CN112898392A - Application of rice PHI1 gene in regulation and control of plant photosynthesis - Google Patents

Abstract

The invention discloses an application of a rice PHI1 gene in regulation and control of plant photosynthesis. The invention provides an application of PHI1 protein or related biological materials thereof in regulating and controlling plant photosynthesis; the related biological material is a nucleic acid molecule for expressing PHI1 protein or an expression cassette, a recombinant vector, a recombinant bacterium or a transgenic cell line containing the nucleic acid molecule; the PHI1 protein is shown in SEQ ID No.1 or the protein with more than 80% of homology and the same function through substitution and/or deletion and/or addition of one or more amino acid residues, or the fusion protein obtained after connecting a label at the N end and/or the C end. Under the normal growth state and the drought condition, the expression level of the PHI1 gene in the target plant is reduced, and the photosynthetic efficiency of the plant can be obviously improved. The invention has important theoretical and practical significance for cultivating the high-light-efficiency rice material under the adverse conditions.

Description

Application of rice PHI1 gene in regulation and control of plant photosynthesis

Technical Field

The invention relates to the field of plant genetic engineering, in particular to application of a rice PHI1 gene in regulation and control of plant photosynthesis.

Background

In China, grain safety is always one of the important problems. Due to the increase of population and the continuous reduction of arable area, for important grain crops such as rice, the cultivation of higher-yield varieties is always the primary breeding target. Each breakthrough of rice yield cannot leave discovery and utilization of new and beneficial resources. In the 50 s of the 20 th century, the discovery of rice dwarf resources and the success of dwarf breeding realize the first leap of rice yield; in the 60-70 th of the 20 th century, the discovery of the sterile cytoplasm of the rice and the successful matching of the three lines enable the rice yield of China to realize the second leap; the cultivation of super hybrid rice successfully increased the yield of Chinese hybrid rice by 20% at the end of the 20 th century. The successful cultivation of super rice mainly utilizes ideal plant types and heterosis. In the ideal plant type, moderate curling of rice leaves is one of the important factors to consider, and the effect is fully reflected in high-yield and ultrahigh-yield breeding. For example, the curliness of the leaf reaches 60 percent and the curliness of the sword leaf reaches 39 percent of the E32 of two lines of super hybrid rice 'Eryou pejiu' and the perbush 64S/E32 backbone parent perbush 64S; the sword leaf curliness of the three-line super rice synergistic 9308 combination is about 15 percent. Research on the influence of the leaf rolling character on the leaf photosynthesis physiology, the group ecological effect and the economic character shows that the proper leaf rolling plays an important role in shaping good plant types of individuals, improving the group quality in the later growth period and increasing the yield. Cultivation of crops with high photosynthetic efficiency is one of the main targets of the planting industry. Currently, it is an effective method to obtain a variety with high photosynthetic efficiency by genetic engineering breeding, and the most key technical bottleneck problem in the method is the screening and function discovery of genes related to photosynthesis.

The constructed mutant library is used for obtaining related high-photosynthetic-efficiency mutants, cloning functional genes for controlling important agronomic characters of rice, providing a foundation for breeding and variety improvement of the rice and providing a theoretical basis for molecular breeding design of the rice. However, little is currently known about the molecular basis for controlling rice leaf traits.

Disclosure of Invention

The invention aims to provide application of a rice PHI1 gene in regulation and control of plant photosynthesis.

In a first aspect, the invention claims the use of PHI1 protein or its related biological material in any one of:

p1, regulating and controlling plant photosynthesis;

p2, regulating and controlling the plant photosynthetic rate;

p3, regulating and controlling the light respiration rate of the plant;

p4, regulating and controlling the plant height;

p5, regulating and controlling the curling degree of the plant leaves;

p6, regulating and controlling the content of phytoalexin;

p7, regulating and controlling the drought resistance of the plant.

The related biological material is a nucleic acid molecule capable of expressing the PHI1 protein or an expression cassette, a recombinant vector, a recombinant bacterium or a transgenic cell line containing the nucleic acid molecule.

The expression cassette refers to a DNA capable of expressing PHI1 in a host cell, which may include not only a promoter for initiating the transcription of PHI1 gene but also a terminator for terminating the transcription of PHI 1. Further, the expression cassette may also include an enhancer sequence. Promoters useful in the present invention include, but are not limited to: constitutive promoters, tissue, organ and development specific promoters, and inducible promoters. Examples of promoters include, but are not limited to: ubiquitin gene ubiqiutin promoter (pUbi); the constitutive promoter of cauliflower mosaic virus 35S; wound-inducible promoter from tomato, leucine aminopeptidase ("LAP", Chao et alHuman (1999) Plant Physiol 120: 979-; chemically inducible promoter from tobacco, pathogenesis-related 1(PR1) (induced by salicylic acid and BTH (benzothiadiazole-7-carbothioic acid S-methyl ester)); tomato proteinase inhibitor II promoter (PIN2) or LAP promoter (both inducible with jasmonic acid ester); heat shock promoters (U.S. patent 5,187,267); tetracycline-inducible promoters (U.S. Pat. No. 5,057,422); seed-specific promoters, such as the millet seed-specific promoter pF128(CN101063139B (Chinese patent 200710099169.7)), seed storage protein-specific promoters (e.g., the promoters of phaseolin, napin, oleosin, and soybean beta conglycin (Beachy et al (1985) EMBO J.4: 3047-3053)). They can be used alone or in combination with other plant promoters. All references cited herein are incorporated by reference in their entirety. Suitable transcription terminators include, but are not limited to: agrobacterium nopaline synthase terminator (NOS terminator), cauliflower mosaic virus CaMV 35S terminator, tml terminator, pea rbcS E9 terminator and nopaline and octopine synthase terminators (see, e.g., Odell et al (I)⁹⁸⁵) Nature313: 810; rosenberg et al (1987) Gene,56: 125; guerineau et al (1991) mol.gen.genet,262: 141; proudfoot (1991) Cell,64: 671; sanfacon et al Genes Dev.,5: 141; mogen et al (1990) Plant Cell,2: 1261; munroe et al (1990) Gene,91: 151; ballad et al (1989) Nucleic Acids Res.17: 7891; joshi et al (1987) Nucleic Acid Res, 15: 9627).

Constructing a recombinant expression vector containing the PHI1 gene expression cassette. The plant expression vector can be a Gateway system vector or a binary Agrobacterium vector, such as pGWB411, pGWB412, pGWB405, pBin438, pCAMBIA1302, pCAMBIA2301, pCAMBIA1301, pCAMBIA1300, pBI121, pCAMBIA1391-Xa or pCAMBIA 1391-Xb. When the GmbHLH664 is used for constructing a recombinant expression vector, any enhanced, constitutive, tissue-specific or inducible promoter can be added in front of the transcription initiation nucleotide, such as a cauliflower mosaic virus (CAMV)35S promoter, a ubiquitin gene Ubiqutin promoter (pUbi) and the like, and the promoters can be used alone or in combination with other plant promoters; in addition, when the gene of the present invention is used to construct plant expression vectors, enhancers, including translational or transcriptional enhancers, may be used, and these enhancer regions may be ATG initiation codon or initiation codon of adjacent regions, etc., but must be in the same reading frame as the coding sequence to ensure proper translation of the entire sequence. The translational control signals and initiation codons are widely derived, either naturally or synthetically. The translation initiation region may be derived from a transcription initiation region or a structural gene.

