KR20190044442A - Use of VP1 gene from Oryza sativa as regulator of yield and environmental stresses - Google Patents

Abstract

The present invention relates to a use of vacuolar pyrophosphatase (VP) genes derived from Oryza sativa as a regulator for yield and environmental stress. More particularly, the present invention relates to a method for producing a plant having enhanced yield and environmental stress tolerance with a recombinant vector comprising VP genes derived from Oryza sativa, and a plant or a seed thereof produced by the method, and a composition for enhancing yield and environmental stress tolerance comprising the VP genes.

Description

[0002] The yield of V P gene from rice and its use as an environmental stress moderator are described in < RTI ID = 0.0 > U. < / RTI >

More particularly, the present invention relates to a method of increasing the yield of rice using VP (vacuolar pyrophosphatase) gene derived from rice ( Oryza sativa ), a method of increasing the yield of VP A method for increasing resistance to environmental stress in rice using VP gene derived from rice, a method for producing transgenic rice having increased environmental stress tolerance using the VP gene, a method for producing transgenic rice, And a biomass thereof and a composition for promoting environmental stress tolerance of rice comprising the VP gene.

With the increase of income and the improvement of culture level, we are living a healthier and more affluent life. In response to these changes in the times, it is necessary to produce food that meets various consumer preferences. In particular, farmers and consumers should be satisfied with the production of high quality varieties with high taste, quality and nutritional value. Recently, the genome sequence of rice has been read to establish a basis for analysis of numerous rice gene functions. It is necessary to develop a technology to isolate useful genes and to produce excellent rice varieties desired by the new era.

Fortunately, most of the genes have been cloned due to the detoxification of the rice genome sequence, and a group of insertion mutants has been constructed, which is a resource for easier analysis of the functions of these genes. Therefore, it is necessary to utilize these resources effectively to analyze useful gene functions and utilize them to secure technology for producing excellent rice varieties.

The present inventors have been continuously studying for the production of superior rice varieties and have found that the recombinant DNA of the present invention can be used for recombinant production of dhar genes derived from rice (Korean Patent Laid-open No. 10-2013-0022564), mdhar (Korean Patent Laid-open No. 10-2013-0022561) We have already filed a patent application for the production of plants with the effect of increasing yield and resistance to stress by transforming the vector into plant cells. In addition, we have continued research on gene functions of rice and various plant-derived gene inserts .

Korean Patent Publication No. 10-2013-0022564 Korean Patent Publication No. 10-2013-0022561

As a result of the above study, the inventors of the present invention have completed the present invention by confirming that the yield of transgenic rice introduced with VP gene derived from rice increased and resistance to environmental stress was increased.

Accordingly, an object of the present invention is to provide a method for producing a plant having an increased yield by using VP gene derived from rice.

Another object of the present invention is to provide a composition for increasing the yield of a plant, which comprises a rice-derived VP gene.

It is another object of the present invention to provide a method for producing a plant having enhanced environmental stress tolerance by using a VP gene derived from rice.

It is another object of the present invention to provide a composition for promoting stress tolerance of a plant, which comprises a VP gene derived from rice.

In order to solve the above problems, the present invention provides a method for producing a plant having an increased yield, comprising transforming a plant cell with a recombinant vector comprising the VP gene derived from the rice, and regenerating the plant from the transformed plant cell Of the present invention.

The present invention also provides transgenic plants and their seeds with increased yields prepared by the above method.

The present invention also provides a composition for increasing the yield of a plant comprising the VP gene.

The present invention also provides a method for producing a plant having increased stress tolerance, comprising transforming a plant cell with a recombinant vector comprising the VP gene derived from the rice, and regenerating the plant from the transformed plant cell to provide.

The present invention also provides transgenic plants and seeds thereof which have been improved by environmental stress tolerance produced by the method.

The present invention also provides a composition for enhancing environmental stress of a plant comprising the VP gene.

