Tea tree CsBZR1 gene and cold-resistant application thereof

Abstract

The invention discloses a tea tree CsBZR1 gene and cold resistance application thereof, the tea tree CsBZR1 gene is a core gene in a tea tree brassinolide signal transduction pathway, and the nucleotide sequence of the tea tree CsBZR1 gene is shown as SEQ ID NO.1 in a sequence table. According to the application of the CsBZR1 gene of the tea tree to regulation and control of cold resistance of the tea tree, the low-temperature tolerance of the tea tree can be reduced by inhibiting expression of the CsBZR1 gene in vivo, and the cold resistance of arabidopsis thaliana is improved by overexpression of the CsBZR1 gene. Expression of CsBZR1 of a tea tree under a low-temperature condition is directly induced to be up-regulated by low-temperature stress of the tea tree, cold resistance of arabidopsis thaliana is improved by overexpression of the gene in the arabidopsis thaliana, and low-temperature damage of the tea tree can be remarkably aggravated by inhibiting expression of CsBZR1 in the tea tree through an antisense oligonucleotide technology. The cloning of the gene not only is beneficial to determining the effects of brassinolide and the signal transduction core factor BZR1 thereof in the cold-resistant process of the tea tree, but also is beneficial to promoting the genetic improvement process aiming at enhancing the cold resistance of the tea tree.

Description

Technical Field

[0001] The invention relates to the technical field of genetic engineering, and in particular to a tea tree CsBZR1 gene and its cold-resistant application. Background Art

[0002] Tea tree (Camellia sinensis (L.) O. Kuntze) is an important leaf-type economic crop in my country, playing a very important role in various aspects such as economy and culture. However, as a perennial evergreen woody plant that likes warmth and fears cold, the growth, development, tea quality and yield of tea trees are significantly affected by low temperature stress, which directly restricts the healthy and sustainable development of the tea industry. In recent years, the global climate has been changeable, and low temperature extreme weather has frequently occurred in tea gardens. Tea trees frequently encounter low temperature damage such as "frost" or "late spring cold" during the wintering period and early spring. Low temperature has become a key environmental factor restricting the growth of tea trees and the high-quality development of the tea industry. In-depth exploration of the cold resistance mechanism of tea trees and the discovery of effective cold resistance regulatory genes have important scientific significance and application value for promoting tea tree cold resistance breeding through molecular design breeding, improving the cold resistance of tea trees, ensuring the safe production of tea, and promoting the sustainable development of the tea industry.

[0003] Brassinosteroids (BRs) have been shown to play an important role in plant growth and development and stress resistance. BZR1, as the core gene in the brassinosteroid signal transduction pathway, has a typical helix-loop-helix protein structure, and shows high conservation and homology in the amino acid sequence. It can specifically bind to the BRRE (CGTGT / CG) and E-box (CANNTG) cis-acting elements in the promoter region of downstream target genes, thereby precisely regulating the expression of target genes and widely participating in various physiological processes and environmental response processes of plants. BZR1 has also been shown to play an important regulatory role in the process of plant response to low temperature stress. In tea plants, previous studies have also found that BZR1, a key signal transduction gene in the brassinosteroid pathway, is significantly induced to upregulate expression under low temperature conditions, suggesting that BZR1 may actively participate in the response of tea plants to low temperature stress. However, the cold resistance of brassinosteroids to tea plants, especially the role of BZR1, a key signal transduction gene in its pathway, in the cold resistance of tea plants is still unclear. Therefore, studying the biological function of BZR1, a key gene in the brassinolide signal transduction pathway, in the tea tree's response to low temperature stress will help to gain a deeper understanding of the molecular regulatory mechanism of the tea tree's response to low temperature stress, provide a solid theoretical basis and key gene resources for the use of molecular design breeding to cultivate new cold-resistant tea varieties, and provide new ideas for the study of tea tree cold resistance mechanisms. Summary of the invention

[0004] The purpose of the present invention is to provide a tea tree CsBZR1 gene and its cold-resistant application, to provide a new idea for the study of the cold-resistant mechanism of tea trees, and to provide a theoretical and application basis for promoting cold-resistant breeding of tea trees by molecular design breeding.

[0005] The present invention provides a tea tree CsBZR1 gene. According to an embodiment of the present invention, the tea tree CsBZR1 gene is a tea tree brassinolide signal transduction core gene, and the nucleotide sequence of the tea tree CsBZR1 gene is shown in the sequence table SEQ ID NO.1.