In order to facilitate the identification and screening of transgenic plant cells or plants, plant expression vectors to be used may be processed, for example, by adding a gene encoding an enzyme or a luminescent compound which can produce a color change (GUS gene, luciferase gene, etc.), an antibiotic marker having resistance (gentamicin marker, kanamycin marker, etc.), or a chemical-resistant marker gene (e.g., herbicide-resistant gene), etc., which can be expressed in plants.

The PHI1 protein is any one of the following proteins:

(A1) protein with an amino acid sequence of SEQ ID No. 1;

(A2) the amino acid sequence shown in SEQ ID No.1 is substituted and/or deleted and/or added by one or more amino acid residues and is derived from the protein with the same function of rice;

(A3) a protein having 99% or more, 95% or more, 90% or more, 85% or more, or 80% or more identity to the amino acid sequence defined in any one of (A1) to (A2) and derived from rice having the same function;

(A4) a fusion protein obtained by attaching a protein tag to the N-terminus and/or C-terminus of the protein defined in any one of (A1) to (A3).

In the above protein, the protein tag (protein-tag) refers to a polypeptide or protein that is expressed by fusion with a target protein using in vitro recombinant DNA technology, so as to facilitate expression, detection, tracking and/or purification of the target protein. The protein tag may be a Flag tag, a His tag, an MBP tag, an HA tag, a myc tag, a GST tag, and/or a SUMO tag, among others.

In the above proteins, identity refers to the identity of amino acid sequences. The identity of the amino acid sequences can be determined using homology search sites on the Internet, such as the BLAST web pages of the NCBI home website. For example, in the advanced BLAST2.1, by using blastp as a program, setting the value of Expect to 10, setting all filters to OFF, using BLOSUM62 as a Matrix, setting Gap existence cost, Per residual Gap cost, and Lambda ratio to 11, 1, and 0.85 (default values), respectively, and performing a calculation by searching for the identity of a pair of amino acid sequences, a value (%) of identity can be obtained.

In the above protein, the homology of 95% or more may be at least 96%, 97%, 98% identity. The homology of 90% or more may be at least 91%, 92%, 93%, 94% identity. The homology of 85% or more may be at least 86%, 87%, 88%, 89% identity. The homology of 80% or more may be at least 81%, 82%, 83%, 84% identity.

In P1-P6, the modulation may be modulation under normal growth conditions or under drought stress conditions.

Under normal growth conditions, the expression level and/or activity of the PHI1 protein or the coding gene thereof in the plant is reduced, the photosynthesis of the plant is enhanced, the photosynthetic rate is increased, the light respiration rate is reduced, the plant height is reduced, the leaf curl is increased and/or the chlorophyll content is increased.

Under drought stress conditions, the expression level and/or activity of the PHI1 protein or the coding gene thereof in the plant is reduced, the photosynthesis of the plant is enhanced, the photosynthetic rate is increased, the light respiration rate is reduced and/or the plant is increased.

In P7, the regulatory plant drought resistance is embodied as: under drought stress conditions, the expression level and/or activity of the PHI1 protein or the coding gene thereof in the plant is reduced, the photosynthesis of the plant is enhanced, the photosynthetic rate is increased, the light respiration rate is reduced and/or the plant is increased.

In a second aspect, the present invention claims the use of the PHI1 protein or its related biological material in plant breeding.

The related biological material is a nucleic acid molecule capable of expressing the PHI1 protein or an expression cassette, a recombinant vector, a recombinant bacterium or a transgenic cell line containing the nucleic acid molecule. The PHI1 protein can be any one of the proteins shown in the above (A1) - (A4).

Further, in such applications, plants that do not contain the PHI1 or that are low in PHI1 can be crossed with other plants to develop plant varieties that "increase photosynthesis, increase photosynthesis rate, decrease photorespiration rate, decrease plant height, increase leaf curliness, and/or increase chlorophyll content under normal growth conditions" and/or "increase photosynthesis, increase photosynthesis rate, decrease photorespiration rate, and/or increase plant height under drought stress conditions" and/or "increase drought resistance".

In a third aspect, the invention claims a method of breeding a plant variety.

The method for breeding a plant variety as claimed in the present invention is a method for breeding a plant variety having at least one of the traits indicated by Q1 to Q3, and may comprise a step of reducing the expression level and/or activity of the PHI1 protein in a recipient plant. The PHI1 protein can be any one of the proteins shown in the above (A1) - (A4).

Q1, increased photosynthesis, increased photosynthetic rate, decreased light respiration rate, decreased plant height, increased leaf curliness and/or increased chlorophyll content under normal growth conditions;

q2, increased photosynthesis, increased photosynthetic rate, decreased light respiration rate and/or increased plant height under drought stress conditions;

q3, drought resistance is enhanced.

Specifically, this can be achieved by hybridization means or by transgenic means.

In a fourth aspect, the invention claims a method of breeding transgenic plant varieties.

The method for breeding a transgenic plant variety claimed by the invention is a method for breeding a transgenic plant with at least any one of the following traits Q1-Q3, and comprises the following steps: inhibiting and expressing a nucleic acid molecule capable of expressing PHI1 protein in a receptor plant to obtain a transgenic plant; the transgenic plant has at least any one of the traits set forth in Q1-Q3 below as compared to the recipient plant. The PHI1 protein can be any one of the proteins shown in the above (A1) - (A4).

Q1, increased photosynthesis, increased photosynthetic rate, decreased light respiration rate, decreased plant height, increased leaf curliness and/or increased chlorophyll content under normal growth conditions;

q2, increased photosynthesis, increased photosynthetic rate, decreased light respiration rate and/or increased plant height under drought stress conditions;

q3, drought resistance is enhanced.

Wherein the inhibition of expression of the nucleic acid molecule capable of expressing the PHI1 protein in the recipient plant is achieved by RNA interference techniques.

Further, the RNA interference vector introduced into the recipient plant is a recombinant expression vector containing an interference fragment.

The interference fragment comprises a segment A, a segment B and a segment C from the 5 'end to the 3' end in sequence; the nucleotide sequence of the segment A is 238 th and 532 nd position of SEQ ID No. 2; the segment C is reverse complementary to the segment A; the segment B is not complementary to both the segment a and the segment C.

In a specific embodiment of the present invention, the interference vector is specifically a recombinant vector obtained by inserting the segment a between SpeI and SacI of pTCK309 vector and inserting the segment C between KpnI and BamHI of pTCK309 vector.