The rice plants transformed with the VP gene of the present invention have high yield and improved environmental stress tolerance and can be very useful for increasing the yield even under conditions adverse to the growth of rice.

FIG. 1A is a vector of the VP gene expressed by the ubiquitin promoter, which is always a promoter, and FIG. 1B is a result of confirming the homologue through genotyping.
FIG. 2 is a photograph of a transformant and a wild-type plant exposed to salt stress for 7 days and recovered for 7 days and then compared with each other.
Figure 3 compares the weight of each plant after the salt stress test.
Figure 4 shows RT-PCR results after salt stress exposure.
FIG. 5 is a photograph of the roots after growth on MS medium containing 200 mM salt.
FIG. 6A shows the number of side roots after the growth on MS medium containing 200 mM of salt, and FIG. 6B shows the result of measuring root length.
FIG. 7 is a graph showing Na contents of roots before and after salt stress.
8 is a graph showing the activity of the liposome of the liposome before and after the salt stress.
FIG. 9 is a graph showing changes in root length of each experimental group in a natural growth state.
Fig. 10 shows a state after a month in the natural growth state (after the transplant).
FIG. 11 shows the results of measurement of the amount of the plant and the amount of dry matter after one month from the natural growth state.
FIG. 12 is a photograph showing the state after culturing for 50 days in a natural growth state.
Fig. 13 shows the result of measuring the change of chlorophyll content in the natural growth state.
Fig. 14 shows an increase in the number of tillers in the natural growth state.
Fig. 15 is a photograph of a plant after culturing for 90 days in a cultivated area (natural growth state, after 90 days).
Figs. 16A and 16B are the results of the analysis of the overall yield, and each element is TPW; Total plant weight, CW; Stem weight, RW; Root weight, NT; total number of tillers, NPT; Effective number of tributaries, TGW; Total seed weight, 1000GW; 1000 grains.

The present invention relates to a method for producing a plant having an increased yield, which comprises transforming a plant cell with a recombinant vector comprising a VP (vacuolar pyrophosphatase) gene derived from oryza sativa and regenerating the plant from the overexpressed plant cell Lt; / RTI >

The VP gene may be present in a multigene family, preferably, but not limited to, the nucleotide sequence of SEQ ID NO: 1. Variants of the above base sequences are also included within the scope of the present invention. Specifically, the gene includes a nucleotide sequence having a sequence homology of 70% or more, more preferably 80% or more, still more preferably 90% or more, and most preferably 95% or more, with the nucleotide sequence of SEQ ID NO: 1 can do. &Quot;% of sequence homology to polynucleotides " is ascertained by comparing the comparison region with two optimally aligned sequences, and a portion of the polynucleotide sequence in the comparison region is the reference sequence for the optimal alignment of the two sequences Or " not including ").

In the method of the present invention, the increase in the yield of the plant may be, but not limited to, an increase in the number of fission of the plant.

The term " recombinant " refers to a cell in which a cell replicates a heterologous nucleic acid, expresses the nucleic acid, or expresses a protein encoded by a peptide, heterologous peptide or heterologous nucleic acid, The modified gene, not the form, is reintroduced intracellularly by artificial means. The gene may be in the sense or antisense form.

In the present invention, the VP gene sequence may be inserted into a recombinant expression vector. The term recombinant expression vector refers to a bacterial plasmid, phage, yeast plasmid, plant cell virus, mammalian cell virus, or other vector. In principle, any plasmid and vector can be used if it can replicate and stabilize within the host. Important characteristics of the expression vector include a replication origin, a promoter, a marker gene, and a translation control element.

VP gene sequences and transcription / translation regulation signals can be constructed by methods known to those skilled in the art. Such methods include in vitro recombinant DNA technology, DNA synthesis techniques, and in vivo recombination techniques. The DNA sequence can be effectively linked to appropriate promoters in the expression vector to drive mRNA synthesis. The expression vector may also include a ribosome binding site and a transcription terminator as a translation initiation site.

