Application of AtCRK10 gene in improving cadmium tolerance of arabidopsis thaliana

Abstract

The invention discloses an application of an AtCRK10 gene in improvement of cadmium tolerance of arabidopsis thaliana. Researches show that the expression of the AtCRK10 gene under cadmium stress is remarkably up-regulated, and the expression is remarkably up-regulated along with the increase of cadmium treatment time. Meanwhile, the AtCRK10 gene is overexpressed in arabidopsis thaliana, so that the symptom of the arabidopsis thaliana under cadmium toxicity can be relieved, the fresh weight, the fresh weight chlorophyll, the root length and the lateral root number of the arabidopsis thaliana can be remarkably increased under cadmium stress, the cadmium toxicity inhibition effect of the plant is relieved, and the growth of the plant is remarkably promoted. The invention finds that the AtCRK10 gene has obvious influence on the growth of arabidopsis thaliana and the effect of relieving cadmium toxicity inhibition, can enhance the cadmium tolerance of plants under the condition of cadmium stress, can well regulate the growth and development of arabidopsis thaliana under the condition of cadmium stress, has important significance on the biological function of adapting to cadmium stress regulation of arabidopsis thaliana, and has broad application prospects. The method is of great significance in improving the growth condition of crops in the cadmium-polluted area and increasing the grain yield.

Description

Technical Field

[0001] The present invention belongs to the technical field of molecular breeding, and more specifically, relates to the application of the AtCRK10 gene in improving the cadmium tolerance of Arabidopsis thaliana. Background Art

[0002] Plant roots absorb cadmium (Cd) from the soil and enter the human body through the food chain. There are many sources of cadmium in the soil, including atmospheric deposition, industrial wastewater and domestic sewage discharge, industrial solid waste and urban garbage dumping, and chemical pesticide application, which can lead to the aggravation of heavy metal pollution in the soil.

[0003] Cd is a highly toxic heavy metal. Excessive Cd will affect the growth and physiological activities of plants: high concentrations of Cd inhibit seed germination, affect the growth of radicles and plumules, and also inhibit the activity of protochlorophyllide synthase required for plant synthesis of chlorophyll, causing the chlorophyll content and chlorophyll a / b ratio in leaves to decrease. At the same time, the efficiency of PSⅠ and PSⅡ also decreases with the increase of Cd concentration; Cd stress will reduce the activity of nitrate reductase in plants, resulting in NO in leaves. 3- Accumulation of Cd has toxic effects on cells; excessive absorption of Cd by plant roots can lead to root browning, reduction of root length and dry weight, changes in root morphology, decreased root absorption capacity, cell damage, and ultimately root death; Cd can increase the permeability of cell membranes, causing overall damage to the plant cell membrane system; Cd can affect the activity of superoxide dismutase, catalase and peroxidase in plants, as well as enzymes related to nitrogen metabolism, thereby indirectly affecting plant photosynthesis, respiration, metabolic activities, etc., and further affecting the normal growth and development of plants, and ultimately affecting plant yield and quality. Therefore, it is of certain significance to understand the functions of genes related to Arabidopsis thaliana's response to Cd toxicity.

[0004] Cysteine-rich receptor-like kinases (CRKs) belong to a large family of DUF26-containing receptor-like kinases (RLKs). 44 CRKs in Arabidopsis and 1074 CRKs in 14 crop species have been discovered through multi-omics and molecular genetics analyses, but only 63 CRKs have been shown to play a role in regulating plant immunity, abiotic stress responses (such as salinity, osmotic, oxidative, and thermal) and growth and development. CRKs control abiotic stress responses and stress-growth balance, and CRKs are involved in regulating Ca 2+Influx of ROS, homeostasis of ROS, activation of MAPK cascade and deposition of hydrazide, thus regulating stomatal closure, pathogenesis-related (PR) gene expression and PCD. Studies have shown that CRK4 regulates stomatal movement and can also regulate ABI1 protein expression in response to drought stress by regulating the protein stability of PUB13; CRK5 positively regulates salt stress; CRK45 regulates the response of Arabidopsis to abiotic stress by regulating ABA biosynthesis; hydrazide is a key regulator of intercellular membrane transport in response to osmotic and salinity stress, and CRK2 can enhance the tolerance of plants to salt stress during the germination stage by promoting the deposition of hydrazide in the intercellular membrane; under abiotic stress treatments such as low temperature, H2O2, and NaCl, the induced expression of CRK10 gene is obvious, and CRK10 plays an important role in coordinating the immune response of plants under different abiotic stress conditions. However, the functional role of CRK genes in cadmium stress is still unclear, and the response to cadmium stress has not been studied. Therefore, exploring the function and role of the Arabidopsis CRK gene in plant Cd tolerance under Cd stress conditions can lay a better foundation for breeding more Cd stress-resistant crop varieties in agricultural production to meet the needs of agricultural production under stress environments. Summary of the invention

