. (gelite) and the like can be used.

As inorganic nutrients, 15 elements, namely C, H, O, N, P, K, S, Ca, Mg, Fe, Mn, Cu, Zn, B, and Mo, are added regardless of form, and organic Carbohydrates, plant growth regulators, vitamins, etc. may be added as nutrients, and myo-inositol, glycine, L-glutamine, etc. may be added as amino acids.

As the carbon source, glucose, fructose, mannose, ribose, xylose, sucrose, melibiose, cellobiose, lactose, amylose, carbohydrate, raffinose, sorbitol, mannitol, glycerol and the like can be added.

As another aspect for achieving the object of the present invention, the present invention provides an expression vector for promoting geodetic development, comprising the PtrTMAX1 gene. The PtrTMAX1, plant geodetic development promotion and the like are as described above.

As used herein, the term “expression vector” refers to a gene construct that is a recombinant vector capable of expressing a peptide of interest in a host cell of interest, and includes a gene construct comprising essential regulatory elements operably linked to express the gene insert. The expression vector includes expression control elements such as initiation codon, termination codon, promoter, operator, etc. The initiation codon and termination codon are generally considered to be part of the nucleotide sequence encoding the polypeptide and the gene construct has been administered. It must be functional in the individual and must be in frame with the coding sequence. The promoter of the vector may be constitutive or inducible. In the present invention, a transgenic plant was prepared in which the geodesic development was promoted using an expression vector prepared by introducing a PtrTMAX1 gene of SEQ ID NO: 2 operably linked to a 35S promoter into a pK2GW7 vector. In addition, in the present invention, the PtrTMAX1 gene may be included in a vector in a state operably linked to a promoter, but is not limited thereto.

As another aspect for achieving the object of the present invention, the present invention provides a method for promoting geodetic development, comprising the step of introducing into the plant expression vector for promoting geodetic development.

Specifically, the method for promoting geodetic development of the plant of the present invention includes the step of introducing the expression vector into the plant where the geodetic development can be promoted. At this time. The promoter used in the method, the PtrTMAX1 gene, the promotion of geodetic development, and the like are the same as described above, and the method can promote geodetic development in any plant that can promote geodetic development.

The present invention promotes the geodetic development of plants by increasing the expression of the PtrTMAX1 protein or the gene encoding the same, thereby enabling the production of geodetic plants rather than wild type. Therefore, the present invention can be utilized in the development of functional crops having various sizes and shapes in the field of flowers and horticulture.

FIG. 1A shows a schematic of the pK2GW7 vector cloned with PtrTMAX1, B and C are the 1-7, 4-18, 16-1 strains and controls of the 3rd generation homozygous 35S :: PtrTMAX1 Arabidopsis transformants Expression level and phenotype of the PtrTMAX1 gene in (WT) is compared.
FIG. 2A is a graph showing the number of geodesic and rosette leaves, B is the area where the geodes are developed (the red arrow indicates the geodetic site), and C is the number of geodesic, secondary and tertiary branches. The graph shown.
3 is a diagram showing the phenotype of the main stem, B is a cross-sectional view of the main stem and root.
4 is a diagram showing the expression level of WUS by quantitative RT-PCR and RNA-sequence analysis, C is a diagram showing the results of transient activation assay analysis, D is a diagram showing the GUS staining results.
FIG. 5A is a diagram illustrating analysis of protein interaction through a yeast protein hybrid assay, and B is a diagram illustrating a result of transient activation assay analysis.
FIG. 6A is a diagram showing the site of PtrTMAX1 expression in various tissues of poplar, B is a diagram analyzing the expression of PtrTMAX1 gene by young growth period in ProTMAX1 :: GUS transgenic Arabidopsis, C is a mature ProPtrTMAX1 :: GUS transformed a view switch analyze expression of genes in Arabidopsis PtrTMAX1, D is a diagram of analyzing the expression of genes in PtrTMAX1 ProPtrTMAX1 :: GUS transgenic poplar.
Figure 7 A and B is the third generation homozygous 35S :: AtTMAX1 Arabidopsis transformants 3-10, 4-11, 9-4, 11-2 lineage and control (WT) expression of AtTMAX1 gene and plant It is a figure showing the phenotype of, C, D and E is a diagram showing the number of geodes, branches and WUS expression of the plants.
8 is a diagram showing the expression level and plant phenotype of PtrTMAX1 gene in 35S :: PtrTMAX1 poplar tree transformants 11, 27 strains and control (BH), B is an enlarged geodetic site of the plants C is a figure showing the size and geodetic number of the plants.