A preferred example of the recombinant vector of the present invention is a Ti-plasmid vector capable of transferring a so-called T-region to a plant cell when present in a suitable host, such as Agrobacterium tumefaciens. Other types of Ti-plasmid vectors (see EP 0 116 718 B1) are currently used to transfer hybrid DNA sequences to plant cells or protoplasts in which new plants capable of properly inserting hybrid DNA into the plant's genome can be produced have. A particularly preferred form of the Ti-plasmid vector is a so-called binary vector as claimed in EP 0 120 516 B1 and U.S. Patent No. 4,940,838. Other suitable vectors that can be used to introduce the DNA according to the invention into the plant host include viral vectors such as those that can be derived from double-stranded plant viruses (e. G., CaMV) and single- For example, from non -complete plant virus vectors. The use of such vectors may be particularly advantageous when it is difficult to transform the plant host properly. The expression vector will preferably comprise one or more selectable markers.

The marker is typically a nucleic acid sequence having a property that can be selected by a chemical method, and includes all genes capable of distinguishing a transformed cell from a non-transformed cell. Examples include herbicide resistance genes such as glyphosate or phosphinothricin, antibiotics such as kanamycin, G418, Bleomycin, hygromycin, chloramphenicol, Resistant gene, aadA gene, and the like, but are not limited thereto.

In the recombinant vector of the present invention, the promoter may be CaMV 35S, actin, ubiquitin, pEMU, MAS, histone promoter, SWPA2 promoter, Clp promoter, but is not limited thereto. The term " promoter " refers to the region of DNA upstream from the structural gene and refers to a DNA molecule to which an RNA polymerase binds to initiate transcription. A " plant promoter " is a promoter capable of initiating transcription in plant cells. A " constitutive promoter " is a promoter that is active under most environmental conditions and developmental conditions or cell differentiation. Constructive promoters may be preferred in the present invention because the choice of transformants can be made by various tissues at various stages. Thus, constitutive promoters do not limit selectivity.

In the recombinant vector of the present invention, conventional terminators can be used. Examples thereof include nopaline synthase (NOS), rice α-amylase RAmy1 A terminator, phaseoline terminator, Agrobacterium tumefaciens (Agrobacterium tumefaciens ) Terminator of the Octopine gene, and the rrnB1 / B2 terminator of E. coli, but the present invention is not limited thereto. Regarding the need for terminators, it is generally known that such regions increase the certainty and efficiency of transcription in plant cells. Therefore, the use of a terminator is highly desirable in the context of the present invention.

Any host cell known in the art may be used as a host cell capable of continuously cloning and expressing the vector of the present invention while being stable in a prokaryotic cell. Also, when transforming the vector of the present invention into eukaryotic cells, As the cells, yeast (Saccharomyce cerevisiae), insect cells, human cells and plant cells and the like can be used. The most preferred host cells are plant cells.

The method of the present invention comprises transforming a plant cell with a recombinant vector according to the invention, said transformation being mediated, for example, by Agrobacterium tumefaciens. In addition, the method of the present invention comprises regenerating a transgenic plant from the transformed plant cell. Any of the methods known in the art can be used for regeneration of transgenic plants from transgenic plant cells.

Transformed plant cells must be regenerated into whole plants. Techniques for the regeneration of mature plants from callus or protoplast cultures are well known in the art for a number of different species (Handbook of Plant Cell Culture, Vol. 1-5, 1983-1989, Momillan, N. Y.).

In addition, the present invention provides an improved transgenic plant produced by the above method and seeds thereof. Preferably, the plant may be a monocot plant but is not limited thereto. Preferably, the monocot plant is rice (Oryza sativa).

