Method for improving cadmium accumulation and transport capability of sedum plumbizincicola

Abstract

The invention discloses a method for improving cadmium accumulation and transport capacity of sedum plumbizincicola, and relates to the field of plant heavy metal pollution remediation. The method comprises the following steps: preparing a selenium solution with the selenium ion concentration of 5 [mu] M, adding the selenium solution into the cadmium-polluted water environment, planting sedum plumbizincicola seedlings in the cadmium-polluted water environment, and carrying out cultivation management; plant pretreatment: culturing sedum plumbizincicola seedlings in non-polluted soil for at least three generations, selecting sedum plumbizincicola plants which are uniform in size and healthy in growth vigor, and pre-culturing the sedum plumbizincicola plants in a Hoaglands nutrient solution for two weeks for rooting; the healthy plants are transferred into Hoagland nutrient solution containing selenium solution to be cultured, and the plants are harvested after 28 days. According to the method, the high-concentration cadmium polluted water environment is simulated, and selenium is applied in the growth period of the sedum plumbizincicola, so that absorption and enrichment of the sedum plumbizincicola on cadmium are promoted, the plant biomass is improved, the toxic effect of cadmium on plants is relieved, and efficient remediation of the sedum plumbizincicola on the cadmium polluted environment is achieved.

Description

Technical Field

[0001] The present invention relates to the field of phytoremediation of heavy metal pollution, and particularly to a method for improving the cadmium accumulation and transport capacity of a hyperaccumulator, Sedum plumbizincicola. Background Art

[0002] Heavy metal pollution caused by human activities such as industrial and agricultural production has become one of the global environmental problems. Cadmium is a heavy metal element that is highly harmful to plant growth and development. Cadmium has the characteristics of high mobility and being easily absorbed by plant roots. When excessive cadmium is ingested by plants, it will inhibit the growth and development of plants and even cause cadmium poisoning. Traditional physical and chemical remediation technologies are not applicable to large-scale cadmium pollution remediation due to their high remediation costs. Phytoremediation technology is considered an eco-friendly cadmium pollution remediation technology, and its remediation effect largely depends on the absorption and accumulation capacity of hyperaccumulator plants for heavy metals. However, due to the characteristics of slow growth and small biomass of plants under heavy metal stress, the phytoremediation efficiency is affected, which to a certain extent hinders the large-scale application of phytoremediation technology. In order to alleviate the toxicity of cadmium to plants and improve the phytoremediation efficiency, it is necessary to start from multiple aspects such as improving the antioxidant level of plants, protecting the photosynthetic organs of plants, and promoting the growth and development of plants to reduce the negative impact of cadmium on plant physiological growth.

[0003] Selenium is a trace element beneficial to plant growth and development. Under heavy metal stress environment, selenium plays an important role in promoting plant growth and development. After plants absorb an appropriate amount of selenium, they can reduce the toxicity of cadmium to plants through various metabolic mechanisms in the plant body. Appropriate selenium can promote the growth and development of plants, increase the content of photosynthetic pigments, improve the antioxidant defense system, enhance the assimilation of carbohydrates and nitrogen, and improve the absorption of other essential elements by plants, thus effectively alleviating the plant stress induced by cadmium. Under medium and low concentration cadmium pollution, selenium can alleviate the adverse effects on the growth and development of crops, but medium and low concentration cadmium pollution will not have a significant impact on the growth and development of hyperaccumulator plants, while it will have an impact on the growth and development of hyperaccumulator plants only under high concentration cadmium pollution. There are few studies on the effect of selenium on alleviating the cadmium toxicity of hyperaccumulator plants and its cadmium absorption and transport under high concentration cadmium pollution environment. It has been found that selenium can activate the plant protection mechanism, reduce oxidative stress, and improve the arsenic absorption of Pteris vittata. So far, there has been no report on the effect of selenium on the growth and development and cadmium accumulation of the hyperaccumulator Sedum plumbizincicola under cadmium pollution environment. Therefore, it is very necessary to explore the effect of selenium on the cadmium accumulation and transport of Sedum plumbizincicola under cadmium treatment. Summary of the Invention

[0004] In view of the limitations such as slow growth and small biomass commonly existing in hyperaccumulator plants in phytoremediation technology, the present invention proposes a method for improving the cadmium accumulation and transportation ability of Sedum plumbizincicola. By simulating a water environment polluted by high-concentration cadmium and applying selenium during the growth period of Sedum plumbizincicola, the absorption and enrichment of cadmium by Sedum plumbizincicola are promoted, the plant biomass is increased, the toxic effect of cadmium on plants is alleviated, and the efficient remediation of cadmium-polluted environment by Sedum plumbizincicola is realized.