In the above method, the introduction of the interference vector into the recipient plant may specifically be: plant cells or tissues are transformed by conventional biological methods using Ti plasmids, Ri plasmids, plant viral vectors, direct DNA transformation, microinjection, conductance, agrobacterium mediation, etc., and the transformed plant tissues are grown into plants.

In the above methods, the transgenic plant is understood to include not only the first to second generation transgenic plants but also the progeny thereof. For transgenic plants, the gene can be propagated in the species, and can also be transferred into other varieties of the same species, including particularly commercial varieties, using conventional breeding techniques. The transgenic plants include seeds, callus, whole plants and cells.

In each of the above aspects, the nucleic acid molecule may be DNA, such as cDNA, genomic DNA, or recombinant DNA; the nucleic acid molecule may also be RNA, such as mRNA, and the like.

Further, the nucleic acid molecule capable of expressing the PHI1 protein can be specifically any one of the following DNA molecules:

(B1) a DNA molecule shown in SEQ ID No.2 or SEQ ID No.3 at position 366-2281;

(B2) a DNA molecule which hybridizes with the DNA molecule defined in (B1) under stringent conditions and encodes the PHI1 protein;

(B3) a DNA molecule which has more than 99%, more than 95%, more than 90%, more than 85% or more than 80% of identity with the DNA sequence limited by (B1) or (B2) and encodes the PHI1 protein.

In the above nucleic acid molecule, the stringent conditions may be as follows: 50 ℃ in 7% Sodium Dodecyl Sulfate (SDS), 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing in2 XSSC, 0.1% SDS at 50 ℃; also can be: 50 ℃ in 7% SDS, 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing at 50 ℃ in 1 XSSC, 0.1% SDS; also can be: 50 ℃ in 7% SDS, 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing in 0.5 XSSC, 0.1% SDS at 50 ℃; also can be: 50 ℃ in 7% SDS, 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing in 0.1 XSSC, 0.1% SDS at 50 ℃; also can be: 50 ℃ in 7% SDS, 0.5M Na₃PO₄Hybridization with 1mM EDTA, rinsing in 0.1 XSSC, 0.1% SDS at 65 ℃; can also be: in a solution of 6 XSSC, 0.5% SDS at 65 ℃ and then washed once with each of 2 XSSC, 0.1% SDS and 1 XSSC, 0.1% SDS.

In the above nucleic acid molecules, homology means the identity of nucleotide sequences. The identity of the nucleotide sequences can be determined using homology search sites on the Internet, such as the BLAST web page of the NCBI home website. For example, in the advanced BLAST2.1, by using blastp as a program, setting the value of Expect to 10, setting all filters to OFF, using BLOSUM62 as a Matrix, setting Gap existence cost, Per residual Gap cost, and Lambda ratio to 11, 1, and 0.85 (default values), respectively, and performing a calculation by searching for the identity of a pair of nucleotide sequences, a value (%) of identity can be obtained.

In the above nucleic acid molecule, the homology of 95% or more may be at least 96%, 97%, 98% identity. The homology of 90% or more may be at least 91%, 92%, 93%, 94% identity. The homology of 85% or more may be at least 86%, 87%, 88%, 89% identity. The homology of 80% or more may be at least 81%, 82%, 83%, 84% identity.

In a fifth aspect, the invention claims any of the following applications:

use of R1, an interfering fragment as hereinbefore described or an interfering vector as hereinbefore described for growing plants having at least any one of the following traits: under normal growth conditions, photosynthesis is enhanced, photosynthetic rate is increased, light respiration rate is reduced, plant height is reduced, leaf curling degree is increased and/or chlorophyll content is increased; under drought stress conditions, photosynthesis is enhanced, photosynthetic rate is increased, photorespiration rate is reduced and/or plants are increased; drought resistance is enhanced;

use of R2, an interfering fragment as hereinbefore described or an interfering vector as hereinbefore described in plant breeding;

r3, use of the method as hereinbefore described in plant breeding.

In the invention, the drought resistance enhancement is embodied in that under the drought stress condition, the photosynthesis is enhanced, the photosynthetic rate is increased, the light respiration rate is reduced and/or the plant is increased.

In the present invention, the chlorophyll is chlorophyll a and/or chlorophyll b.

In each of the above aspects, the plant may be a monocot or a dicot.

Further, the monocot plant may be a plant of the order gramineae.

Further, the plant of the order gramineae may be a gramineae.

More specifically, the gramineous plant may be a plant of the genus oryza, such as rice.

In a particular embodiment of the invention, the rice is in particular a japonica rice.

The invention screens and obtains a mutant scd1 with obviously improved photosynthetic efficiency from the constructed rice T-DNA inserted mutant library, the mutant has the advantages that the photosynthetic efficiency is improved, the plant morphology is represented as half dwarfing (3/4 of the wild plant height), the leaves are curled inwards to be semi-cylindrical, the chlorophyll content is obviously improved compared with the wild plant, and the photosynthetic efficiency of the plant is further improved. Genetic analysis shows that the mutant character is controlled by a recessive single gene, a gene for controlling the mutant character is separated by using a site cloning method, and a wild type gene of the gene is a RCD1 family gene (named PHI 1). Meanwhile, the invention uses a complementary method to carry out function complementary research on the gene and restore the phenotype of the mutant.

Experiments prove that under the normal growth state, the expression level of the PHI1 gene in a target plant is reduced, and the photosynthetic efficiency of the plant can be obviously improved. In addition, under the drought condition, the reduction of the expression of the PHI1 gene can also improve the plant height and the photosynthetic efficiency of the plant. The invention has important theoretical and practical significance for cultivating high-photosynthetic-efficiency rice materials, particularly for cultivating the high-photosynthetic-efficiency rice materials under adverse conditions.

Drawings

FIG. 1 shows the results of plant height and leaf camber in example 1.

FIG. 2 is a graph showing the statistical results of the chlorophyll content in example 1. .

FIG. 3 is a map-based clone of the PHI1 gene in example 2.

FIG. 4 is a gene structural diagram of the PHI1 gene in example 2.

FIG. 5 shows the results of the detection of the transgenic complementation expression plants in example 2.

FIG. 6 shows the result of testing the transgenic plants with PHI1 loss-of-function in example 3.

FIG. 7 shows that the PHI1 deficient plants in example 4 can increase the plant height and fruit of the plants under drought conditions.

Detailed Description

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

The experimental procedures in the following examples, unless otherwise indicated, are conventional and are carried out according to the techniques or conditions described in the literature in the field or according to the instructions of the products. Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.

Japonica rice (Oryza sativa l. ssp. japonica): department of seed Ministry of Chinese academy of agricultural sciences.

Mutant scd 1: mutant scd1 was obtained by performing T-DNA insertion mutation on rice nippon (Oryza sativa l. ssp. japonica), reference: wan S, Wu J, Zhang Z, et al. activation tagging, an effective tool for functional analysis of the edge gene [ J ]. Plant Molecular Biology,2009,69(2): 69-80.; the public is available from the institute of biotechnology, academy of agricultural sciences, china.