In addition, the present invention provides a composition for increasing the yield of a plant comprising a VP (vacuolar pyrophosphate) gene derived from rice (Oryza sativa). Preferably, the VP gene may comprise the nucleotide sequence of SEQ ID NO: 1. The composition includes a VP gene derived from rice as an active ingredient, and the yield of the plant can be increased by transforming the gene into a plant.

The present invention also provides a method for increasing the environmental stress tolerance of a plant comprising the step of overexpressing a VP gene by transforming a plant cell with a recombinant vector comprising VP (vacuolar pyrophosphate) gene derived from rice (Oryza sativa) . Preferably, the VP gene may comprise the nucleotide sequence of SEQ ID NO: 1.

In the method of the present invention, the environmental stress may be salt, low temperature or oxidative stress, but is not limited thereto.

The present invention also relates to a method for producing a plant having an environmental stress tolerance, comprising transforming a plant cell with a recombinant vector comprising VP (vacuolar pyrophosphate) gene derived from rice (Oryza sativa) and regenerating the plant from the transformed plant cell Thereby providing an increased plant production method. Preferably, the VP gene may comprise the nucleotide sequence of SEQ ID NO: 1.

In the method for producing a plant of the present invention, the environmental stress may be salt, low temperature or oxidative stress, but is not limited thereto.

The present invention also provides transgenic plants and seeds thereof with increased environmental stress tolerance produced by the above method. Preferably, the plant may be a monocot plant but is not limited thereto.

In addition, the present invention provides a composition for enhancing environmental stress tolerance of a plant, which comprises a VP (vacuolar pyrophosphate) gene derived from rice (Oryza sativa). Preferably, the VP gene may comprise the nucleotide sequence of SEQ ID NO: 1. The composition contains a VP gene derived from rice as an active ingredient, and the gene can be transformed into a plant to enhance tolerance to environmental stress of the plant.

Hereinafter, the present invention will be described in more detail with reference to Examples. It will be apparent to those skilled in the art that these embodiments are merely illustrative of the present invention and that the scope of the present invention is not limited to these embodiments.

Example 1 Rice Transformation Using VP Genes

<1-1> Vector construction and rice transformation

Total RNA was isolated from rice ( oryza sativa ) leaves using the RNeasy Plant Mini Kit (Qiagen, Germany). The full-length cDNA encoding the VP gene was obtained from the total RNA of the rice by RT-PCR (OVP_NheI-F: 5'-CGGCTAGCGCAATGGCGGCGATACTGC-3 ': SEQ ID NO: 2, OVP_NheI-R; 5'-CGGCTAGCCCCTTGGGAGCTCGTAAATC- : SEQ ID NO: 3). Two homologous lines were finally selected and designated as OVP1 and OVP2, respectively. The cDNA was cloned between the always-expressed ubiquitin promoter and the nos terminator of the binary vector pGA1611 (Kim et al. 2003) using KpnI restriction enzyme and HindIII, and the vector was named pOsAVP1. The nucleotide sequence of the promoter and OVP was determined to confirm no frame shift or nucleotide change in the open reading frame (ORF). (Figs. 1A and 1B)

&Lt; 1-2 > Selection of Salt-resistant Transgenic Rice

To obtain homozygotes with OVP expression, 69 event lines (T0 generation) were produced by transforming rice with an OVP gene construct (pGA1611 :: OVP) regulated by the maize ubiquitin promoter and the nos terminator. To further investigate whether transgenic plants overexpressing OVP have salt tolerance to abiotic stress, primary T0 transformants were screened in the presence of hygromycin coded by the hph gene. T1 seeds were harvested from 69 independent T0 rice plants: 20 T1 seeds per T1 transgenic line were germinated in the greenhouse and harvested after cultivation in the paddy field (T2 seeds).