Hereinafter, the present invention will be described in more detail with reference to Examples and Experimental Examples. However, these Examples and Experimental Examples are for illustrative purposes only and the scope of the present invention is not limited to these Examples and Experimental Examples.

Example 1: Plant Materials and Growth Conditions

In the present invention, Arabidopsis thaliana , ecotype Columbia (Col-0) was used as a wild type or transformant. The Arabidopsis cultivars were grown in soil at 5 to 7 per pot and grown at 23 ± 2 ° C. and 14 hours of light (illuminance, 150 μmol * m ⁻² * sec ⁻¹ ) or 1% (w / v) water. Cultivation was done in half-strength MS medium (Murashige and Skoog, Sigma) containing 0.8% (w / v) agar containing appropriate antibiotics for cross and screening. In the embodiment of the present invention, hybrid poplar ( Poplus alba x P. tremula var. Glandulosa , clone BH) was used as wild type poplar. The plants were acclimated to soil and grown in growth chambers under controlled conditions.

Example 2: Genomic DNA Extraction and DNA Sequencing

Genomic DNA was extracted from the leaves of the three week old Arabidopsis using Edward buffer (200 mM Tris pH 7.5, 250 mM NaCl, 25 mM EDTA, 0.5% SDS). 1 μl of the extracted genomic DNA was used as a template for PCR (10 μl reaction), and PCR was performed with pK2GW7 vector specific primers. PCR products prepared from the PCR reaction were electrophoresed on 1% agarose gel, extracted using a gel extraction kit (GeneAll ExpinTM Gel SV, Seoul, Korea) and sequenced (Macrogen http://dna.macrogen.com). / kor /). Poplar genomic DNA was extracted directly from the leaves using DNA extraction buffer after 1 week of poplar acclimation.

Example 3: Vector Construction and Preparation of Transgenic Plants

Full- length genomic DNA encoding PtrTMAX1 (potri.005G232000.1) was amplified by PCR in Poplar and inserted into the 35S promoter downstream of the pK2GW7 vector using the Gateway cloning system to construct the 35S :: PtrTMAX1 construct. It was. The construct in the vector was introduced into Agrobacterium tumefaciens strain C58, which transformed Arabidopsis by using the floral-dip method, and transformed poplar by the leaf disk transformation-regeneration method. Transformed callus derived from poplar leaves was 1.0 mg / L 2,4-dichlorophenoxyacetic acid (2, 4-D), 0.1 mg / L benzyl aminopurine, 0.01 mg / L 1-naphthyl acetic acid (naphthylacetic acid) : NAA), 500 mg / L Cytoxim, and 50 mg / L kanamycin containing MS medium. Shoots were generated from callus cultured in wood medium containing 1.0 mg / L zeatine, 0.1 mg / L benzyl adenine, and 0.01 mg / L NAA. All the incubations were performed at 25 ± 2 ° C. and 16 hours of light (roughness, 150 μmol * m ⁻² * sec ⁻¹ ).

Example 4 Cross Section Fabrication

The stems and roots of the 80-day-old Arabidopsis immersed in FAA (formaldehyde-acetic acid-ethanol) fixative for about 1 week before making cross sections. Then, all the geodetic shoots and rosette leaves were removed and then the main stems and roots were separated. Then, an intermediate region of approximately 5 mm between the stem and the roots was excluded. The frequency section was performed at the bottom of the stem and at the top of the root.

Example 5: RNA extraction and semi-quantitative RT-PCT, quantitative RT-PCR

Total RNA of Arabidopsis leaf was extracted according to a slightly modified method using TRIZOL reagent (Ambion, CA, USA). Total RNA extraction of poplar leaves was performed using RNAPrep Pure Tissue Purification kit (TIANGEN biotech Co., Beijing, China). 0.5 or 1 μg of total RNA was reverse transcribed using a Superscript II reverse transcriptase (Invitrogen, CA, USA) in a 20 μl reaction. RT-PCR was performed in 1 μl after 2-fold dilution of the cDNA product. Amplified DNA fragments were isolated using 1% agarose gel. Quantitative real-time PCT (qRT-PCR) was performed using the CFX96 ™ Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) and iQ ™ SYBR® Supermix (Bio-Rad). Poplar ACTIN2 gene was used as a control group, ^2- calculated the relative expression levels by the ^△△ Ct method (Pfaffl2001). All primers used in the examples were designed using Primer3 software (http://fokker.wi.mit.edu).