Wild type indica rice variety durar (oryzasatival l. ssp. indica): the public is available from the institute of biotechnology, academy of agricultural sciences, china.

Example 1 acquisition of PHI1 protein and Gene encoding the same

Phenotypic analysis of mutant scd1

The plant height and leaf curl of the mutant scd1 and the wild rice Nipponbare are measured, and the chlorophyll A content, the chlorophyll B content and the photosynthetic rate are detected at the same time.

Measuring the plant height: maturity is measured as the height from the ground surface to the top of the highest ear of an individual (without miscanthus). The same holds true for the data of the various sections (sections from top to bottom).

Curl (Leaf rolling index LRI) — maturity time determination, [ Lw (post-expansion Leaf-edge spacing) -Ln (natural distance from Leaf to Leaf edge measured at the same site) ]/Lw (post-expansion Leaf-edge spacing) × 100%.

Photosynthetic rate determination: one week after the ear initiation, selecting a sunny day with the air temperature (daily average temperature) higher than 30 ℃,9 in the morning of each day: 00, measuring the photosynthetic rate of the sword leaves of the plant to be measured, wherein the measured part is the front l/3 part of the sword leaves; each measurement took 1 minute; the measurement of each group of materials is completed within 1 hour in time; 13 on the same day: 00 start to repeat the measurement of the morning; repeating the measurement for 2 times within the next week; the photosynthetic determination system is a single U.S. Li-CO 6400 portable photosynthetic rate determinator, so that the determinator can automatically determine net photosynthetic rate, light respiration rate and CO₂Conductance of the air hole. The results are the mean of multiple replicates, each index being measured more than 5 times.

Measuring the chlorophyll content: the chlorophyll content was measured by acetone extraction. The method comprises the following specific steps: shearing rice leaves at the same growth part, accurately weighing, shearing into 2mm small fragments, and placing in acetone: soaking in 1:1 (volume ratio) ethanol solution (10 ml). Soaking for 24 hr to whiten the leaves, measuring OD of chlorophyll extractive solution with mixed extractive solution of acetone and ethanol as blank control_663nmAnd OD_645nm. The chlorophyll extraction and determination are carried out under low-temperature dark conditions. Each sample was set to 3 replicates. Absorbance of chlorophyll solution at 663nm and 645nm according to Lambert-Beer's law (A)₆₆₃And A₆₄₅) And the chlorophyll a, b and total concentration in the solution can be expressed by the following equations:

the chlorophyll a content was calculated using the following formula: chlorophyll a (mg/L) ═ 12.7 XA₆₆₃nm－2.69×A₆₄₅nm) x dilution factor;

the chlorophyll b content was calculated using the following formula: chlorophyll b (mg/L) ═ 22.9 XA₆₄₅nm－4.64×A₆₆₃nm) x dilution factor;

the total chlorophyll content was calculated using the following formula: chlorophyll a + b (mg/L) ═ 20.2 XA₆₄₅nm+8.02×A₆₆₃nm) x dilution factor.

The results are shown in fig. 1, fig. 2 and table 1. FIG. 1 shows the results of plant height and leaf camber. In FIG. 1, A is the observation result of phenotype, B in FIG. 1 is the transverse section of the leaf in the mature period of the wild type, C in FIG. 1 is the transverse section of the leaf in the mature period of the mutant scd1, D in FIG. 1 is the statistical result of plant height (I-IV are the number of each node of the rice stem), and E in FIG. 1 is the statistical result of the curling degree of the leaf. FIG. 2 is a graph showing the statistical results of chlorophyll content. Table 1 shows photosynthetic efficiency, CO₂And (5) counting the stomatal conductance and the light respiration rate.

The results show that mutant scd1 strain is 3/4 (a and D in fig. 1) of wild type (Wt) and about 70 cm in height. Mutant scd1 leaf curliness was 14% (E in fig. 1). The chlorophyll a and chlorophyll B content of mutant scd1 was significantly increased compared to Wild Type (WT) (fig. 2), and the photosynthetic efficiency of mutant scd1 was also significantly increased (table 1).

TABLE 1 photosynthetic efficiency, CO₂Statistical results of stomatal conductance and light respiration rate

Note: compared to wild type, represents P <0.01, with very significant difference.

Second, location cloning of PHI1 Gene

F is obtained by hybridizing the mutant scd1 and indica rice Dular (Oryzasatival L. ssp. indica) as parents₁Generation, F₁Generation selfing to produce F₂And (4) separating the populations. F₂And after the generation plant grows for 90 days, taking the leaf of the mutant plant to extract genome DNA for gene localization. First, 100F are taken₂The mutant plants are roughly positioned, molecular markers with better polymorphism are selected on 12 linkage groups of rice approximately every 20cM, linkage analysis is carried out on 40 mutant plants, and after the molecular markers linked with scd1 are found, further verification is carried out by 200 mutant plants. After the verification is correct, the large population carries out chromosome walkingAnd fine positioning the target gene in one step. The molecular markers used for localization are shown in Table 2 (5 '-3'). A total of 3000F₂Mutant plants localize scdl in the 45K segment, a total of 4 complete ORFs are predicted in this segment, and the 4 ORFs are subjected to gene structure analysis and all sequencing analysis to determine candidate genes. The screening process and results are shown in fig. 3 and 4. In FIG. 3, the thick horizontal line represents the chromosome, the short vertical line represents the molecular markers on the chromosome, Indel1-1 to Indel1-4 are the names of the primary localization molecular markers, and 2252, 2269, 2285, 2290, 2301 are the fine localization molecular markers. AC018929, AC027037 and AC018727 are BAC clones on chromosome 4 of rice. In FIG. 4, A in FIG. 4 is a diagram showing the structure of the target gene and the position of the inserted gene. In FIG. 4, B is the amino acid change caused by the three domains and insertion sites of the target protein.

TABLE 2 molecular markers for localization

Three, PHI1 protein and coding gene thereof

Through the research, the protein PHI1 related to the plant photosynthetic rate and the coding gene PHI1 thereof are discovered. The protein shown in SEQ ID No.1 is named as protein PHI1 and consists of 591 amino acid residues.

The gene encoding the protein PHI1 was designated as PHI1 gene. The PHI1 gene is shown in SEQ ID No. 2.

Example 2 construction of transgenic complementation expression plants and detection of their traits

1. Construction of recombinant expression vector pCambia23A-PHI1

The SEQ ID No.3 (derived from rice cDNA) is amplified by TWI1-F and TWI1-R, the obtained product is cut by SmaI/XbaI, then is cut by XbaI and SmaI enzyme with pCambia23A vector (Changsheng Biotechnology Limited liability company, Beijing) for connection, and the pCambia23A-PHI1 recombinant expression vector is obtained and is verified to be correct by sequencing.