[0006] In another aspect of the present invention, the present invention provides an application of the tea plant CsBZR1 gene to regulate the cold resistance of tea plants. According to an embodiment of the present invention, the CsBZR1 gene is inhibited from being expressed in plants to reduce the low temperature tolerance of tea plants, while the overexpression of CsBZR1 is used to improve the cold resistance of Arabidopsis thaliana.

[0007] Compared with the prior art, the present invention has the following beneficial effects:

[0008] In the present invention, the brassinolide signal transduction core gene BZR1 gene (i.e. CsBZR1) that regulates the response of tea trees to low temperature stress is cloned and verified for the first time. The gene plays an important regulatory role in the process of tea trees responding to low temperature stress, and directly affects the formation of cold resistance of tea trees. The present invention also provides a recombinant plasmid containing the BZR1 gene, a transgenic Arabidopsis thaliana, etc. The present invention enriches the functional research of brassinolide signal transduction genes in tea trees, provides a new idea for the cold resistance mechanism of tea trees, and provides a theoretical and application basis for promoting cold resistance breeding of tea trees using molecular breeding. BRIEF DESCRIPTION OF THE DRAWINGS

[0009] Figure 1 The gene sequence information of tea plant CsBZR1 in Examples 1 and 2 of the present invention and its expression pattern diagram in different tissues and under 4°C low temperature treatment, wherein Figure 1 A is the basic sequence information of the gene, Figure 1 B is the expression level of the gene in different tea plant tissues, Figure 1 C is the expression of the gene under low temperature stress;

[0010] Figure 2 The figure shows the effect of silencing the gene CsBZR1 in tea plants on the cold resistance of tea seedlings in Example 3 of the present invention, wherein A, B, C, D, and E are respectively the expression level change graph of plants inhibited by antisense oligonucleotide CsBZR1 in Example 3 of the present invention, the chlorophyll fluorescence graph of damaged tea seedling leaves, the Fv / Fm value change graph of tea seedlings after CsBZR1 expression inhibition, the superoxide dismutase content change graph, and the malondialdehyde content analysis graph;

[0011] Figure 3This is the effect of overexpression of the gene CsBZR1 in vitro on the cold resistance of Arabidopsis in Example 4 of the present invention, wherein A, B, C, and D are respectively expression analysis graphs of CsBZR1-overexpressing Arabidopsis strains, cold resistance phenotype analysis graphs of wild-type Arabidopsis and CsBZR1-overexpressing Arabidopsis under low temperature treatment, and survival rate and relative conductivity change graphs of Arabidopsis overexpressing strains under low temperature treatment. DETAILED DESCRIPTION

[0012] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0013] The terms used in the present invention, unless otherwise specified, generally have the meanings commonly understood by those of ordinary skill in the art.

[0014] In the following examples, various processes and methods not described in detail are conventional methods known in the art. The primers used are indicated when they first appear, and the same primers used thereafter are the same as those indicated for the first time.

[0015] Example 1

[0016] The tea tree CsBZR1 gene is a brassinolide signal transduction gene, and its cloning and sequence structure analysis are as follows:

[0017] (1) Plant materials and RNA extraction

[0018] The national-level improved tea variety "Shucha Zao" was used for sequence cloning. This variety was planted in the National High-tech Agricultural Park of Anhui Agricultural University, Luyang District, Hefei City, Anhui Province (31°36′N, 117°12′E), and young leaves were used for RNA extraction. Total RNA was extracted according to the operating instructions of the FastPure Universal Plant Total RNA Isolation Kit (Vazyme, Nanjing, China), and the RNA content and quality were tested and used for subsequent analysis.

[0019] (2) Target gene cloning

[0020] Based on the reference genome of tea tree "Shucha Zao", the reference sequence of CsBZR1 gene was obtained, PCR primers were designed using Primer 5, and PCR reaction was carried out using cDNA obtained by reverse transcription as template. The 50μl PCR reaction system was: ddH2O 17μl, 2×Phanta Max Buffer 25μl, dNTP Mix 1μl, 2μl of upstream and downstream primers, Phanta Max Super-FidelityDNA Polymerase 1μl, and 2μl of template. The PCR reaction program was: 95℃3min, 95℃15s, 68℃15s, 72℃1min, and 35 cycles. After the PCR product was purified and recovered, it was connected to the cloning vector PGEX4T-1 and transformed into Escherichia coli DH5α for sequencing identification. Among them, the upstream primer used for gene cloning was 5'-ATGACTGCCGGCGGATCA-3', and the downstream primer was 5'-TTAAGTGTGGGCTTTTCCA-3'.