[0005] The technical problem to be solved by the present invention is to overcome the unclear functional role of the existing CRK gene in cadmium stress and the lack of research on the response to cadmium stress, and provide the application of the AtCRK10 gene in improving the cadmium tolerance of Arabidopsis thaliana.

[0006] The purpose of the present invention is to provide application of AtCRK10 gene in alleviating cadmium stress in Arabidopsis thaliana.

[0007] Another object of the present invention is to provide the use of a preparation for promoting the expression of the AtCRK10 gene.

[0008] Another object of the present invention is to provide a method for alleviating cadmium stress in Arabidopsis thaliana or promoting the growth of Arabidopsis thaliana under cadmium stress.

[0009] Another object of the present invention is to provide a method for cultivating cadmium-resistant plants.

[0010] The above-mentioned purpose of the present invention is achieved through the following technical solutions:

[0011] The present invention clarifies the expression of Arabidopsis thaliana AtCRK10 gene (its nucleotide sequence is shown in SEQ ID NO:1, the encoded protein sequence is shown in SEQ ID NO:2, and the gene number is AT4G23180) under cadmium stress. The AtCRK10 gene is significantly upregulated under cadmium stress, and is significantly upregulated with the increase of cadmium treatment time. At the same time, overexpression of the AtCRK10 gene in Arabidopsis thaliana can alleviate the symptoms of Arabidopsis thaliana under cadmium poisoning, and significantly increase the fresh weight, fresh weight chlorophyll, root length and lateral root number of Arabidopsis thaliana under cadmium stress, relieve the inhibitory effect of cadmium poisoning on plants, and significantly promote the growth of plants. And after the AtCRK10 gene is mutated, the opposite result is presented, and the crk10 mutant material is more sensitive to cadmium poisoning. The results showed that the AtCRK10 gene had a significant effect on the growth of Arabidopsis and the elimination of the inhibitory effect of cadmium toxicity. The AtCRK10 gene enhanced the cadmium tolerance of plants under cadmium stress conditions, which was of great significance to the biological function of Arabidopsis in adapting to cadmium stress regulation. It could well regulate the growth and development of Arabidopsis under cadmium stress, and was of great value in improving the growth of crops in cadmium-contaminated areas and increasing grain yield, and was worthy of promotion and application.

[0012] Therefore, the present invention provides the use of the AtCRK10 gene in alleviating cadmium stress in Arabidopsis thaliana.

[0013] The present invention provides application of the AtCRK10 gene in preparing a preparation for alleviating cadmium stress in Arabidopsis thaliana.

[0014] The present invention provides application of the AtCRK10 gene in improving the tolerance of Arabidopsis thaliana to cadmium stress.

[0015] The present invention provides application of AtCRK10 gene in promoting growth of Arabidopsis thaliana under cadmium stress.

[0016] The present invention provides application of the AtCRK10 gene in cultivating Arabidopsis plants resistant to cadmium stress.

[0017] The present invention provides use of a preparation for promoting AtCRK10 gene expression in alleviating plant cadmium stress or in preparing a product for alleviating plant cadmium stress.

[0018] The invention provides use of a preparation for promoting AtCRK10 gene expression in promoting plant growth under cadmium stress, or in preparing a product for promoting plant growth under cadmium stress.

[0019] The invention provides application of a preparation for promoting AtCRK10 gene expression in cultivating plants resistant to cadmium stress.

[0020] Furthermore, the plant is Arabidopsis thaliana.