TWI1-F-5'SmaI：5'-TTCCCGGGGTGTAAATAGTAGGCTTGTTGGAG-3'；

TWI1-R-3'XbaI：5'-CCTCTAGAAAGCTTCTCTCCACTAGTCAAGTC-3'。

2. And (2) introducing the pCambia23A-PHI1 recombinant expression vector obtained in the step (1) into Agrobacterium AGL1 (Beijing Dingguo Changsheng biotechnology, Limited liability company) to obtain recombinant Agrobacterium.

3. Introducing the recombinant agrobacterium obtained in the step 2 into a transformation mutant scd1 by an electric excitation method, and harvesting T₀And (5) plant generation.

4. For the wild type rice Nipponbare, the mutant scd1 and the T constructed in the step 3₀The generation plants (CP1, CP2 and CP3) were subjected to phenotypic observation and PHI1 gene expression amount analysis at the flowering stage.

PHI1 gene expression level analysis: extracting total RNA of the plants and carrying out reverse transcription to obtain cDNA, carrying out real-time fluorescence quantitative PCR detection on the expression quantity of the PHI1 gene in each plant by taking the cDNA as a template, taking an Actin gene as an internal reference gene, taking the expression quantity of the PHI1 gene in wild Nipponbare as 1, and calculating the relative expression quantity of the PHI1 gene in other plants.

The primer sequences for identifying the PHI1 gene are as follows:

146dlF：5’-ATGTAAGCGTGGCAATGCA-3’；

146dlR：5’-GAGTAATTCACACAGGTATTGGAGC-3’。

the primer sequences for identifying the Actin gene are as follows:

actinF：5’-TGCTATGTACGTCGCCATCCAG-3’；

actinR：5’-AATGAGTAACCACGCTCCGTCA-3’。

the results are shown in FIG. 5. In FIG. 5, A is the phenotypic observation. B in FIG. 5 is the statistical result of the relative expression of PHI1 gene. The result shows that the phenotype of the transgenic plant CP1-CP3 is the same as that of the wild type, and the relative expression quantity of the PHI1 gene has no significant difference.

Example 3 establishment and phenotypic Observation of PHI1 loss-of-function transgenic plants

First, construction of interfering plants

1. Taking cDNA of wild rice Nipponbare as a template, and carrying out PCR amplification by adopting a primer P5utrF and a primer P5utrR to obtain a PCR amplification product.

P5utrF：5’-GGGGTACCACTAGTGCTTTACCTAAGCGAATTCT-3’；

P5utrR：5’-CGGGATCCGAGCTCATACATTATTACCATTTCC-3’。

In primer P5utrF, the underlined kpnI and SpeI restriction sites are marked;

in primer P5utrR, the BamH I and Sac I sites are underlined.

2. And (3) connecting the PCR amplification product obtained in the step (1) with a pEasy vector (Beijing Ding Guoshang biotechnology, Inc.) to obtain a connection product pEasy-PHI 1.

3. The ligation product pEasy-PHI1 of step 2 was double-digested with SpeI and SacI restriction enzymes, and an digested product of about 0.3Kbp was recovered.

4. The pTCK309 vector (Invitrogen) was digested with SpeI and SacI restriction enzymes to give a vector backbone of about 8 Kbp.

5. And (4) connecting the enzyme digestion product obtained in the step (3) with the vector skeleton obtained in the step (4) to obtain the recombinant plasmid pTCK 309-X.

6. The ligation product pEasy-PHI1 of step 2 was digested simultaneously with KpnI and BamHI restriction enzymes, and an digested product of about 0.3Kbp was recovered.

7. The recombinant plasmid pTCK309-X obtained in step 5 was digested with KpnI and BamHI restriction enzymes to obtain a vector fragment of about 8 Kbp.

8. And (3) connecting the enzyme digestion product obtained in the step (6) with the vector fragment obtained in the step (7) to obtain an interference vector RNAi-309, and verifying the correctness through sequencing.

9. And (3) introducing the interference vector RNAi-309 obtained in the step (8) into Agrobacterium AGL1 to obtain recombinant Agrobacterium.

10. Transforming the recombinant agrobacterium obtained in the step 9 into wild rice Nipponbare through an electric excitation method, and harvesting T₀The generations interfered with the plants.

Second, construction of empty vector plants

Replacing interference vector RNAi-309 with pTCK309 vector, and performing operation according to 9 and 10 in step one to obtain empty vector T₀And (5) plant generation.

Third, character identification

And (3) the plant to be detected: wild type rice Nipponbare, mutant scd1 and T constructed in step one₀And (3) generation interference plants (randomly selecting 3 strains numbered as RNAi-1, RNAi-2 and RNAi-3) and empty transfer vector plants constructed in the second step.

1. And observing the phenotype of the flowering period of the plant to be detected.

2. And detecting the relative expression quantity of the PHI1 gene of the plant to be detected (see example 2 for a specific method).

3. Detecting net photosynthetic rate, light respiration rate and CO of plant to be detected₂Conductance of the air hole. See example 1 for a specific method).

The results are shown in FIG. 6 and Table 3. In FIG. 6, A is a photograph of a phenotype; b is relative expression level of PHI1 gene.

TABLE 3 net photosynthetic Rate, Photorespiratory Rate, intercellular CO₂Concentration detection result

Note: compared to wild type, represents P <0.01, with very significant difference.

Compared with the wild type, the photosynthetic rate of the interference plants (RNAi-1, RNAi-2 and RNAi-3) is improved to different degrees, the photosynthetic efficiency of the empty vector transfer plant has no obvious difference with the wild type, and the result shows that the photosynthetic efficiency can be improved by inhibiting the expression of PHI1 gene.

Example 4 PHI1 function-deficient plants can improve the high photosynthetic efficiency of seedlings under drought conditions

T of 3 high-generation transgenic lines is selected₂The generation plants (3 strains of RNAi-1 strain, RNAi-2 strain and RNAi-3 strain, each strain) and Nipponbare (3 strains) of the rice variety were subjected to the following drought resistance identification (mannitol treatment test), respectively:

1. plants with consistent germination and growth states are taken out, transferred into 0, 150 and 200mM mannitol solution, and replaced by new mannitol aqueous solution every 3 days, at which time the height of seedlings can be observed.

2. When the seedlings grow to 8 days, the growth conditions of seedlings of 3 interference plants (RNAi-1, RNAi-2 and RNAi-3) are obviously different from those of the wild plants, pictures are taken, the picture is shown as A in figure 7, the statistical result of the average seedling length is shown as B in figure 7, and the result of the seedling growth condition is shown in table 4.

Table 4 growth conditions of the respective lines under different concentrations of mannitol (plant height: cm)

	0mM	150mM	200mM
				Nippon nitrile control WT	15.361	5.493	1.113
RNAi-1	15.89	11.2	3.583**
				RNAi-2	16.1	8.833	2.817**
RNAi-3	15.856	7.65	1.4*

Note: p <0.01, represents significant and the difference is very significant compared to wild type.