<1-3> Genomic DNA isolation and PCR analysis

Genomic DNA was isolated from the leaves according to the alkaline treatment method described by Klimyuk et al. (1993). (5'-TGCCTTCATACGCTATTTATTGCTTGG-3 ': SEQ ID NO: 4) and OVP-R (5'-GCGTGCTGGATCTCGGAGCACTTCTCG-3': SEQ ID NO: 5) to confirm the DNA introduction of the ubiquitin promoter and the OVP transgene in the transformants ) Primer set. The PCR reaction contained DNA and the primers in a total volume of 20 쨉 l and the conditions were as follows: initial denaturation at 93 째 C for 3 min, followed by 30 sec at 93 째 C, 30 sec at 50 째 C, 40 sec at 72 째 C 34-36 cycles, final expansion step 72 ° C 5 min. Finally, the PCR product was isolated on a 1.2% agarose gel.

 <1-4> Semi-quantitative RT-PCR and Quantitative RT-PCR Analysis

To confirm whether the OVP gene was properly expressed under the control of the ubiquitin promoter in the transformant, RNA was extracted from 4-week-old leaf tissues treated with 200 mM NaCl for 12 days and quasi-quantitative RT-PCR was performed.

To determine whether the mechanisms of OVP adapt to transgenic plants, photosynthetic ability was examined in rice fields under environmental conditions. Rice leaves were extracted from seedlings grown for 60 days in rice fields (2011). Total RNA was isolated from leaf tissue using RNeasy Plant Mini Kit (Qiagen, Germany).

Quasi-quantitative RT-PCR was performed according to the manufacturer's instructions using a One-Step RT-PCR PreMix Kit (Intron, South Korea) and a primer set to encode the sequence of each gene. Each of the quasi-quantitative RT-PCR reactions was carried out in a total volume of 20 은 by mixing the DNA and the primers: OVP-F (5'-TTTGGCCTTGCTCTGGGATA 3 ': SEQ ID NO: 6) and OVP-R (5'-TGCTGGAGGAATTGCTGAGA- 7), and the conditions are as follows. Initial denaturation at 45 ° C for 30 minutes followed by 30 seconds at 94 ° C, 30 seconds at 54 ° C, 26-28 cycles at 72 ° C for 40 seconds, and final expansion step at 72 ° C ° C for 5 minutes. Finally, the PCR product was isolated on a 1.2% agarose gel.

Tubulin gene was used as a control and amplified using the following primer sets: Tubulin-F 5'-TACCGTGCCCTTACTGTTCC-3: SEQ ID NO: 8 'and tubulin R5'-CGGTGGAATGTCACAGACAC-3: SEQ ID NO .: 9. StepOne- Plus ™ Real Quantitative real-time PCR was performed three times at each time point using -Time PCR System (Applied Biosystems, USA) and the results were analyzed using StepOne Software v2.1 (Applied Biosystems, USA). Standardization and relative quantification were performed using the 2-ΔΔCt method (Ferreira et al., 2006).

The transcription was carried out under the normal conditions of Gunyu Campus (36 ° 24'N, 128 ° 53'E, 95m asl) at Kyungpook National University, and all transgenic and wild type (WT, control) rice grown for 60 days Leaf, and this was used for subsequent experiments.

&Lt; Example 2 > Increase in yield and stress resistance of VP transformants

<2-1> Identification of salt tolerance to salt stress

To assess whether OVP overexpression affects overall plant growth and development, total biomass (including roots and shoots) of transgenic and wild-type rice plants were compared. For this purpose, wild-type and transformants were grown in a greenhouse at 28-32 ° C for 4 weeks, exposed to 200 mM NaCl for 7 days, and then restored for 7 days in NaCl-irrigated soil pots. (Wang et al., 2001; Ohta et al., 2002). The rice was dried at 80 DEG C for 72 hours to measure the dry weight (DW). The effect of salt stress on growth was measured by comparing the amount and dry weight of the experimental and control groups.

As a result, as shown in FIG. 2 , it was confirmed that the transgenic rice plants were more resilient to stress than the rice rice, which was a control, at both stresses. Also, as shown in FIG. 3 , the freshness and dry weight of the experimental group and the control group were higher than those of the control group.