[0021] Preferably, the agent for promoting the expression of the AtCRK10 gene is a plasmid, a vector, or a recombinant bacterium that overexpresses the AtCRK10 gene, which is constructed by conventional methods in the art.

[0022] More preferably, the preparation is an AtCRK10 gene overexpression vector.

[0023] The invention provides a method for alleviating cadmium stress of Arabidopsis thaliana or promoting the growth of Arabidopsis thaliana under cadmium stress, and the preparation containing overexpressed AtCRK10 gene is used to treat the plant.

[0024] The present invention also provides a method for cultivating cadmium-resistant plants, which comprises over-expressing the AtCRK10 gene in the plant, or introducing a recombinant vector containing the over-expressed AtCRK10 gene into the plant.

[0025] The present invention has the following beneficial effects:

[0026] The invention clarifies the function and effect of Arabidopsis CRK gene under cadmium stress. Studies show that Arabidopsis AtCRK10 gene is significantly upregulated under cadmium stress and significantly upregulated with the increase of cadmium treatment time. Overexpression of AtCRK10 gene in Arabidopsis can alleviate the symptoms of Arabidopsis under cadmium poisoning, and significantly increase the fresh weight, fresh weight chlorophyll, root length and lateral root number of Arabidopsis under cadmium stress, relieve the cadmium toxicity inhibition of plants, and significantly promote the growth of plants. And after the AtCRK10 gene is mutated, the opposite result is presented, and the crk10 mutant material is more sensitive to cadmium poisoning, indicating that the AtCRK10 gene has a significant effect on the growth of Arabidopsis and the removal of the cadmium toxicity inhibition. The AtCRK10 gene enhances the cadmium tolerance of plants under cadmium stress conditions, has important significance for the biological function of Arabidopsis adapting to cadmium stress regulation, can well regulate the growth and development of Arabidopsis under cadmium stress, has great value in improving the growth condition of crops in cadmium-polluted areas and increasing grain yield, and is worthy of promotion and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 These are the quantitative PCR results of Arabidopsis thaliana AtCRK10 gene under cadmium treatment conditions (cadmium treatment concentration was 75 μM CdCl2; 0, 6, and 24 h represent samples collected without treatment, 6 h after treatment, and 24 h after treatment, respectively; data are the means and standard errors of 4 biological replicates, and different letters indicate significant differences at the P < 0.05 level (one-way ANOVA and Turkey's test)).

[0028] Figure 2The results of identification of mutant materials (ab) and the relative expression of AtCRK10 in different materials (c) (the quantitative PCR results in the figure are the mean and standard error of 4 biological replicates, and different letters indicate significant differences at the P<0.05 level (one-way ANOVA and Turkey's test)).

[0029] Figure 3 The diagram shows the effects of different genetic materials after 10 days of normal treatment (CK) and cadmium treatment (Cd: 75 μM CdCl2) (the overexpression materials are CRK10_OE-1_X (abbreviated as OE-1_X) and CRK10_OE-2_1 (abbreviated as OE-2_1), the mutant materials are crk10-1 and crk10-2 and the Columbia wild-type material (Col-0); the scale bar in the figure is 2 cm).

[0030] Figure 4 This is the result graph of the effects of normal treatment and cadmium treatment on fresh weight, fresh weight chlorophyll, root length and lateral root number of different materials after 10 days (the data in the figure are the mean and standard error of 16 biological replicates; different letters indicate significant differences among different materials under the same treatment at the P<0.05 level (one-way ANOVA and Turkey's test)). DETAILED DESCRIPTION

[0031] The present invention is further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0032] Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0033] Example 1 Expression pattern of AtCRK10 gene under different cadmium treatment time

[0034] 1. Design of quantitative PCR primers

[0035] The genomic sequence of the Arabidopsis thaliana cysteine-rich receptor-like kinase (CRK) AtCRK10 gene (gene number: AT4G23180) was downloaded from the website, and its nucleotide sequence is shown in SEQ ID NO: 1, and its protein sequence is shown in SEQ ID NO: 2. According to the genomic sequence, the quantitative amplification primers specific to the AtCRK10 gene were designed and synthesized: AtCRK10_qF (SEQ ID NO: 3): 5'-TGGTACCTATGGTTACATGTCTC-3', AtCRK10_qR (SEQ ID NO: 4): 5'-CCACGAGTTCTAATGGTCGC-3'.