3. When the plant grows to the ten-leaf stage, the LiCOR6400 photosynthetic instrument is used for carrying out photosynthetic parameters, and the net photosynthetic rate, the light respiration rate and the CO of the plant to be detected are detected₂See example 1 for specific methods for porosity conductance). The results are shown in Table 5.

TABLE 5 Net photosynthetic Rate, Photorespiring Rate, CO₂Air hole conductivity detection result

Note: compared to wild type, represents P <0.01, with very significant difference.

Compared with the wild type, the photosynthetic rate of the interfering plants (RNAi-1, RNAi-2 and RNAi-3) is improved to different degrees, and the result shows that the photosynthetic efficiency can be improved by inhibiting the expression of PHI1 gene under drought conditions (Table 5). The PHI1 gene is shown to have potential application prospect under drought and other conditions (mannitol).

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

<110> institute of biotechnology of Chinese academy of agricultural sciences

Application of PHI1 gene of rice in regulation and control of plant photosynthesis

<130> GNCLN210532

<160> 3

<170> PatentIn version 3.5

<210> 1

<211> 591

<212> PRT

<213> Oryza sativa

<400> 1

Met Ala Ala Met Asn Glu Lys Val Leu Asp Lys Cys Gly Arg Asn Ile

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Ser Ser Leu Lys Arg Lys Arg Asp Asn Pro Ala Ala Arg Cys Ala Asp

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Ala Gly Asn Thr Ser Lys Leu His Lys His Pro Ala Asp Asn Ser Val

35 40 45

Val Arg Phe Tyr Val Asp Glu Gly His Lys Ala Lys Ile Lys Cys His

50 55 60

Phe Lys Met Gln Ile Ile Gln Ser Tyr Gln Asn Phe Met Thr Ser Ala

65 70 75 80

Leu Pro Lys Arg Ile Leu Leu Arg Gln Gly Gly Glu Trp Lys Asp Phe

85 90 95

Pro Lys Gln Ile Val Lys Leu Ala His Ser Asp Phe Arg Thr Lys Lys

100 105 110

Thr Ile Thr Glu Gly Glu His Gln Thr His Leu Phe Leu Leu Asp Phe

115 120 125

Val His Met Thr Phe Ile Asp Ser Lys Thr Gly Leu Gln Arg Pro Ile

130 135 140

Ala Trp Ile Asp Glu Asn Gly Lys Gln Tyr Phe Pro Glu Phe Phe Ile

145 150 155 160

Glu Asp Lys Thr Leu Tyr Arg Lys Lys Glu Leu Gly Asn Gly Asn Asn

165 170 175

Val Tyr Ile Ile Val Glu Pro Asn Gly Thr Gln Glu Met Asn Asp His

180 185 190

Phe Gly Thr Ser Glu Ser Ser Ala Glu Ser Ser Asn Phe Glu Ser Ser

195 200 205

Thr Asp Asp Val Ser Ser Pro Lys Arg Ala Lys Ala Glu Arg Ser Val

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Ala Gly Asn Lys Thr Gly Gly Val Lys Glu Thr Ile Gly Glu Asn Glu

225 230 235 240

Pro His Ala Leu Leu Pro Ile Pro Cys Arg Ser Leu Pro Gln Asp Lys

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Leu Gly Asp His Ser Arg Val Gln Leu Ala Ile Ser Ala Val Gln Lys

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

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Gly Ile Tyr Arg Thr Pro Ala Val Asp Asn His Lys Glu Phe Arg Tyr

290 295 300

Asn Leu Phe Lys Lys Gln Ala Glu His Thr Lys Cys Lys Arg Gly Asn

305 310 315 320

Ala Asn Val Arg Tyr Ala Trp Leu Ala Cys Ser Lys Asp Ala Val Asp

325 330 335

Glu Met Met Leu Asn Gly Val Met His Phe Glu Lys Thr Val Lys Cys

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Pro Asp Tyr Gly Ile Gly Thr Ile Leu Ala Pro Ala Asn Cys Ser Asn

355 360 365

Thr Cys Val Asn Tyr Ser Asp Val Asp Glu Asn Gly Ile Val His Met

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Met Leu Cys Arg Val Val Met Gly Asn Val Glu Ile Val His His Gly

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Ser Lys Gln His Arg Pro Ser Asn Glu Tyr Phe Asp Ser Gly Val Asp

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Asp Ile Lys Asn Pro Gln His Tyr Ile Val Trp Asp Met Asn Val Asn

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Ser His Ile Tyr Ser Glu Phe Val Val Thr Ile Lys Leu Pro Ser Arg

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Val Lys Asp Ser Pro Ala Thr Glu Glu Asp Cys His Asn Leu Ser Glu

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Val Ser Ser Leu Ile Leu Ser Ser Gly Ser Pro Asp Ser Val Ser Gln

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Asp Met Asn Leu Gln Ala Ser Pro Ala Leu Gly Gly His Tyr Glu Ala

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Pro Met Leu Gly Asp Lys Val Glu Arg Ala Pro Ser Thr Pro Trp Met

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Pro Phe Ser Met Leu Phe Ala Ala Ile Ser Thr Lys Val Ser Ala Glu

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Asn Met Asp Met Val Asn Ser Cys Tyr Glu Glu Phe Lys Ser Lys Lys

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Ile Ser Arg Val Asp Leu Val Lys Lys Leu Arg His Ile Val Gly Asp

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Arg Met Leu Ile Ser Thr Ile Met Arg Leu Gln Asp Lys Leu Pro Pro

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Met Ser Arg His Glu Ala Pro Lys His Val Gly Gln Asp Asp Gly

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<210> 2

<211> 1776

<212> DNA

<213> Oryza sativa

<400> 2

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aagcatcccg ccgataactc tgttgttagg ttttatgttg atgaaggtca caaggcaaag 180