In addition, semi-quantitative RT-PCR was performed to examine whether the expression of the inserted gene was increased by extracting total RNA after the salt stress treatment. Experimental results show that the expression of VP gene in rice of transgenic rice plants is increased compared with that of control rice rice regardless of the presence or absence of stress. (Figure 4)

<2-2> Comparison of root growth under salt stress

The salt treatment experimental group and the control plants performed in 2-1 above were grown in MS medium (for plant culture) for 7 days and then roots were compared. The root length itself tends to be longer in the control than in the whole root, but the number of the root is higher in the transformant. This is because the plants are more focused on length growth in order to find water when they are stressed by salt, and they are more resistant to stress than those grown in stress tolerant plants. (Figs. 5 and 6)

The contents of Na in roots and the activity of vacuola pyrophosphatase were examined before and after stress. Since Na + ratio in vacuolate is known to be a key element of salt tolerance, Na + concentration was measured under salt stress condition in order to determine whether salt accumulation occurs in the whole tissues of transgenic plants. As a result, the sodium content increased after stress rather than before the stress, but the sodium content was higher in the transformant than in the control (FIG. 7). In addition, the phospholipase activity of vacuoles was also found to be higher in the transformants than in the control before and after stress (FIG. 8). It seems that it was able to tolerate salt stress by increasing the sodium content in the vacuole due to the active action of phosphorylase.

<2-3> Comparison of adaptive capacity and growth state in natural growth state .

Root observation, chlorophyll content and number of tillers were analyzed to compare the adaptability and growth state in the natural growth state. First, root length was measured every 4 days after transplantation, and it was found that the root length was longer in the transformant than in the control. (Figs. 9 and 10).

As shown in FIG. 11, the transformed OVP was significantly heavier than that of the wild type, as a result of comparing the dry weight and the dry weight of the transgenic and WT rice plants. From these results, it can be seen that in the case of the transgenic rice, the yield is significantly increased, which is a result that can be expected to increase the total yield.

<2-4> Comparison of yields in cultivated land

To investigate the effect of overexpression of OVP on agricultural characteristics, transgenic plants were cultivated during the cultivation period of 2010 (T2 generation) and 2011 (T3 generation) at Kyungpook National University (Gunwi campus, 36 ° 24 'N, 128 °) Field trials were conducted to investigate. Transgenic and WT rice plants grown for 90 days after germination in the greenhouse were transplanted into rice paddies and matured. Each plot contains 20 rice plants per row, with a row spacing of 0.2 m and a row spacing of 0.2 m. Control WT rice plants were placed between them (ie, every two plots). In the late reproductive stage, yield parameters of transformants were evaluated based on phenotype and agronomic traits. Yield parameters were evaluated based on 12 rice plants per plot and harvested by hand after the plants were matured. The immature and mature rice plants were independently isolated, counted, and weighed. The total plant biomass (g), stem weight (g), root weight (g), number of spikelets per plant, number of spikelets per spike, Grain filling rate (%), total grain weight (g) and average grain weight (MGW, g) were measured.

Fig. 15 is a visual observation of the rice grown in the cultivation area. It is evident that the OVP transgenic group shows better growth than the control group. As a result of the above experiment, it can be confirmed that the total plant biomass, stem, root, number of spines, seed weight, etc. are all significantly increased in the transformant. (Figs. 16A and 16B)

Statistical analysis

Significant differences between the transgenic and WT rice plants in the measured parameters were determined by ANOVA. Comparisons between individual data points were made using Student's t-test and P <0.05 was considered significant. All experiments were repeated 3 times and all results were expressed as mean ± standard deviation (SD).

The present invention has been described with reference to the preferred embodiments. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Therefore, the disclosed embodiments should be considered in an illustrative rather than a restrictive sense. The scope of the present invention is defined by the appended claims rather than by the foregoing description, and all differences within the scope of equivalents thereof should be construed as being included in the present invention.