[0021] The nucleotide sequence of the CsBZR1 gene finally obtained is shown in the sequence table SEQ ID NO.1, which is as follows:

[0022] ATGACTGCCGGCGGATCATCAGGGCGTTTACCGACCTGGAAGGAGAGAGAGAACAACAAGAGGAGAGAGAGGAGGAGAAGAGCTATTGCTGCCAAGATATTTACAGGTCTCAGAGCTCAGGGAAACTACAAGCTTCCCAAACACTGCGACAACAACGAAGTCTTGAAAGCTCTCTGTTCCGAAGCTGGTTGGGTCGTCGAAGACGATGGCACCACTTATCGCAAGGGTTGCAGACCTCCCCCAAATCAAACTGAGATTGCAGGCAATACAACAAACATAAGTAACATCAGTGCATGTTCATCGATCCAACCGAGCCCAAATTCCTCATCCTTTCCAAGTCCTGTACCGTCTTACCACGCCAGCCCAACATCCTCTTCCTTCCCAAGCCCCACACGTTTTGACGGAAACCCCTCATCTTACATTCTGCCTTTCCTCCGAAACTTAGCCTCCATTCCTTCGTCTCTCCCTCCCCTCCGCATATCCAGCAGTGCCCCTGTAACTCCGCCTCTTTCTTCCCCAACTTCCCGAGGTTCAAAGCGAAAGGCCGATTGGGAATCATTTTCCAATGGCTCATTGAACACATTCCGTCACCCCCTTTTCGCTGCCTCTGCCCCAGCAAGTCCCACTCGCCGCCACCATCTTACACCGGCCACAATTCCAGAATGTGATGAGTCCGATGCCTCCAGTGTTGATTCGGGTCAGTGGGTCAGTTTCCAGACAGGGGCAGCTCCGGCTGCTCCATCGTCCCCAACATTTAACCTTGTAAAAGCTGTTGCTCAGCAGAGCTCCCTCCAAAATGGAACTGATGGGCATGAAGAATTGGGCTGGGGAGTAGCAGCACAAAGAGGGCGTGGCTCTGAATTTGAGTTTGAGAGTGGCACAGTGAAAGCGTGGGAGGGTGAGATAATTCATGATATTGGAGTGGACGATTTGGAGCTCACGCTTGGGAGTGGAAAAGCCCACACTTAA。

[0023] The protein sequence encoded by the CsBZR1 gene is shown in SEQ ID NO.2, and is as follows:

[0024] MTAGGSSGRLPTWKERENNKRRERRRRAIAAKIFTGLRAQGNYKLPKHCDNNEVLKALCSEAGWVVEDDGTTYRKGCRPPPNQTEIAGNTTNISNACSSSSIQPSPNSSSFPSPVPSYHASPTSSSFPSPTRFDGNPSSYILPFLRNLASIPSSLPPLRIS SSAPVTPPLSSPTSRGSKRKADWESFSNGSLNTFRHPLFAASAPASPTRRHHLTPATIPECDESDASSVDSGQWVSFQTGAAPAAPSSPTFNLVKAVAQQSSLQNGTDGHEELGWGVAAQRGRGSEFEFESGTVKAWEGEIIHDIGVDDLELTLGSGKAHT.

[0025] Example 2

[0026] Analysis of tissue and low temperature stress expression patterns of CsBZR1 gene:

[0027] (1) CsBZR1 gene expression in different tissues of tea plants

[0028] The expression pattern of CsBZR1 gene was analyzed in 8 representative tissues and organs of Shucha Zao variety. These 8 tissues and organs are tender shoots, flowers, fruits, tender leaves, mature leaves, old leaves, roots and stems. After extracting total RNA from these samples, quantitative PCR was used to analyze the expression of CsBZR1. The reverse transcription product was diluted 30 times as a template and the expression of CsBZR1 was analyzed using Hieff TM qPCR Green Master Mix (No Rox) (Yeasen, Shanghai, China), prepare 20 μl reaction system: 2.0 μl reverse transcription product diluted 30 times, 0.4 μl upstream and downstream primers (10 pmol / μl), 10 μl Hieff TM qPCR Green Master Mix, 7.2 μl ddH20, 3 replicates for each reaction. Then, run 39 cycles on the Bio-rad CFX-96 instrument with the following program: ① 95°C 5 min; ② 95°C 10 sec, 60°C 30 sec, 72°C 30 sec; ③ From 65°C to 95°C, draw a melting curve at 0.1°C / sec. The upstream primer is 5'-CACGTTTTGACGGAAACCCC-3', and the downstream primer is 5'-CTGGAGGCATCGGACTCATC-3'. The tea tree ACTIN gene was used as the internal reference, the upstream primer was 5'-GCCATCTTTGATTGGAATGG-3'), and the downstream primer was 5'-GGTGCCACAACCTTGATCTT-3'. The relative expression levels of CsBZR1 in different tissues were calculated using the instrument's own analysis software (2 -deltaCT ).