[0005] To achieve the above object, the technical solution of the present invention is as follows: A method for improving the cadmium accumulation and transportation ability of Sedum plumbizincicola, comprising the following steps: configuring a selenium solution with a selenium ion concentration of 5 μM and adding it to the cadmium-polluted water environment, planting Sedum plumbizincicola seedlings in the cadmium-polluted water environment, and performing cultivation management; in the selenium solution, the selenium ion concentration is added in the form of Na2SeO3 and calculated as Se; (1) Plant pretreatment: After taking Sedum plumbizincicola seedlings and culturing them in non-polluted soil for at least three generations, selecting Sedum plumbizincicola plants with uniform size and healthy growth and pre-culturing them in Hoaglands nutrient solution for two weeks to root; (2) Hydroponic treatment: Selecting healthy plants that have been pre-cultured for two weeks and have the same growth trend and transferring them to Hoagland nutrient solution containing 0-100 μM selenium solution for cultivation, and harvesting the plants after 28 days; The Sedum plumbizincicola plants of the healthy plants are: plants with 4-8 true leaves and a plant length of 6-7 cm; The cultivation environment for the plant pretreatment and hydroponic treatment is: a 16-hour light cycle, with day and night temperatures of 26°C and 20°C respectively, a relative humidity of 70%, a light intensity of 300 µmol / m 2 / s, maintaining continuous aeration for 24 hours, changing the Hoagland nutrient solution every 3 days, and adjusting the pH of the Hoagland nutrient solution to 5.8 with 0.1 mol / L NaOH solution or 0.1 mol / L HCl solution.

[0006] Compared with the prior art, the beneficial effects are as follows: The present invention provides a method for improving the cadmium accumulation and transportation ability of Sedum plumbizincicola, and uses the low-promotion and high-inhibition effect of selenium on plant growth and development and cadmium absorption to increase the cadmium content and biomass of Sedum plumbizincicola, thereby improving the cadmium accumulation and transportation ability of Sedum plumbizincicola and effectively promoting the efficient remediation of cadmium-polluted areas by Sedum plumbizincicola. Description of the Drawings

[0007] Figure 1 Effects of different cadmium and selenium treatments on the biomass of Sedum plumbizincicola; Figure 2 Effects of different cadmium and selenium treatments on the cadmium concentration of Sedum plumbizincicola; Figure 3Cadmium transport coefficient of Sedum plumbizincicola under different cadmium and selenium treatments; Figure 4 Effects of different cadmium and selenium treatments on photosynthetic pigment contents of Sedum plumbizincicola; Figure 5 Effects of different cadmium and selenium treatments on the activities of catalase (A), superoxide dismutase (B) and peroxidase (C) of Sedum plumbizincicola; Figure 6 Effects of different cadmium and selenium treatments on malondialdehyde content of Sedum plumbizincicola; Figure 7 Transmission electron micrographs (2.0 μm) of root tip cells of Sedum plumbizincicola under control (A), 100 μM cadmium (B) and 100 μM cadmium plus 5 μM selenium (C); where: CW represents cell wall; N represents nucleus; M represents mitochondrion; GB represents Golgi body; Figure 8 Statistical count of the number of differentially expressed genes in different comparison groups; Figure 9 KEGG classification analysis of CK vs Cd comparison group; Figure 10 KEGG enrichment analysis of CK vs Cd comparison group; Figure 11 KEGG classification analysis of Cd vs Cd / Se5 comparison group; Figure 12 KEGG enrichment analysis of Cd vs Cd / Se5 comparison group.

[0008] Figures 1-6 Among them, the data are the mean plus standard error of 3 replicates (n = 3). The data were analyzed by Duncan's multiple comparison, and different lowercase letters in each figure represent significant differences under different concentrations of cadmium or selenium solutions (p < 0.05). Se0, Se5, Se25, Se50 and Se100 represent selenium concentrations of 0, 5, 25, 50 and 100 μM. Cd0, Cd50, Cd100 and Cd150 represent cadmium concentrations of 0, 50, 100 and 150 μM respectively. Specific implementation manners

[0009] The following describes the specific implementation manners in detail with reference to the accompanying drawings, but it should be understood that the protection scope of the present invention is not limited by the specific implementation manners. The raw materials and reagents used in the examples are commercially available unless otherwise specified.