Example 6 Transient Activation Assay

Transient transfection of Arabidopsis thaliana leaf protoplasts Preparation of reporter and effector constructs is [Plant J. 60, 649-665. To prepare the effector construct, the full length cDNA of PtrTMAX1 was linked between the CaMV 35S promoter and the nofarin synthase terminator after removing GUS from the pTrGUS vector. The reporter construct was constructed by removing the 35S promoter from the pTrGUS vector and placing the promoter fragment (AtWUSpro) in front of the GUS reporter gene. Plasmid DNA was prepared using the Plasmid Plus Maxi kit (QIAGEN, Valencia, Calif., USA) and 5 μg of reporter plasmid and 5 μg of effector plasmid were used for transfection. 1 μg of PtrNAN plasmid was added to use as an internal control for GUS activity normalization. Then 11 μl plasmid mixture (15 μg) and 100 μl of protoplasts were transferred to 2 ml microcentrifuge tubes according to the method described in Nat Protoc ., 2, 1565-1572.

β-Glucuronidase and NAN enzyme assays are described in Plant J , 32, 391-400. NAN and GUS activity was determined for MU standards on a Hoefer TK 100 fluorometer (excitation wavelength: 355 nm, emission wavelength: 460) using MUN (Sigma-Aldrich Co.) and MUG (Sigma-Aldrich Co.) as substrates, respectively. Measured. The ratio of GUS and NAN activity was calculated in relative GUS / NAN units. The experiments were performed in three replicate experiments.

Example 7: Yeast Protein Hybrid Assay (Yeast Two Hybrid)

PtrTMAX1 -OX were amplified from the cDNA of the cDNA PtrTMAX1 Arabidopsis transformants was purified using a PCR purification. The PtrTMAX1 cDNA template was prepared by specific primers to make additional two nucleic acids at the N-terminus for correct C-terminal fusion to the UAS binding domain. In addition, PtrTMAXl -ΔC, a PtrTMAXl cDNA template from which the C terminus without the homeodomain and unanimous activation site was removed, was prepared. STM was cloned from cDNA of Arabidopsis wild-type and poplar clone BH and amplified with specific primers for C-terminal fusion. All the above templates were inserted into pGBKT7 or pGADT7 by Gateway cloning method. Each vector was transformed into yeast strain AH109 using electroporation. Yeasts with bait and prey were selected in selection medium containing 10 mM 3-amino-1,2,4'-triazole (3-AT).

Example 8: GUS Staining

Tissues were stored in 90% acetone at 4 ° C. for 30 minutes before staining. The tissue was then rinsed once with 50 mM sodium phosphate and immersed in a GUS staining solution containing 5 mM potassium ferricyanide. Next, the samples were stored under vacuum for at least 30 minutes and incubated at 37 ° C. for 12 to 24 hours. Prior to observation, the samples were soaked in diacid 70% ethanol for 24 hours to remove greens.

Example 9: RNA Sequencing Assay

RNA was sown in soil at 10-day intervals and extracted by TRIZOL from wild-type and PtrTALE12-OX rosette leaf tissue including shoot apical meristem (SAM) from 30 to 50 days. Each sample was sequenced in Macrogen. As a result of sequencing, a raw fastq file was obtained, which was divided by 20 quality scores or less. The summarized data was analyzed by de novo sequencing using Trinity. Finally, the results of the differentially expressed genes were used to calculate additional fold changes.

Experimental Example 1: PtrTMAX1  Confirmation of phenotype induction with increased geodetic formation by overexpression of gene (Potri.005G232000)

To determine whether the PtrTMAX1 gene (Potri.005G232000) induces geodetic formation of plants, the following experiment was performed.

Specifically, poplar by inserting a cloned from PtrTMAX1 (Populus trichocarpa) in pK2GW7 vector having a kanamycin resistance region was produced PtrTMAX1 construct (Fig. 1A). The PtrTMAX1 overexpression state induced using the construct was named PtrTMAX1- OX or 35S :: PtrTMAX1 . Using this system, T1 generation of 16 strains were produced, and three strains (# 1, # 4, # 15) with dwarf characterization and many geodes were found. Of these, the homozygote of # 15 could not produce seeds, so # 16 was selected, indicating a weak geodetic phenotype similar to that of the wild type. Eventually, three T3 homozygote lines of PtrTMAXl- OX, # 1-7, # 4-18 and # 16-1 were obtained.