<110> Kyungpook National Univercity <120> Use of VP1 gene from Oryza sativa as regulator of yield and          environmental stresses <130> PN1710-378 <160> 9 <170> KoPatentin 3.0 <210> 1 <211> 2742 <212> RNA <213> Oryza sativa <400> 1 gtcgaatcct cctccgagaa gaggagagta catgaatcct agcgcgagga tctcgcaggt 60 agcaatggcg gcgatactgc cggacctggc gacgcaggtg ctggtcccgg cggcggccgt 120 ggtcggcatc gcgttcgcgg tggtgcagtg ggtgctggtg tccaaggtga agatgaccgc 180 ggagcggcgc ggcggggagg ggtcccccgg cgcggccgcc gggaaggacg gcggcgccgc 240 cagcgagtac ctcatcgagg aggaggaggg gctcaacgag cacaacgtcg tcgagaagtg 300 ctccgagatc cagcacgcca tctccgaagg agcaacttct ttcctcttca ctgaatacaa 360 gtatgttgga ctattcatgg gcattttcgc ggtcctgatc ttcctcttcc ttggatctgt 420 tgagggattc agcacaaaga gtcaaccttg ccactacagc aaggatagga tgtgcaagcc 480 tgcccttgca aatgctatct tcagtactgt tgcctttgtg cttggtgcag tcacctctct 540 tgtatctggt ttccttggaa tgaagattgc aacttatgca aatgccagga caacccttga 600 ggcaaggaag ggtgttggga aggcattcat tactgctttc cgctctggtg ctgtcatggg 660 tttcctgctt gctgcaagtg gccttgttgt tcttttacatt gccatcaact tgttcggaat 720 ttattatggt gatgactggg aaggtctttt tgaggctatc actgggtatg gtcttggtgg 780 ttcttctatg gccctatttg gccgtgtagg tggcggtatt tacacgaaag ctgctgatgt 840 tggtgctgac ctagttggga aagtagaaag gaatatccct gaagatgacc caagaaaccc 900 agctgtcatt gcagataatg ttggtgacaa tgttggagat attgctggaa tggggtcaga 960 tctctttggt tcctatgctg agtcatcttg tgctgctctt gtcgttgctt caatctcttc 1020 ttttggaatt aaccatgaat tcactcctat gctgtacccg ctcctgatta gctcagttgg 1080 tatcattgct tgtctgataa ccacactttt cgccactgat ttttttgaga tcaaggctgt 1140 tgatgaaatt gagcctgccc tcaagaagca acttataatt tccaccgttg tgatgacggt 1200 tggcattgca cttgtcagtt ggttaggcct cccatattcc ttcacaatat tcaactttgg 1260 tgcccagaag actgtgtata actggcaact tttcttgtgt gtggcggttg gtctttgggc 1320 tggactaatc atcggatttg tgacagagta ttacacaagc aacgcctaca gccctgtcca 1380 agacgttgct gattcctgca gaaccggagc tgctactaat gtcatctttg gccttgctct 1440 gggatacaaa tctgtcatta ttcctatttt tgctattgct tttagtattt tcctcagctt 1500 cagccttgcc gcaatgtacg gtgtagctgt ggctgccctt ggaatgctta gcacaattgc 1560 aactggtctt gccattgatg cctatggccc tatcagtgac aatgctggag gaattgctga 1620 gatggctggc atgagtcaca gaattcgtga gagaactgat gctctggatg ctgcaggaaa 1680 caccactgct gccatcggga agggtttcgc cattggctcg gcagccttgg tgtctcttgc 1740 cctctttggt gcctttgtga gccgtgccgc catctccact gtcgatgttc tgacacctaa 1800 agttttcatt ggactgattg ttggtgctat gctcccatac tggttctctg ctatgaccat 1860 gaagagtgtt ggcagcgcag cgctcaagat ggtggaggaa gtccgtaggc agttcaactc 1920 catccctggg ctcatggagg gcacaaccaa gcctgactac gcaacctgtg tgaagatctc 1980 cactgatgca tccatcaagg agatgattcc cccaggtgct cttgtcatgc tcagccctct 2040 cattgttggg atcttctttg gtgtcgagac cctctcagga