[0010] In the example, the hydroponic treatment was carried out using a polyethylene plastic box for cultivation. The diameter of the plastic box was 17.5 cm and the height was 8.5 cm.

[0011] In the examples, sodium selenite (Na2SeO3) and cadmium chloride (CdCl2·2.5H2O) were both purchased from Sinopharm Chemical Reagent Co., Ltd.

[0012] The basic composition of the Hoagland nutrient solution used in the examples was: Ca(NO3)2·4H2O 2.00 mM, CuSO4·5H2O 0.20 μM, KH2PO4 0.10 mM, MgSO4·7H2O 0.50 mM, KCl 0.10 mM, K2SO4 0.70 mM, H3BO3 10.00 μM, MnSO4·H2O 0.50 μM, ZnSO4·7H2O 0.50 μM, (NH4)6Mo7O 24 0.01 μM.

[0013] Example 1 To simulate the cadmium-polluted water environment, a hydroponic experiment was conducted. Each culture box was filled with selenium and cadmium solutions at different concentrations and 1 L of Hoagland nutrient solution.

[0014] Sedum plumbizincicola was collected from an abandoned lead-zinc mining area in Tianli Village, Qujiang District, Quzhou City, Zhejiang Province. After growing Sedum plumbizincicola seedlings in non-polluted soil for at least three generations, healthy Sedum plumbizincicola plants with uniform size and growth vigor were pre-cultured in Hoagland nutrient solution for two weeks to take root. Healthy plants with the same growth vigor were selected as experimental materials, and 6 plants were cultured in each pot. Cadmium solutions with concentrations of 0 (control group), 50, 100, and 150 μM were prepared by dissolving CdCl2·2.5H2O in deionized water to mimic the pollution of water environment caused by cadmium metal ions. The experiment was carried out in 4 groups, with 3 parallels in each group, and a total of 12 pots were cultured. After 28 days of hydroponics, the samples were harvested and processed.

[0015] The laboratory culture environment (i.e., plant pretreatment and hydroponic treatment) was as follows: The pH of the Hoagland nutrient solution was adjusted to 5.8 with 0.1 mol / L NaOH solution or 0.1 mol / L HCl solution, and it was changed every 3 days to maintain continuous aeration for 24 h. The Sedum plumbizincicola undergoing hydroponics was cultured in an artificial climate chamber with day and night temperatures of 26°C and 20°C respectively, a light cycle of 16 hours, a light intensity of 300 μmol / m 2 / s, and a humidity of about 70%. To keep the volume of the nutrient solution constant, the volume lost due to transpiration was supplemented with Hoagland nutrient solution every day.

[0016] Example 2 Based on the operation of Example 1, dissolve Na2SeO3 in deionized water to prepare a selenium solution with a concentration of 5 μM. Similarly, set four cadmium solutions with concentrations of 0, 50, 100, and 150 μM and mix them. Conduct experiments in 4 groups, with 3 replicates in each group, and cultivate a total of 12 pots.

[0017] Example 3 Based on the operation of Example 1, dissolve Na2SeO3 in deionized water to prepare a selenium solution with a concentration of 25 μM. Similarly, set four cadmium solutions with concentrations of 0, 50, 100, and 150 μM and mix them. Conduct experiments in 4 groups, with 3 replicates in each group, and cultivate a total of 12 pots.

[0018] Example 4 Based on the operation of Example 1, dissolve Na2SeO3 in deionized water to prepare a selenium solution with a concentration of 50 μM. Similarly, set four cadmium solutions with concentrations of 0, 50, 100, and 150 μM and mix them. Conduct experiments in 4 groups, with 3 replicates in each group, and cultivate a total of 12 pots.

[0019] Example 5 Based on the operation of Example 1, dissolve Na2SeO3 in deionized water to prepare a selenium solution with a concentration of 100 μM. Similarly, set four cadmium solutions with concentrations of 0, 50, 100, and 150 μM and mix them. Conduct experiments in 4 groups, with 3 replicates in each group, and cultivate a total of 12 pots.