Next, as a result of confirming the RT-PCR results of the three lines, the geodetic phenotype and the PtrTMAX1 expression level matched (Fig. 1B). Specifically, the amount woke PtrTMAX1 appear most strongly expressed in the # 4-18, showed strong volume, followed by expression of PtrTMAX1 # 1-7, # 16-1 showed the weakest PtrTMAX1 expression levels, but also geodetic formed phenotype This is consistent with the trend of the RT-PCR results (FIG. 1C). In addition, the two strong phenotypes, # 1-7 and # 4-18, were bushy and dwarf phenotypes. Looking more closely at these three lineages, there were only three sideways at 60 days for the wild type, but the line with the two strong phenotypes had 8 to 12 geodes. It can be seen that the rosette leaves also showed a similar tendency with the geodetic number (FIGS. 2A and B). In addition, when the branching area was subdivided into geodetic, secondary and tertiary branches, and the number of branches was counted, the total number of branches occurring in PtrTMAX1 -OX was higher than that of wild type, but in wild type and PtrTMAX1 -OX, It was confirmed that the branches of the three regions occurred in the same pattern (FIG. 2C). Thus, based on this, it can be seen that the main factor of the unique bush-type phenotype of PtrTMAX1- OX is the number of geodesic.

In addition, another feature of PtrTMAXl- OX is a very short main stem that cannot grow more than 2-3 centimeters, which may be one cause for many geodesy (Figure 3A). Therefore, cross sections were examined to see if there were anatomical differences in the short main stems. As a result, it was confirmed that the main stem of # 1-7 and # 4-18 reduced the amount of wood fiber cells (Fig. 3B). This suggests that PtrTMAX1- OX does not have to support long lengths, resulting in a reduced number of wood fiber cells.

Experimental Example 2: PtrTMAX1  By overexpression of gene (Potri.005G232000) WUSCHEL  Confirm overexpression induction

In general, WUSCHEL ( WUS ) was known to be required for geodetic development of the rosette region, and quantitative PCR and RNA sequencing were performed to confirm the expression level of WUS in the three homozygous lines.

As a result, it was confirmed that the expression level of WUS in the three homozygous lines increased proportionally to the phenotype of 35S :: PtrTMAX1 Arabidopsis transformants (FIGS. 4A and B).

In addition, the expression level of WUS was confirmed in vitro using the Transient Activation Assay. Specifically, PtrTMAX1 was inserted into the pTrOX vector as an effector and Arabidopsis WUSCHEL ( AtWUS ) promoter was inserted into the pTrGUS vector as a reporter. Both vectors were transformed into protoplasts extracted from wild-type rosette mouth and incubated overnight at 25 ° C.

As a result of the experiment, it was confirmed that the GUS signal of the PtrTMAX1 and AtWUS promoters appeared clearly at 6 hours (FIG. 4C). From this, it can be seen that TMAX1 positively regulates the transcription process of WUS from an early stage. .

In addition, upregulation of WUS by overexpression of PtrTMAX1 was confirmed by GUS staining in vivo. Comparing F1 of proWUS :: GUS and proWUS :: GUS X PtrTMAX1-- OX showed increased blue signal in crossed F1 seedlings (FIG. 4D). Since the signal only appeared in tissues expressing WUS , it means that WUS is not a direct target of PtrTALE12.

Experimental Example 3: PtrTMAX1 Wow STM Of interaction and WUS Association with

Using a yeast protein hybrid assay, the following experiments were performed to identify the interaction partners of PtrTMAX1 belonging to the BELL-like group known to interact with the genes of the KNOX group of the Poplar TALE family. It was.

First, STM is known to function as a key factor in initiating geodetic meristem formation, first confirming that PtrTMAX1 interacts with STM . Specifically, by the Gateway cloning, the cloning PtrTMAX1 the pGBKT7 vector, and both the STM (PagSTM) derived from the STM-derived (AtSTM) and poplar, BH from Arabidopsis thaliana was cloned into the pGADT7 vector. In addition, since the C-terminus of the BELL group of the TALE family is known to have a self-activating domain, a PtrTMAX1 construct was constructed in which the C-terminus was removed as bait. Without Prey, the PtrTMAX1 full-length protein allows the yeast to survive in the selection medium, which may explain the self-activation of PtrTMAX1 . However, yeast with only PtrTMAX1_ΔC could not survive in selection medium without Prey. However, after the addition of STM / pGBKT7, the yeast was able to survive showing PtrTMAX1 interacted with STM at the protein level (Figure 5A). AtKNAT3 was used as an internal control.