[0036] 2. Planting and processing of Arabidopsis materials

[0037] Arabidopsis thaliana (Columbia wild type) seeds of the same period were selected and disinfected with 75% and 95% alcohol respectively, and then dried in a clean bench; the disinfected seeds were evenly sown on the surface of the solid culture medium (1 / 2MS) and placed in a dark environment at 4°C for 2-3 days to break the seed dormancy. Then they were placed in an artificial climate chamber (16 hours of light at 24°C / 8 hours of darkness at 24°C) for 4-5 days. When the seeds germinated and the leaves slightly opened, they were transplanted into the treatment medium for treatment: cadmium treatment (Cd: 75μM CdCl2), 6 biological replicates. Samples were taken after 0h, 6h and 24h of treatment, and appropriate amounts of roots and leaves were cut, frozen in liquid nitrogen, and stored at -80°C.

[0038] 3. Extraction of Total RNA

[0039] Weigh about 0.2g of plant sample into a 2mL centrifuge tube and immediately freeze it with liquid nitrogen; add a steel ball and 1mL Trizol to the centrifuge tube, place it in a grinder and grind for 2min, then let it stand at room temperature for 5 minutes; after grinding, add 200μL of chloroform, shake vigorously, let it stand for 5min, and centrifuge at 4℃, 13000rpm for 15min; after centrifugation, aspirate 600μL of the upper supernatant to a clean 1.5mL centrifuge tube, add an equal volume of isopropanol, shake evenly, and let it stand for 10min; after standing, centrifuge at 4℃, 13000rpm for 10min, remove the supernatant; add 500μL 70%-75% ethanol, centrifuge at 4℃, 13000rpm for 5min, repeat this step once; after aspirating the supernatant, air-dry the precipitate in a fume hood, add 20μL of sterile water, dissolve the precipitate and measure the OD value.

[0040] 4. Preparation and detection analysis of real-time fluorescence quantitative PCR samples

[0041] A one-step method was used to remove genomic DNA and perform RNA reverse transcription. The reaction system and reaction conditions of reverse transcription are shown in Tables 1 and 2.

[0042] Table 1 RNA reverse transcription system

[0043]

[0044] Table 2 RNA reverse transcription conditions

[0045]

[0046]

[0047] A portion of the reverse transcribed cDNA was mixed in equal volumes, and then diluted 3 times in sequence, and diluted to 7 points in total as the standard sample required for the standard curve. The remaining cDNA was diluted 10 times and used as a template for quantitative PCR, AtACTIN2 was used as an internal reference gene, and AtCRK10 gene-specific quantitative amplification primers were used to perform real-time quantitative PCR according to the reaction system and reaction conditions in Tables 3 and 4.

[0048] Table 3 Quantitative PCR reaction system (20 μL reaction system)

[0049]

[0050] Table 4 Quantitative PCR reaction conditions

[0051]

[0052] After the amplification process is completed, the standard curve R2 is adjusted to not less than 0.99, and the efficiency value is between 70% and 120%, and then the data is exported, the relative expression is calculated, and the difference analysis is performed.

[0053] The results are as follows Figure 1 As shown, the expression of AtCRK10 gene in Arabidopsis increases with the increase of cadmium treatment time; and the relative expression of AtCRK10 after 24h of cadmium treatment is 2.78 times that of untreated conditions and 2.22 times that of 6h of treatment. This shows that the expression of AtCRK10 gene in Arabidopsis is significantly upregulated by cadmium stress and is significantly upregulated with the increase of cadmium treatment time.

[0054] Example 2 Construction of AtCRK10 gene overexpression vector

[0055] 1. Amplify the target fragment

[0056] Using Arabidopsis cDNA as a template, forward and reverse specific primers AtCRK10_OE_F (SEQ ID NO: 5): 5'-ATGAGAAGAAACACAGATCAAGAAAG-3', AtCRK10_OE_R (SEQ ID NO: 6): 3'-TCGAGGATGTATATCTGTGATGGA-5' of the target gene AtCRK10 were used to amplify the full-length CDS sequence of the target gene according to the system and conditions specified in the instruction manual of the high-fidelity enzyme.