ctacttgctg gtgctctcgt 2100 ttctggtgtt cagattgcca tctctgcatc aaacactggt ggtgcctggg acaacgcaaa 2160 gaagtacatt gaggctggtg catctgagca tgcaaggacc ctgggcccga aaggctcaga 2220 ctgccacaag gctgctgtga tcggcgacac cattggggat cctctcaagg acacctcagg 2280 cccctcgctc aacatcctca tcaagctgat ggctgtcgag tcgctcgtgt ttgccccgtt 2340 cttcgccacc catggaggca tcctcttcaa gtggttttag atggtgacga tagatttacg 2400 aggacgttat ttttgccagg agggcgctgg aaatcatcca tatattatag agttcatcga 2460 gttgacacat atagttccct ccgtttccta tttgttatta ctgtctgtcc atcagtcgat 2520 catgctaagc ttattagcat catacctgtg tctagttttt ttgagtctgg tagttgtagc 2580 cgatgggagc aaatcactgg ccacctacag gtttggacaa gagtaccttg aatttgcgtt 2640 ttcctgtagg ttggatgata cgatgatggt gatgattatt gtacgtgcac ggaatgcacc 2700 tgtctcagtg aaattttccc gggaaggttc tgctgtttct tt 2742 <210> 2 <211> 27 <212> DNA <213> Oryza sativa <400> 2 cggctagcgc aatggcggcg atactgc 27 <210> 3 <211> 28 <212> DNA <213> Oryza sativa <400> 3 cggctagccc cttgggagct cgtaaatc 28 <210> 4 <211> 27 <212> DNA <213> Oryza sativa <400> 4 tgccttcata cgctatttat tgcttgg 27 <210> 5 <211> 27 <212> DNA <213> Oryza sativa <400> 5 gcgtgctgga tctcggagca cttctcg 27 <210> 6 <211> 20 <212> DNA <213> Oryza sativa <400> 6 tttggccttg ctctgggata 20 <210> 7 <211> 20 <212> DNA <213> Oryza sativa <400> 7 tgctggagga attgctgaga 20 <210> 8 <211> 20 <212> DNA <213> Oryza sativa <400> 8 taccgtgccc ttactgttcc 20 <210> 9 <211> 20 <212> DNA <213> Oryza sativa <400> 9 cggtggaatg tcacagacac 20

Claims (9)

Transforming a plant cell with a recombinant vector comprising a VP (vacuolar pyrophosphatase) gene derived from rice ( oryza sativa );
And regenerating the transformed plant. &Lt; RTI ID = 0.0 &gt; 21. &lt; / RTI &gt; Wherein the VP gene comprises the nucleotide sequence of SEQ ID NO: 1.  Transgenic plants or seeds produced by the method of claim 1. A composition for increasing the yield of a plant comprising a VP (vacuolar pyrophosphatase) gene derived from rice ( oryza sativa ). Transforming a plant cell with a recombinant vector comprising a VP (vacuolar pyrophosphatase) gene derived from rice ( oryza sativa );
And regenerating the transgenic plant, wherein the plant is resistant to environmental stress. Wherein the VP gene comprises the nucleotide sequence of SEQ ID NO: 1. 6. The method of claim 5, wherein the environmental stress is salt, cold or oxidative stress. Transgenic plants or seeds having increased tolerance to environmental stress produced by the method of claim 5. A composition for enhancing environmental stress tolerance of a plant, comprising a VP (vacuolar pyrophosphatase) gene derived from rice ( oryza sativa ).

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