Next, to confirm whether the interaction with STM affects the upregulation of WUSCHEL by PtrTMAX1 , a Transient Activation Assay with AtSTM / pTrOX as an effector was performed. As a result, the addition of AtSTM did not enhance GUS expression compared with the case of only PtrTMAX1 effects (Fig. 5B). Thus, it can be seen that the interaction with STM is not related to the overexpression of WUS by PtrTMAX1, which may be related to the nuclear function or other function of the transcription factor.

Experimental Example 4: In Poplar PtrTMAX1  Identify the location of gene expression

RNA-sequencing data of various tissues were analyzed to confirm in which tissues the PtrTMAX1 gene was expressed in poplar, and it was confirmed that the PtrTMAX1 gene was mainly expressed in leaves and human eyes (FIG. 6A).

In addition, pro PtrTMAX1 :: GUS transformed Arabidopsis transformed by the expression of the PtrTMAX1 gene for each growth period, it was confirmed that GUS staining in the SAM and leaf origin, PtrTMAX1 gene is strongly expressed in apical meristem It can be seen that (Fig. 6B).

In mature plants, the T2 generation of pro PtrTMAX1 :: GUS was found to show blue signals in rosette axilla and stem leaf axilla, leaf vein and apical meristem, where new geodes were present (FIG. 6C), where the PtrTMAX1 gene It can be seen that the expression, ProTMAX1 :: GUS transgenic poplar tree also confirmed the expression pattern similar to the Arabidopsis (Fig. 6D).

Experimental Example 5: PtrTMAX1  Arabidopsis homologous gene of the gene (Potri.005G232000) AtTMAX1 Phenotypic induction with increased geodetic formation by overexpression of (AT1G75430)

In order to check whether geodesic formation is increased even when overexpressing the Arabidopsis homologous gene AtTMAX1 (AT1G75430) of the PtrTMAX1 gene (Potri.005G232000), the following experiment was performed.

Specifically, the BLH11 gene, which is the closest homologue of the PtrTMAX1 gene, was named AtTMAX1 .

The AtTMAX1 has 58% identity and 76% similarity when compared to the protein amino acid sequence of PtrTMAX1.

The gene was cloned into the pB2GW7 vector to construct an AtTMAX1 construct, which was used to obtain 19 T1 lines. Four of the lines (# 3, # 4, # 9, and # 11) showing expressions with many features were selected. Therefore, by screening the T3 homozygotes of the selected strains, the strains (# 3-10, # 4-11, # 9-4, and # 11-2) in which overexpression of AtTMAX1 was confirmed by RT-PCR were selected. Observed.

As a result, the # 4-11 lineage showed the strongest dwarf phenotype, apical dominant was also broken, and the phenotype was excessively formed (FIGS. 7A and B).

Next, the number of geodesic and total number of branches of the four strains were counted, which was consistent with the tendency of AtTMAX1 expression level confirmed by RT-PCR of FIG. 7A (FIGS. 7C and D). As a result of confirming the WUS expression level, it was found that the amount of WUS expression was most upregulated in the # 4-11 line in which the geodetic phenotype was the strongest (FIG. 7E).

Based on the above results, BHL11 gene of Arabidopsis thaliana can be seen that the homologous gene of the gene PtrTMAX1.

Experimental Example 6: PtrTMAX1  Phenotypic induction with increased geodetic formation in poplars overexpressing the gene (Potri.005G232000)

In order to determine whether the PtrTMAX1 gene identified as inducing geodetic formation through the above experimental example induces geodetic formation in poplar, the following experiment was performed.

Specifically, by using a PtrTMAX1 construct was created by inserting the pK2GW7 vector having resistance to kanamycin area PtrTMAX1 truck poplar tree by transforming the (x P.tremula Poplus alba var. Glandulosa, Clone BH), the 35S gene-overexpressed PtrTMAX1 :: A PtrTMAX1 poplar transformant was produced. Next, RT-PCR was used to select strains (# 11 and # 27) overexpressing the PtrTMAX1 gene from the transformant strains prepared above, and the selected strains were cultivated for 60 days (FIG. 8A).