[0029] (2) Analysis of the expression pattern of CsBZR1 under low temperature stress

[0030] The "Shucha Zao" material used for low temperature treatment was collected from the asexual tea seedling breeding base of Chaohu Niansheng Agricultural Ecology Co., Ltd. (31°37′N, 117°55′E). Select tea seedlings with consistent growth, healthy and free of pests and diseases and place them in a low temperature incubator for 4°C low temperature treatment, with a light intensity of 9000-10000 lux, a light duration of 16h, and a humidity of 75%. At the corresponding treatment time (0h, 4h, 8h, 12h, 24h and 48h), the first or second tender leaves of the control group and the treated plants were collected, three biological replicates were set up, and they were frozen with liquid nitrogen and stored in a -80°C refrigerator for analysis of CsBZR1 gene expression. RNA extraction and quantitative PCR methods are the same as above.

[0031] Figure 1 A is the sequence information of CsBZR1 cloned from tea plants. It can be seen that the length of CsBZR1 mRNA is 969 bp, the length of the protein sequence is 322 amino acids, and the molecular weight is approximately 34.4 kDa. Figure 1 B shows the expression changes of CsBZR1 in different tissues of tea plants under low temperature treatment of 4°C. It was found that CsBZR1 was constitutively expressed in different tissues as a whole, and tended to be highly expressed in tender buds, flowers and young leaves. Figure 1 C is the expression pattern of CsBZR1 under low temperature stress. The results show that CsBZR1 is significantly induced to upregulate its expression by low temperature, suggesting that CsBZR1 may play an important role in the response of tea plants to low temperature stress.

[0032] Example 3

[0033] Effects of inhibiting CsBZR1 expression in tea plants on their cold resistance:

[0034] (1) Inhibition of CsBZR1 expression in tea plants

[0035] Antisense oligonucleotide primers (two pairs in total) were designed and synthesized according to the CsBZR1 sequence. The primer design was completed through the website http: / / sfold.wadsworth.org / cgi-bin / soligo.pl. The primer sequences are as follows:

[0036] P1: 5'-TTTGGGGGAGGTCTGCAACC-3';

[0037] P2: 5'-TTTCCGTCAAAACTGTGGG-3';

[0038] P3: 5'-TGTGGCCGGTAAGATGGT-3';

[0039] P4: 5'-TTTGGAGGGAGCTCTGCTGA-3';

[0040] Dissolve in sterile water to prepare in vitro oligonucleotide antisense inhibitory primers, and the blank is sterile water; select healthy, pest-free and disease-free one-bud and two-leaf tea leaves, insert them into a 1.5ml centrifuge tube containing 1ml 100μM primer solution, and ensure that the end of the leaf is completely immersed in the solution. Place the centrifuge tube in a 25℃ light incubator for light culture at 16h / darkness 8h. Samples were taken from the experimental group and the control group after treatment for 0h, 6h, 12h, 24h, and 48h, and one bud and two leaves were treated with liquid nitrogen and stored in a -80℃ refrigerator. Total RNA was extracted from the treated and control samples, and then reverse transcribed, and the expression of the CsBZR1 gene was analyzed using quantitative PCR. The gene expression level of CsBZR1 in the control and treated samples was detected, and the results showed that oligonucleotide antisense inhibition could significantly interfere with the expression level of CsBZR1, and the inhibition effect was best at 6h.

[0041] (2) Effects of oligonucleotide antisense inhibition on CsBZR1 gene expression and physiological parameters under low temperature stress in tea plants

[0042] In order to study the role of CsBZR1 in low temperature stress of tea plants, the samples treated with oligonucleotide antisense inhibitory primer solution for 6 hours were placed at -2℃ for 1 hour, and then restored to room temperature for 30 minutes to observe the changes in chlorophyll fluorescence and Fv / Fm values ​​as well as changes in physiological indicators. The malondialdehyde content after low temperature treatment was determined according to the malondialdehyde (MDA) content detection kit (Cat. No.: BC0020) (Solarbio, Beijing, China). The absorbance of each sample at 450nm, 532nm and 600nm was measured, and ΔA450=A450 was calculated respectively. 测定 -A450 空白 , ΔA532=A532 测定 -A532 空白 , ΔA600=A600 测定 -A600 空白 .