[0057] 2. Restriction digestion of vector pCAMBIA1300 and ligation

[0058] Use single or double enzyme digestion method, according to the reaction temperature of restriction enzyme, digest the vector successively or simultaneously, connect the target fragment with linearized vector according to the instruction of recombinant cloning enzyme, and connect the target gene to pCAMBIA1300 vector.

[0059] 3. Transformation of E. coli

[0060] Pipette 10μL of the ligation product, blow it into the competent E. coli, mix gently, place it on ice for 30min, place it in a 42℃ water bath for 1min, and then transfer it to ice again for 2-3 minutes. Then add 500μL of LB culture solution to the clean bench and culture it on a 37℃ shaker at 220rpm for 1h. Centrifuge the bacterial solution at 8000rpm for 1min, aspirate 500μL of the supernatant on the clean bench, gently pump and mix the remaining LB culture solution and bacteria, and evenly apply the bacterial solution to the LB solid culture medium containing the corresponding antibiotics, culture it at 37℃ overnight (about 12-16h), pick a single clone and shake it (at least 4h), and detect it. Pick a single clone with the correct detection band position and send it to a biological company for sequencing. After the sequencing results are correct, add glycerol and store it in a -80℃ refrigerator for use or for the next experiment.

[0061] 4. Transformation of Agrobacterium

[0062] Add the corresponding antibiotics and an appropriate amount of correctly sequenced Escherichia coli monoclonal bacterial solution to a 50mL centrifuge tube that has been sterilized at high temperature and high pressure, extract the plasmid according to the instructions of the plasmid extraction kit, and determine the concentration of the resulting plasmid solution. Pipette 10-100μg of Escherichia coli plasmid, add it to Agrobacterium competent medium (GV3101), ice bath for 5min, liquid nitrogen quick freeze for 5min, heat shock at 37℃ for 5min, and then place on ice for 2-3min. Add 800μL YEP liquid culture medium to the clean bench, culture at 28℃ shaker at 220rpm for 2-3h, draw an appropriate amount of bacterial solution and apply it to solid YEP solid culture containing the corresponding antibiotics, culture at 28℃ for 24-48h, pick a single clone and shake bacteria and test, select one of the monoclonal clones with the correct detection band position, add glycerol and store at -80℃ to obtain the AtCRK10 gene overexpression vector.

[0063] Example 3 Arabidopsis genetic transformation

[0064] 1. Cultivation of wild-type Arabidopsis

[0065] Wild-type Arabidopsis seeds (Col-0) were evenly sown on the surface of the nutrient pots filled with substrate, and some water was sprayed, then covered with plastic film to keep moisture, and placed in 4℃ dark conditions for 2-3 days to break seed dormancy. Then placed in an artificial climate room (16 hours of light at 24℃ / 8 ​​hours of darkness at 24℃) for cultivation, and after a week, they were transplanted into nutrient pots, 3-4 plants per pot. When the first bolt was 1cm, the main stem was cut off to increase the number of branches. When the Arabidopsis was in full bloom next time, Agrobacterium liquid could be used for transformation.

[0066] 2. Transformation of Arabidopsis thaliana inflorescence with Agrobacterium liquid

[0067] The Agrobacterium GV3101 preservation solution carrying the target vector was activated by streaking. First, it was inoculated onto a 50mL YEP (Kan+Rif) culture plate and cultured at 28°C for 2 days. After the bacteria grew, a single clone was picked and 2mL YEP (Kan+Rif) liquid was shaken in a small tube. Then, the Agrobacterium solution was expanded and cultured in 350mL YEP (Kan+Rif) liquid, and cultured at 200rpm at 28°C for 20 hours. The bacterial solution was centrifuged at 4000rpm for 10 minutes, the supernatant was discarded, and then the Agrobacterium was resuspended with one volume of fresh infiltration solution (5% sucrose + MgCl2 2.03g / L + Silwet L-77 0.02%) and then transferred to a 500mL beaker. Immerse the aerial part of the flowering Arabidopsis in the Agrobacterium solution for 10 seconds and stir gently, then take out the soaked plant, cover the transformed Arabidopsis with a plastic lid, and place it on its side for 16-24 hours to ensure high humidity. The next day, remove the plastic lid and place it in the Arabidopsis culture room for growth and harvest the T0 generation seeds when mature.