As a result of observing the cultivated poplar transformant and the control group, it was confirmed that compared to the control group (BH), the number of geodes from the leaf axle increased significantly. In addition, geodesic formation was started at node # 11 at the 15th node, and geodesic formation was started at the node 19 at # 27, but BH did not start geodetic formation at the 20th node even after 2 months of cultivation. 8B). In addition, # 11 had more geodesic numbers than the control group, but was shorter in length (FIG. 8C), indicating that the dwarf of the PtrTMAX1- OX Arabidopsis transformants was consistent with the phenotype.

The present inventors confirmed that the experimental example can produce a transgenic plant whose geodetic development is promoted by overexpression of PtrTMAX1 and its homologous protein AtTMAX1.

From the above description, those skilled in the art will understand that the present invention can be implemented in other specific forms without changing the technical spirit or essential features. In this regard, it should be understood that the embodiments described above are exemplary in all respects and not restrictive. The scope of the present invention should be construed that all changes or modifications derived from the meaning and scope of the following claims and equivalent concepts rather than the detailed description are included in the scope of the present invention.

<110> University-Industry Cooperation Group of Kyung Hee University <120> A gene that regulate the axillary shoot development of plant and          uses according <130> KPA180762-KR <160> 4 <170> KoPatentIn 3.0 <210> 1 <211> 463 <212> PRT <213> Artificial Sequence <220> <223> PtrTMAX1 <400> 1 Met Val Ser Gln Asp Ser Ser Ser Asn Ser Ala Ser Ser Met Leu His   1 5 10 15 Gln Phe Ile Phe Ser Asp Ser Ile Thr Ser Gln Asn Gln Phe Gln Asn              20 25 30 Gln Asn Phe Asp Ala Leu Val Gly Ser Asn Thr Phe Pro Gln Ser His          35 40 45 Gly Val Leu Pro Ser Ile Gln Ser Leu Glu Glu Arg Met Ser Arg Ser      50 55 60 Ile Asp Leu Val Gln Ala Pro Ser Val Ala Gln Glu Ser Glu Ile Ser  65 70 75 80 His Thr Arg His Leu Met Ser Leu Leu Gly Ala Ala Asn Glu Thr Asn                  85 90 95 Arg Gln Ala Gln Arg Leu Ser Leu Ser Leu Gly Ser His Met Leu Val             100 105 110 Pro Gln Val Gln Tyr Arg Gln Arg Ser Phe Asn Ser Asp Leu Met Ser         115 120 125 Pro Ser Tyr Leu Ile Pro Arg Glu Glu Glu Ala Arg Glu Ala Cys Asn     130 135 140 Pro Gly Gly Glu Gln Ala Asn Asn Asp Tyr Ser Leu Ile Gly Ser Gly 145 150 155 160 Phe Pro Ser Ser Pro Ala Ser Leu Ser Arg Arg Ser Thr Thr Ala Tyr                 165 170 175 Gly Thr Glu Ser Phe Ala Val Ala Ile Glu Asn Ser Arg Tyr Leu Lys             180 185 190 Pro Ala Gln Ser Leu Leu Glu Glu Thr Val His Val Ser Cys Lys Ala         195 200 205 Val Glu Ile Ser Asn Glu Lys Tyr Val Arg Arg Leu Ile Arg Cys Arg     210 215 220 Gly Ser Leu Gly Leu Ser Ser Glu Leu Lys Ala Glu Leu Trp Gly Asn 225 230 235 240 Gly Leu Val Gln Ala Glu Lys His Glu Val Gln Leu Lys Ile Ala Lys                 245 250 255 Leu Ile Ala Leu Leu Glu Glu Val Glu Gly Arg Tyr Glu Lys Tyr Tyr             260 265 270 His Gln Met Glu Glu Val Val Ser Ser Phe Glu Glu Met Ala Gly Leu         275 280 285 Gly Ala Ala Lys Ser Tyr Thr Ala Leu Ala Leu Gln Ala Met Ser Lys     290 295 300 His Phe Cys Asn Leu Arg Asp Ala Ile Val Ser Gln Ile Asn Glu Thr 305 310 315 320 Arg Arg Lys Phe Ser Gln Asp Leu Pro Arg Thr Ser Ser Gly Leu Ser                 325 330 335 Pro Leu Ser Phe Phe Asp Lys Glu Thr Lys His Asn Arg Met Ser Leu             340 345 350 Gln Gln Leu Gly Met Thr Gln Ser Gln Arg Gln Ala Trp Arg Pro Ile         355 360 365 Arg Gly Leu Pro Glu Thr Ser Val Ala Ile Leu Arg Ser Trp Leu Phe     370 375 380 Glu His Phe Leu His Pro Tyr Pro Asn Glu Ser Glu Lys Leu Met Leu 385 390 395 400 Ala Ser Gln Thr Gly Leu Thr Lys Asn Gln Val Ser Asn Trp Phe Ile                 405 410 415 Asn Ala Arg Val Arg Leu Trp Lys Pro Met Ile Glu Glu Met Tyr Lys             420 425 430 Val Glu Phe Ala Asp Ser Ser Glu Asp Ser Asn Pro Leu Pro Gly Ser         435 440 445 Ser Phe Ile Thr Arg Glu Gly Val Thr Asp His Ser Glu Asp ***     450 455 460 <210> 2 <211> 1389 <212> DNA <213> Artificial Sequence <220> <223> PtrTMAX1 <400> 2 atggtttcac aagattcatc ttcaaattca gcttctagca