atcaagtgcc atttcaaaat gcagattata cagagttatc agaatttcat gactagtgct 240

ttacctaagc gaattctgct tcgtcaaggt ggtgaatgga aagactttcc aaagcaaatt 300

gtcaagcttg cccacagtga ttttaggact aagaagacaa tcacagaagg tgaacaccag 360

acccatcttt ttttgctgga ttttgtgcac atgacattca tagactcaaa gacaggcctt 420

cagaggccaa tagcttggat tgatgagaat gggaagcagt acttcccaga attttttatt 480

gaagataaga cactgtatag gaaaaaggaa ttgggaaatg gtaataatgt atacataatt 540

gtcgagccca atgggacaca agaaatgaac gaccactttg gaacatcaga aagttcagca 600

gaaagctcga attttgaatc tagcactgat gatgtttcca gccctaagag agccaaggct 660

gagaggagtg tcgcaggaaa caaaactggt ggtgtcaaag aaactattgg agagaatgag 720

ccacacgcct tgctgcctat tccctgcaga tctctaccac aagataagct gggtgaccat 780

tcacgtgttc aactagctat ttctgctgtt cagaaattgt tgctgcaggg tctgggcact 840

gttcttggtt ccaaggacat cgttgggatc tacagaacac cagcagtaga taaccataaa 900

gaatttcgtt ataatctttt caaaaagcag gcagagcata ctaaatgtaa gcgtggcaat 960

gcaaatgtcc gctatgcctg gcttgcttgc tcaaaagatg ctgtggatga aatgatgttg 1020

aatggtgtca tgcactttga gaagactgtt aagtgcccag actacggaat tggcacaatc 1080

cttgctccag caaattgctc caatacctgt gtgaattact ctgatgttga tgaaaatggc 1140

attgtccata tgatgttgtg ccgtgttgta atggggaacg tagaaatagt tcaccatgga 1200

tctaagcagc atcggcctag taacgagtat ttcgatagtg gtgtagatga cattaaaaac 1260

cctcaacatt acattgtgtg ggatatgaat gtgaatagtc acatctattc tgaatttgta 1320

gtcaccatca aattgccttc tagagtcaaa gattctcctg ccaccgaaga agattgtcac 1380

aatttatcag aggtttcatc actgatcttg agctctggtt cgcctgatag tgtatcacag 1440

gacatgaacc ttcaggcttc tccagcattg ggaggtcatt atgaagcccc catgttagga 1500

gataaagtgg aaagggctcc aagcacgcct tggatgcctt tctccatgtt atttgcagcg 1560

atttcaacca aagtttctgc tgagaatatg gacatggtca acagttgtta cgaagaattc 1620

aagagcaaga aaataagccg agtcgaccta gtgaagaagt taaggcatat agttggtgac 1680

agaatgctta tatcaacaat aatgcgactc caagataagt tgccccctat gtcgaggcat 1740

gaagcaccca aacacgtggg ccaagatgat ggctaa 1776

<210> 3

<211> 2553

<212> DNA

<213> Oryza sativa

<400> 3

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cctactgcct aaagtaccac acaaactacg tttatgctag taacagtggc tattgttttg 180

agcgactttg gcatgcagtc tgtcgtattg atgatttaaa atattgtgtt tttgaagtgt 240

tctctaatat ctggctttca ttttcaggat catttcgctt ctgcctcttt ctgacccgac 300

tctttttggt ttttgcctcg tcgataatga acttgagtac actcagaagc ttgcctatta 360

gcttagtgta aatagtaggc ttgttggagt ttagtttgtt agaaattatt agtctctggt 420

tcatttcagc tagcaaccat ggctgcgatg aacgaaaagg tattggataa atgtggaaga 480

aacataagta gcctcaagag aaagcgggac aacccagctg cacgttgcgc tgatgccggc 540

aacacctcta aattgcataa gcatcccgcc gataactctg ttgttaggtt ttatgttgat 600

gaaggtcaca aggcaaagat caagtgccat ttcaaaatgc agattataca gagttatcag 660

aatttcatga ctagtgcttt acctaagcga attctgcttc gtcaaggtgg tgaatggaaa 720

gactttccaa agcaaattgt caagcttgcc cacagtgatt ttaggactaa gaagacaatc 780

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gactcaaaga caggccttca gaggccaata gcttggattg atgagaatgg gaagcagtac 900

ttcccagaat tttttattga agataagaca ctgtatagga aaaaggaatt gggaaatggt 960

aataatgtat acataattgt cgagcccaat gggacacaag aaatgaacga ccactttgga 1020

acatcagaaa gttcagcaga aagctcgaat tttgaatcta gcactgatga tgtttccagc 1080

cctaagagag ccaaggctga gaggagtgtc gcaggaaaca aaactggtgg tgtcaaagaa 1140

actattggag agaatgagcc acacgccttg ctgcctattc cctgcagatc tctaccacaa 1200

gataagctgg gtgaccattc acgtgttcaa ctagctattt ctgctgttca gaaattgttg 1260

ctgcagggtc tgggcactgt tcttggttcc aaggacatcg ttgggatcta cagaacacca 1320

gcagtagata accataaaga atttcgttat aatcttttca aaaagcaggc agagcatact 1380

aaatgtaagc gtggcaatgc aaatgtccgc tatgcctggc ttgcttgctc aaaagatgct 1440

gtggatgaaa tgatgttgaa tggtgtcatg cactttgaga agactgttaa gtgcccagac 1500

tacggaattg gcacaatcct tgctccagca aattgctcca atacctgtgt gaattactct 1560

gatgttgatg aaaatggcat tgtccatatg atgttgtgcc gtgttgtaat ggggaacgta 1620

gaaatagttc accatggatc taagcagcat cggcctagta acgagtattt cgatagtggt 1680

gtagatgaca ttaaaaaccc tcaacattac attgtgtggg atatgaatgt gaatagtcac 1740

atctattctg aatttgtagt caccatcaaa ttgccttcta gagtcaaaga ttctcctgcc 1800

accgaagaag attgtcacaa tttatcagag gtttcatcac tgatcttgag ctctggttcg 1860

cctgatagtg tatcacagga catgaacctt caggcttctc cagcattggg aggtcattat 1920

gaagccccca tgttaggaga taaagtggaa agggctccaa gcacgccttg gatgcctttc 1980

tccatgttat ttgcagcgat ttcaaccaaa gtttctgctg agaatatgga catggtcaac 2040

agttgttacg aagaattcaa gagcaagaaa ataagccgag tcgacctagt gaagaagtta 2100

aggcatatag ttggtgacag aatgcttata tcaacaataa tgcgactcca agataagttg 2160

ccccctatgt cgaggcatga agcacccaaa cacgtgggcc aagatgatgg ctaaaccatg 2220

aagtcgtcgg tgcggatggt gagagaagca actggccgac ttgactagtg gagagaagct 2280

tatctcacat aagttgtctg cgtctgtata tgatgcgggt ttgtcttttc ggtcctagga 2340

ttcatgttgt caagatttct gtcatcggtt tccgatgcga aatgatgatt tcagctttac 2400

tcgtggacct gaggttacac ataaaattga taaaagttcc ttgtttaccc tcctgtttct 2460

ttttcttctc tttgcataaa aacatgtaat gttatataca tattgattaa tgttattgtt 2520

tcgccatagt cgtggtgttg tgctttaatg tta 2553

Claims (10)

1. Use of the PHI1 protein or related biological material thereof in any one of:

   p1, regulating and controlling plant photosynthesis;

   p2, regulating and controlling the plant photosynthetic rate;

   p3, regulating and controlling the light respiration rate of the plant;

   p4, regulating and controlling the plant height;

   p5, regulating and controlling the curling degree of the plant leaves;

   p6, regulating and controlling the content of phytoalexin;

   p7, regulating and controlling the drought resistance of the plant;

   the related biological material is a nucleic acid molecule capable of expressing the PHI1 protein or an expression cassette, a recombinant vector, a recombinant bacterium or a transgenic cell line containing the nucleic acid molecule;

   the PHI1 protein is any one of the following proteins:

   (A1) protein with an amino acid sequence of SEQ ID No. 1;