[0043] MDA content (nmol / g) = 5*[6.45*(ΔA532-ΔA600)-1.29*ΔA450] / W.

[0044] Superoxide dismutase activity was determined using a superoxide dismutase (SOD) activity assay kit (Solarbio, Beijing, China). Prepare the SOD assay working solution, mix thoroughly, and incubate in a 37°C water bath for 30 min. The absorbance value A of each tube was measured at 560 nm. Calculate ΔA 测定 =A 测定 -A 对照 , ΔA 空白 =A1 空白 -A2 空白 If there is precipitation at the bottom, mix it and then measure it. (Only 1 to 2 blank tubes 1 and 2 need to be made; each sample has a control tube.) Inhibition percentage = (ΔA 空白 -ΔA 测定 )÷ΔA 空白 ×100%. According to the sample mass:

[0045] SOD activity (U / g mass) = [inhibition percentage ÷ (1-inhibition percentage) × V 反总 ]÷(W×V 样 ÷V 样总 )×F=11.11×inhibition percentage ÷(1-inhibition percentage) ÷ W×F.

[0046] like Figure 2 As shown in A, compared with the control group, the expression level of CsBZR1 was significantly reduced after 6 hours of inhibition, and the inhibition rate of expression reached about 50%. Figure 2As shown in B, the expression inhibition samples were further subjected to low temperature treatment (1°C for 1 hour) and the low temperature damage of the inhibition samples was detected. It was found that the CsBZR1 expression inhibition tea seedlings were more seriously damaged than the control group under low temperature conditions. Figure 2 As shown in Figure CE, by measuring physiological indicators such as maximum photosynthesis rate (Fv / Fm), SOD activity, and malondialdehyde content (MDA), it was found that the Fv / Fm and SOD activity of the CsBZR1 expression inhibition sample decreased significantly, while the malondialdehyde content increased significantly, indicating that the expression inhibition of CsBZR1 reduced the tolerance of tea seedlings to low temperature stress. The above research results show that CsBZR1 plays a positive regulatory role in tea plants' response to low temperature stress.

[0047] Example 4

[0048] Effects of CsBZR1 overexpression in vitro on low temperature tolerance in Arabidopsis thaliana:

[0049] (1) Construction of CsBZR1-PCAMBIA1305 vector

[0050] Using the PGEX4T-1::CsBZR1 plasmid as a template, specific primers were designed for PCR amplification, followed by purification and elution. The PCAMBIA1305 vector was double-digested and linearized, with Spe1 and BamH1 as restriction sites. The digestion product was confirmed to have been cut by 1% agarose gel electrophoresis, and then purified and eluted. The linearized PCAMBIA1305 vector and the purified PCR product were recombined using a recombinase (GenRec Assembly MasterMix Kit, GENERAL BIOL), and then transformed into Escherichia coli competent cells DH5α (WEIDI, Shanghai, China). After the positive clones were grown and verified by colony PCR, the correct single clones were picked for sequencing to obtain the CsBZR1-PCAMBIA1305 vector.

[0051] (2) CsBZR1 transformation into Arabidopsis

[0052] Take an appropriate amount of wild-type Arabidopsis seeds and add deionized water. Sow them after 72 hours of vernalization treatment, place them under appropriate conditions (humidity 60%; temperature 23°C; photoperiod 16h light / 8h dark) for germination and transplanting. The target gene is cloned into the PCAMBIA1305 vector, and then transformed into Escherichia coli DH5α for sequence identification; then the plasmid is extracted and further transformed into Agrobacterium GV3101, and verified by colony PCR; then the GV3101 Agrobacterium containing the target gene is amplified and cultured to at least 200 ml, and cultured at 28°C and 200 rpm for 16 to 20 hours. Then the bacteria are collected by centrifugation, and the bacteria are resuspended in 5% sucrose solution to an OD600 of about 0.8, and then 0.1% silwet L-77 is added for mixing and prepared for infection.