tgctacacca attcattttc 60 tcggactcca ttactagcca aaaccaattt caaaaccaga attttgatgc tttagttggc 120 agtaacacat ttcctcagtc tcatggtgtg ttgcctagca tacagtctct tgaggaaaga 180 atgtctaggt cgatagacct tgtacaagct ccctcagtag cgcaagaatc cgagattagc 240 cacacaagac acttaatgag tcttcttgga gcggcaaatg agaccaatcg tcaagctcaa 300 aggctatcac tgtctcttgg ttctcacatg cttgttcctc aagttcagta caggcagaga 360 tcctttaact cggatctcat gagtcctagt tacttaattc ctagagaaga agaagcaagg 420 gaagcttgta atcctggagg tgaacaagca aacaacgact attctctcat tggtagtgga 480 tttccatcat caccagcctc gttaagtaga cgttctacta ctgcttacgg aacagaatct 540 tttgctgttg ctatagagaa ttcaaggtat ttaaagcctg ctcagtccct actagaagaa 600 accgtgcatg taagttgcaa ggcagttgaa attagcaatg aaaaatatgt ccgaaggtta 660 attcgttgta gaggatctct tggactttct tctgaattaa aagcagaact gtggggaaat 720 gggcttgtgc aagctgagaa acacgaagtc cagcttaaaa ttgcaaagct tatagcattg 780 ttggaggagg ttgagggaag atacgagaaa tactaccatc aaatggaaga agtggtatca 840 tcatttgagg agatggcagg tttaggagct gccaaatctt acacagctct agcgctccaa 900 gccatgtcca agcacttctg caacttaaga gatgccatag tgtcccagat caatgagaca 960 agaagaaaat tctctcaaga tttaccaaga accagctcag gattgtcacc acttagcttt 1020 tttgataaag agaccaaaca taatcggatg tctcttcaac agcttggaat gacgcaaagc 1080 caaaggcaag catggagacc catcagagga ttgccagaga cttctgtagc catccttcgc 1140 tcttggcttt ttgaacactt cttgcatccg tatccaaatg aatctgagaa gctaatgtta 1200 gcatcgcaga caggcttgac aaagaatcaa gtctcaaact ggttcataaa tgctcgagtc 1260 cggctatgga agcctatgat agaagaaatg tacaaagtgg agttcgcaga ttcttctgaa 1320 gactcgaacc ccttacctgg cagttctttc ataacaaggg aaggtgtaac agatcattca 1380 gaggactga 1389 <210> 3 <211> 291 <212> PRT <213> Artificial Sequence <220> <223> AtTMAX1 <400> 3 Met Glu Asp Phe Arg Val Arg His Glu Cys Ser Ser Leu Arg Gly Thr   1 5 10 15 Leu Leu Asp Ser Arg Tyr Ala Lys Ala Val Gln Cys Leu Val Glu Glu              20 25 30 Val Ile Asp Ile Gly Gly Arg Glu Val Glu Leu Cys Asn Asn Ile Leu          35 40 45 Ile Asn Gln Leu Phe Pro Gly Arg Arg Arg Pro Gly Phe Ala Leu Ser      50 55 60 Ser Glu Ile Lys Ser Glu Leu Cys Ser Ser Gly Phe Met Ser Leu Pro  65 70 75 80 Glu Asn His Glu Ile His Ile Lys Ile Thr Lys Leu Leu Ser Leu Leu                  85 90 95 Gln Gln Val Glu Glu Arg Phe Glu Gln Tyr Cys Asn Gln Leu Glu Gln             100 105 110 Val Ile Ser Ser Phe Glu Glu Ile Ala Gly Glu Gly Ser Ser Lys Val         115 120 125 Tyr Thr Gly Leu Ala Leu Gln Ala Met Thr Arg His Phe Gly Ser Leu     130 135 140 Glu Glu Ala Ile Ile Ser Gln Leu Asn Ser Val Arg Arg Arg Phe Ile 145 150 155 160 Ile Ser His Gln Asp Val Pro Lys Ile Ile Ser Ser Gly Leu Ser Gln                 165 170 175 Leu Ser Leu Phe Asp Gly Asn Thr Thr Ser Ser Ser Le Le Gln Arg Leu             180 185 190 Gly Leu Val Gln Gly Pro Gln Arg His Ala Trp Lys Pro Ile Arg Gly         195 200 205 Leu Pro Glu Thr Ser Val Ala Ile Leu Arg Ala Trp Leu Phe Gln His     210 215 220 Phe Leu His Pro Tyr Pro Asn Glu Ala Glu Lys Leu Val Leu Ala Ser 225 230 235 240 Gln Thr Gly Leu Ser Lys Asn Gln Val Ser Asn Trp Phe Ile Asn Ala                 245 250 255 Arg Val Arg Leu Trp Lys Pro Met Ile Glu Glu Met Tyr Arg Glu Glu             260 265 270 Phe Gly Asp Ser Leu Asp Glu Ser Met Gln Arg Glu Ala Asn Asp Asp         275 280 285 Ser Asn ***     290 <210> 4 <211> 873 <212> DNA <213> Artificial Sequence <220> <223> AtTMAX1 <400> 4 atggaggatt ttagggtaag acatgagtgt tcatctctcc gtggtacttt gttggattca 60 agatatgcta aggcagtgca atgccttgta gaagaagtta tcgatatagg tggtagagag 120 gttgagcttt gcaacaacat tctaatcaat cagctgtttc caggaaggag aagacccggt 180 tttgctttgt cttccgaaat caaatccgaa ctttgcagtt ctggtttcat gtctttgcct 240 gagaatcatg agattcatat caaaatcaca aagcttctca gtttgttgca acaggtagaa 300 gaaagatttg agcagtactg caatcaactg gagcaagtga tttcatcatt tgaggaaata 360 gcaggagaag gatcatctaa agtgtacacc ggtttagctc tccaagccat gactcgacat 420 ttcggtagtc ttgaagaagc aataatctct cagctaaact ctgttcgtcg gagattcatc 480 atttctcatc aagatgttcc caagatcatc agctctggct tatcacagct cagcttgttc 540 gatgggaaca ccacttcatc atcacttcag cgacttggtt tggttcaagg accacagaga 600 catgcttgga aacccatcag aggcttgcct gagacttccg ttgcaattct ccgagcttgg 660 ctctttcaac atttcttgca cccttatccg aatgaagcag agaagctggt gttggcgtct 720 caaacaggac tttcgaagaa ccaagtatca aattggttta taaatgctcg agttcgactc 780 tggaaaccga tgatagaaga gatgtataga gaggaatttg gagattcctt agatgaatca 840 atgcaaagag aagccaatga tgattcaaac taa 873