[0068] 3. Identification of transgenic Arabidopsis offspring

[0069] Sterilize the mature T0 generation seeds, first with 75% alcohol, then with 100% alcohol, and finally blow dry the alcohol; sow the sterilized seeds on 1 / 2MS medium (containing hygromycin × 1000), screen the T1 generation transgenic Arabidopsis, and the Arabidopsis seedlings with long taproots indicate that they are hygromycin resistant and may be transgenic materials; then transfer the Arabidopsis seedlings with long taproots to the matrix for growth, and harvest the T1 generation transgenic material seeds. Then repeat the previous step, use hygromycin to screen, see the length of the Arabidopsis seedling taproot, and whether the number meets 3:1 (single copy insertion). If it meets, it can be determined that it is a transgenic material, and harvest the T2 generation Arabidopsis seeds when mature. The same method is used to screen the T2 generation Arabidopsis seeds for fully resistant T3 generation seeds, which are the final homozygous line materials.

[0070] Finally, the overexpression CRK10 strains CRK10_OE-1_X and CRK10_OE-2_1 were screened, and crk10 materials with T-DNA insertion mutations were purchased from the company, including crk10 mutant strains crk10-1 and crk10-2. After the four different materials were identified, the results were as follows Figure 2 As shown, the crk10 mutant was generated by inserting T-DNA into the CRK10 exon region ( Figure 2 a), and PCR analysis confirmed the insertion of T-DNA ( Figure 2 b). The relative expression of CRK10 in different materials was identified by quantitative PCR. Compared with the wild type, the relative expression of CRK10 in CRK10_OE-1_X and CRK10_OE-2_1 materials was 46.7 times and 39.3 times that of the wild type, respectively ( Figure 2 c); the relative expression levels of CRK10 in the mutant materials crk10-1 and crk10-2 were 3% and 17% of the wild type, respectively.

[0071] Example 4 Functional analysis of overexpression of CRK10 and crk10 mutants

[0072] 1. Seed disinfection

[0073] Place mature and plump Arabidopsis seeds (CRK10_OE-1_X, CRK10_OE-2_1, crk10-1, crk10-2 and wild-type materials) into 2 mL tubes respectively. On a clean bench, suck 1 mL of 75% alcohol into the 2 mL tube, cover the seeds and stir evenly, then suck out the alcohol. The operation time should not exceed 1 minute. Then, disinfect them for the second time with 95% alcohol, repeat the previous step, and then put them on the clean bench to let the seeds dry.

[0074] 2. Seed germination

[0075] The sterilized seeds were sown on a 1 / 2MS plate with normal nutrition, and then placed in a dark condition at 4°C for 2-3 days to break the seed dormancy, and then placed in an artificial climate chamber (16 hours light at 24°C / 8 hours dark at 24°C) for cultivation.

[0076] 3. Cadmium treatment

[0077] Three days after the seeds germinated, Arabidopsis seedlings with the same growth potential were transferred to solid culture medium plates under different conditions for treatment: normal treatment (CK) and cadmium treatment (Cd: 75 μM CdCl2), and cultured in an artificial climate chamber (16 hours of light at 24°C / 8 hours of darkness at 24°C); after 10 days of treatment under different conditions, the relevant physiological indicators (fresh weight, fresh weight chlorophyll, main root length, and number of lateral roots) after treatment with different materials were counted.

[0078] The growth phenotype results of Arabidopsis materials under different conditions are as follows Figure 3 As shown, it can be seen that compared with the normal treatment, the fresh weight of Arabidopsis wild-type materials under cadmium treatment was reduced, the fresh weight chlorophyll was reduced, the main root length was shortened, and the number of lateral roots was reduced, while the CRK10 overexpression material alleviated the symptoms under cadmium stress, and the crk10 mutant material was more sensitive to cadmium toxicity.