[0053] Select Arabidopsis seedlings about 4 weeks old for infection. Place the prepared transformation solution in a container, completely soak the inflorescence of Arabidopsis for 1 minute, then place it in the dark for 24 hours and continue to culture under normal conditions to harvest seeds. The obtained seeds are sown on a 1 / 2MS solid culture medium containing hygromycin b to screen positive transgenic strains. When the true leaves of the screened positive plants grow and are green in color, they are transferred to the soil and continue to grow for 2 to 3 weeks. Subsequently, the genome of the Arabidopsis plant leaves is extracted, and the successful integration of the target gene is identified using specific primers. Arabidopsis strains identified as positive are further planted and cultivated. Arabidopsis seedling RNA is extracted, and PCR is performed using gene-specific primers to detect the expression of the target gene.

[0054] (3) Detection of low temperature tolerance in overexpressed Arabidopsis

[0055] The transgenic plants were cultured at -8°C for 2.5 h, then the culture dishes were removed and subsequently transferred to a normal culture room for culture. The survival of the seedlings was observed after 3 days, and the survival rate and relative conductivity of Arabidopsis were also calculated.

[0056] Figure 3 The expression and low temperature phenotype analysis of CsBZR1 in Arabidopsis thaliana overexpression lines and wild type. Figure 3 As shown in A, compared with wild-type Arabidopsis, the expression level of CsBZR1 in overexpression lines OE-15 and OE-18 was significantly increased, indicating that CsBZR1 has been successfully overexpressed in Arabidopsis. Phenotypic analysis of Arabidopsis under low temperature stress found that under normal temperature (25°C), there was no significant phenotype difference between overexpressed CsBZR1 and wild-type Arabidopsis. However, when subjected to -6°C, 3h low temperature treatment and restored growth for 3 days, it was found that the survival rate of Arabidopsis plants overexpressing CsBZR1 was significantly higher than that of wild-type Arabidopsis (such as Figure 3The survival rate of Arabidopsis thaliana overexpressing CsBZR1 was found to be over 75%, while the survival rate of wild-type plants was less than 50% (as shown in Figure 2A). Figure 3 C). Figure 3 As shown in D, by detecting the ion permeability of overexpressed CsBZR1 and wild-type Arabidopsis under low temperature stress, it was found that the ion permeability of the overexpressed plants was significantly lower than that of the wild-type plants, indicating that overexpression of the CsBZR1 gene significantly improved the tolerance of Arabidopsis to low temperature stress.

[0057] In summary, CsBZR1 was significantly induced to up-regulate expression by low temperature. Overexpression of CsBZR1 in Arabidopsis significantly reduced the ion permeability of Arabidopsis under low temperature conditions and increased the survival rate of Arabidopsis. In tea plants, the inhibition of CsBZR1 expression by antisense oligonucleotides significantly reduced the maximum photosynthesis rate of tea seedlings under low temperature conditions, increased the content of malondialdehyde and superoxide dismutase activity, and reduced the low temperature tolerance of tea seedlings, indicating that CsBZR1 plays an important role in the low temperature response of tea plants. The cloning and functional verification of this gene will not only be beneficial to the study of the role of brassinolide and its signal transduction genes in the cold resistance of tea plants, but also help promote the genetic improvement process aimed at enhancing the cold resistance of tea plants and promote the sustainable development of the tea industry, which has important theoretical and application value.

[0058] In the present invention, the brassinolide signal transduction gene CsBZR1 that regulates the response of tea trees to low temperature stress was cloned and verified for the first time, and it was found that the gene plays an important regulatory role in the process of tea trees responding to low temperature stress. The present invention also provides a recombinant plasmid and a transgenic engineering bacterium containing the CsBZR1 gene. The present invention not only enriches the understanding of tea tree cold resistance, provides new ideas for the study of tea trees and even plants cold resistance mechanism, but also provides an important target gene for tea tree cold resistance breeding, and provides a theoretical and practical reference basis for promoting tea tree cold resistance breeding using molecular design breeding.

[0059] The above contents are merely examples and explanations of the structure of the present invention. The technicians in this technical field may make various modifications or additions to the specific embodiments described or replace them in a similar manner. As long as they do not deviate from the structure of the present invention or exceed the scope defined by the claims, they should all fall within the protection scope of the present invention.

Claims

1. A tea plant CsBZR1 gene, characterized in that: The nucleotide sequence of the tea tree CsBZR1 gene is shown in the sequence table SEQ ID NO.

1.

2. Application of tea tree CsBZR1 gene to regulate cold resistance of tea tree, characterized in that: The CsBZR1 gene described in claim 1 is used to reduce the low temperature tolerance of tea plants by inhibiting its expression in vivo, and is used to improve the cold resistance of Arabidopsis thaliana by overexpressing it in vitro.

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