Claims (13)

A transgenic plant with enhanced geodetic development, with increased expression of the Potri.005G232000 gene or PtrTMAX1 protein encoded thereby.
delete The transgenic plant of claim 1, wherein the PtrTMAX1 protein consists of SEQ ID NO: 1.
The transgenic plant of claim 1, wherein the Potri.005G232000 gene consists of SEQ ID NO: 2. 6.
The transgenic plant of claim 1, wherein the expression increase is increased by introducing a Potri.005G232000 gene.
The transgenic plant of claim 5, wherein the Potri.005G232000 gene to be introduced is operably linked to a promoter.
The transgenic plant of claim 6, wherein the promoter is a 35S promoter.
The transformed plant of claim 1, wherein the plant is Arabidopsis thaliana or Populus trichocarpa .
A method for producing a transgenic plant with improved geodetic development, comprising increasing the expression of the Potri.005G232000 gene or the PtrTMAX1 protein encoded thereby.
10. The method of claim 9, further comprising the step of culturing the transgenic plant in soil or medium.
Expression vector for promoting geodetic development, including the Potri.005G232000 gene.
The expression vector of claim 11, wherein the Potri.005G232000 gene is operably linked to a promoter.
A method for promoting geodetic development, comprising the step of introducing the expression vector of claim 11 or 12 into a plant.

Images