[0079] Further comparison of the specific physiological index values ​​of different materials showed that Figure 4 As shown in the figure, under cadmium treatment, the CRK10 overexpression material increased the plant fresh weight, fresh weight chlorophyll, taproot length, and the number of lateral roots. The fresh weight of the two overexpression materials increased by 18.0% and 18.5% respectively compared with the wild type material; the fresh weight chlorophyll increased by 43.8% and 29.3% respectively; the taproot length increased by 98.2% and 113.4% respectively; and the number of lateral roots was 2.8 times and 2.45 times that of the wild type, respectively. The fresh weight of the two crk10 mutant materials decreased by 33.3% and 29.8% respectively compared with the wild type material; the fresh weight chlorophyll decreased by 20.2% and 23.2% respectively; the taproot length decreased by 22.6% and 23.6% respectively; and the number of lateral roots decreased by 82.5%. It shows that overexpressing the AtCRK10 gene in Arabidopsis can significantly increase the fresh weight, fresh weight chlorophyll, taproot length and lateral root number of Arabidopsis under cadmium toxicity stress, while the mutant showed the opposite result. This indicates that the AtCRK10 gene may be involved in alleviating the stress and detoxifying cadmium under cadmium toxicity conditions in plants, and overexpressing the AtCRK10 gene can alleviate cadmium stress in Arabidopsis, eliminate the inhibitory effects of cadmium toxicity, and promote plant growth.

[0080] In summary, the study showed that the expression of Arabidopsis AtCRK10 gene was significantly upregulated by cadmium stress, and was significantly upregulated with the increase of cadmium treatment time. Overexpression of AtCRK10 gene in Arabidopsis can alleviate the symptoms of Arabidopsis under cadmium toxicity, and significantly increase the fresh weight, fresh weight chlorophyll, root length and lateral root number of Arabidopsis under cadmium stress, relieve the inhibitory effect of cadmium toxicity on plants, and significantly promote the growth of plants. After the AtCRK10 gene was mutated, the opposite result was presented, and the crk10 mutant material was more sensitive to cadmium toxicity. It shows that the AtCRK10 gene has a significant effect on the growth of Arabidopsis and relieves the inhibitory effect of cadmium toxicity. The AtCRK10 gene enhances the Cd tolerance of plants under Cd stress conditions, which is of great significance to the biological function of Arabidopsis in adapting to cadmium stress regulation. It can well regulate the growth and development of Arabidopsis under cadmium stress, and has great value in improving the growth of crops in cadmium-polluted areas and increasing grain yield. It is worthy of promotion and application.

[0081] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.

Claims

1. Use of the AtCRK10 gene as shown in SEQ ID NO.1 in alleviating cadmium stress in Arabidopsis thaliana.

2. Use of the AtCRK10 gene as shown in SEQ ID NO.1 in preparing a preparation for alleviating cadmium stress in Arabidopsis thaliana.

3. Use of the AtCRK10 gene as shown in SEQ ID NO.1 in improving the tolerance of Arabidopsis thaliana to cadmium stress.

4. Use of the AtCRK10 gene as shown in SEQ ID NO.1 in promoting the growth of Arabidopsis thaliana under cadmium stress.

5. Use of the AtCRK10 gene as shown in SEQ ID NO.1 in cultivating Arabidopsis plants resistant to cadmium stress.

6. Use of a preparation for promoting the expression of the AtCRK10 gene as shown in SEQ ID NO. 1 in alleviating plant cadmium stress, or in preparing a product for alleviating plant cadmium stress.

7. Use of a preparation for promoting the expression of the AtCRK10 gene as shown in SEQ ID NO. 1 to promote plant growth under cadmium stress, or in the preparation of a product for promoting plant growth under cadmium stress.

8. Use of a preparation for promoting the expression of the AtCRK10 gene as shown in SEQ ID NO. 1 in cultivating plants resistant to cadmium stress.

9. A method for alleviating cadmium stress in Arabidopsis thaliana or promoting the growth of Arabidopsis thaliana under cadmium stress, characterized in that: The plants were treated with a preparation containing an overexpressed AtCRK10 gene, the gene sequence of AtCRK10 being shown as SEQ ID NO.

1.

10. A method for cultivating cadmium-resistant plants, characterized in that: The AtCRK10 gene is overexpressed in the plant, or a recombinant vector containing the overexpressed AtCRK10 gene is introduced into the plant, wherein the gene sequence of the AtCRK10 is shown in SEQ ID NO.1.

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