   (A2) the amino acid sequence shown in SEQ ID No.1 is substituted and/or deleted and/or added by one or more amino acid residues and is derived from the protein with the same function of rice;

   (A3) a protein having 99% or more, 95% or more, 90% or more, 85% or more, or 80% or more identity to the amino acid sequence defined in any one of (A1) to (A2) and derived from rice having the same function;

   (A4) a fusion protein obtained by attaching a protein tag to the N-terminus and/or C-terminus of the protein defined in any one of (A1) to (A3).
2. 2. Use according to claim 1, characterized in that: in P1-P6, the modulation is modulation under normal growth conditions or under drought stress conditions;

   under normal growth conditions, the expression level and/or activity of the PHI1 protein or the coding gene thereof in the plant is reduced, the photosynthesis of the plant is enhanced, the photosynthetic rate is increased, the light respiration rate is reduced, the plant height is reduced, the leaf curl is increased and/or the chlorophyll content is increased;

   under drought stress conditions, the expression level and/or activity of the PHI1 protein or the coding gene thereof in the plant is reduced, the photosynthesis of the plant is enhanced, the photosynthetic rate is increased, the light respiration rate is reduced and/or the plant is increased;

   in P7, the regulatory plant drought resistance is embodied as: under drought stress conditions, the expression level and/or activity of the PHI1 protein or the coding gene thereof in the plant is reduced, the photosynthesis of the plant is enhanced, the photosynthetic rate is increased, the light respiration rate is reduced and/or the plant is increased.
3. The use of the PHI1 protein or related biological material thereof in plant breeding;

   the related biological material is a nucleic acid molecule capable of expressing the PHI1 protein or an expression cassette, a recombinant vector, a recombinant bacterium or a transgenic cell line containing the nucleic acid molecule;

   the PHI1 protein is any one of the following proteins:

   (A1) protein with an amino acid sequence of SEQ ID No. 1;

   (A2) the amino acid sequence shown in SEQ ID No.1 is substituted and/or deleted and/or added by one or more amino acid residues and is derived from the protein with the same function of rice;

   (A3) a protein having 99% or more, 95% or more, 90% or more, 85% or more, or 80% or more identity to the amino acid sequence defined in any one of (A1) to (A2) and derived from rice having the same function;

   (A4) a fusion protein obtained by attaching a protein tag to the N-terminus and/or C-terminus of the protein defined in any one of (A1) to (A3).
4. 4. A method for breeding a plant variety having at least one of the traits Q1-Q3, comprising the step of reducing the expression level and/or activity of a PHI1 protein in a recipient plant;

   q1, increased photosynthesis, increased photosynthetic rate, decreased light respiration rate, decreased plant height, increased leaf curliness and/or increased chlorophyll content under normal growth conditions;

   q2, increased photosynthesis, increased photosynthetic rate, decreased light respiration rate and/or increased plant height under drought stress conditions;

   q3, drought resistance is enhanced;

   the PHI1 protein is any one of the following proteins:

   (A1) protein with an amino acid sequence of SEQ ID No. 1;

   (A2) the amino acid sequence shown in SEQ ID No.1 is substituted and/or deleted and/or added by one or more amino acid residues and is derived from the protein with the same function of rice;

   (A3) a protein having 99% or more, 95% or more, 90% or more, 85% or more, or 80% or more identity to the amino acid sequence defined in any one of (A1) to (A2) and derived from rice having the same function;

   (A4) a fusion protein obtained by attaching a protein tag to the N-terminus and/or C-terminus of the protein defined in any one of (A1) to (A3).
5. 5. A method of breeding a transgenic plant having at least any one of the traits Q1-Q3 comprising the steps of: inhibiting and expressing a nucleic acid molecule capable of expressing PHI1 protein in a receptor plant to obtain a transgenic plant; the transgenic plant has at least any one of the traits set forth in Q1-Q3 below as compared to the recipient plant;

   q1, increased photosynthesis, increased photosynthetic rate, decreased light respiration rate, decreased plant height, increased leaf curliness and/or increased chlorophyll content under normal growth conditions;

   q2, increased photosynthesis, increased photosynthetic rate, decreased light respiration rate and/or increased plant height under drought stress conditions;

   q3, drought resistance is enhanced;

   the PHI1 protein is any one of the following proteins:

   (A1) protein with an amino acid sequence of SEQ ID No. 1;

   (A2) the amino acid sequence shown in SEQ ID No.1 is substituted and/or deleted and/or added by one or more amino acid residues and is derived from the protein with the same function of rice;

   (A3) a protein having 99% or more, 95% or more, 90% or more, 85% or more, or 80% or more identity to the amino acid sequence defined in any one of (A1) to (A2) and derived from rice having the same function;

   (A4) a fusion protein obtained by attaching a protein tag to the N-terminus and/or C-terminus of the protein defined in any one of (A1) to (A3).
6. 6. The method of claim 5, wherein: the inhibition of expression of the nucleic acid molecule capable of expressing the PHI1 protein in the recipient plant is achieved by RNA interference techniques.
7. 7. The method of claim 6, wherein: the RNA interference vector introduced into the receptor plant is a recombinant expression vector containing an interference fragment.

   The interference fragment comprises a section A, a section B and a section C from the 5 'end to the 3' end in sequence; the nucleotide sequence of the segment A is 238 th and 532 nd position of SEQ ID No. 2; the segment C is reverse complementary to the segment A; the segment B is not complementary to both the segment a and the segment C.
8. 8. Use or method according to any of claims 1-7, wherein: the nucleic acid molecule capable of expressing the PHI1 protein is any one of the following DNA molecules:

   (B1) a DNA molecule shown in SEQ ID No.2 or SEQ ID No.3 at position 366-2281;

   (B2) a DNA molecule which hybridizes with the DNA molecule defined in (B1) under stringent conditions and encodes the PHI1 protein;

   (B3) a DNA molecule which has more than 99%, more than 95%, more than 90%, more than 85% or more than 80% of identity with the DNA sequence limited by (B1) or (B2) and encodes the PHI1 protein.
9. 9. Any of the following applications:

   use of R1, the interfering fragment of claim 7 or the interfering vector for growing plants having at least any one of the following traits: under normal growth conditions, photosynthesis is enhanced, photosynthetic rate is increased, light respiration rate is reduced, plant height is reduced, leaf curling degree is increased and/or chlorophyll content is increased; under drought stress conditions, photosynthesis is enhanced, photosynthetic rate is increased, photorespiration rate is reduced and/or plants are increased; drought resistance is enhanced;

   use of R2, the interfering fragment of claim 7, or the interfering vector in plant breeding;

   use of R3 or the method of any one of claims 4 to 8 in plant breeding.
10. 10. Use or method according to any of claims 1-9, wherein: the plant is a monocotyledon or a dicotyledon;

    further, the monocotyledonous plant is a plant of the order gramineae;

    further, the plant of the order gramineae is a gramineae;

    more specifically, the gramineous plant is a plant of the genus oryza;

    preferably, the plant of the genus oryza is rice.

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