[0020] The treatment concentrations of the above Examples 1-5 are shown in Table 1: Table 1 Cadmium and selenium concentrations in different examples After cultivating the samples in Examples 1-5 for 28 days, harvest and process the samples. Before collecting and processing the samples, soak the plant roots in a 20 mM EDTA-Na2 solution for 15 min to remove the metal ions adsorbed on the root surface; then rinse the plants with deionized water multiple times and dry the remaining moisture on the surface. Separate the samples into roots, stems, and leaves. Among them, part of the samples are quickly frozen in liquid nitrogen and stored in a -80°C ultra-low temperature refrigerator for physiological index analysis; another part of the samples are placed in an oven at 105°C for 30 min to inactivate enzymes, and then dried at 60°C to constant weight for standby, and record the dry weight (DW) for biomass determination (the results are as Figure 1 shown). After grinding the dried samples, they are used to determine the cadmium content in the roots, stems, and leaves of the plants.

[0021] Under stress, plant biomass is one of the important indicators reflecting plant health. From Figure 1It can be seen that when 5, 25, 50, and 150 μM selenium were added, the biomass of Sedum plumbizincicola increased under different cadmium concentration treatments. Only under the treatment of 100 μM selenium and 150 μM cadmium, the biomass of Sedum plumbizincicola decreased by 12.93%. This indicates that within the above concentration range, adding 5 μM selenium has the best promoting effect on the growth of Sedum plumbizincicola.

[0022] Determination of cadmium content: According to the national food safety standard "Determination of cadmium in foods" (GB 5009.15 - 2014), the cadmium content in Sedum plumbizincicola was determined. Weighed plant samples were digested with a mixed solution of nitric acid - perchloric acid (v:v = 4:1). After the sample digestion solution was made up to volume, the absorbance value of cadmium in the test solution was measured using a flame atomic absorption spectrometer (TAS - 990, Beijing Purkinje General Instrument Co., Ltd., China). Through the standard curve method, the content of cadmium was quantitatively analyzed and the cadmium transport coefficient was calculated. The results are as Figure 2 and Figure 3 shown.

[0023] It can be Figure 2 seen that when 5 μM selenium was added, under different cadmium concentration treatments, the cadmium content in the leaves of Sedum plumbizincicola increased significantly, and there were differences in the change range of cadmium content in the stems and roots; under the treatment of 5 μM selenium and 100 μM cadmium, the increase in cadmium content in the leaves of the plant was the most significant, increasing by 27.62%. When 25, 50, and 100 μM selenium were added, under different cadmium concentration treatments, the cadmium content in the leaves of Sedum plumbizincicola decreased, but the cadmium content in the roots increased significantly. By calculating the change in the cadmium transport coefficient of Sedum plumbizincicola, it can be Figure 3 seen that when 5 μM selenium was added, under the treatment of 100 μM cadmium, the transport coefficient of stem / root increased by 1.26%; when 5 μM selenium was added, under the treatment of 50 μM cadmium, the transport coefficient of leaf / stem increased by 2.06%. When high - concentration selenium (50 or 100 μM) was added, under different cadmium concentration treatments, the cadmium transport coefficient of Sedum plumbizincicola decreased. Thus, it can be seen that within the above concentration range, adding 5 μM selenium under different cadmium concentration treatments increased the cadmium content in the roots, stems, and leaves of Sedum plumbizincicola and improved the cadmium transport ability.

[0024] Determination of photosynthetic pigment content: The photosynthetic pigment content was determined by the 80% acetone extraction method. The fresh plant leaves were washed and dried, avoiding the leaf veins. 0.20 g of leaves at the same position were weighed and placed in a porcelain mortar, and 2 mL of 80% acetone was added and ground into a homogenate. The homogenate was filtered into a 25 mL brown volumetric flask, and the pigments remaining on the filter paper were washed into the volumetric flask with 80% acetone, made up to the mark, and mixed well for measurement. The absorbance values were measured at 663 nm, 646 nm, and 470 nm using a microplate reader. The contents of chlorophyll a, chlorophyll b, chlorophyll, and carotenoids were calculated using the following formula, and the results are as Figure 4 shown.

[0025] It can be seen from Figure 4 that when 5, 25, and 50 μM selenium were added, the contents of chlorophyll a and chlorophyll b in Sedum plumbizincicola were significantly increased under the treatments of 50 and 100 μM cadmium concentrations. Among them, when 5 μM selenium was added, the chlorophyll content reached the highest value of 1.17 mg / g under the treatment of 50 μM cadmium concentration. When different concentrations of selenium were added, the carotenoid content in Sedum plumbizincicola was decreased under the treatments of 100 and 150 μM cadmium. Only when 5 and 50 μM selenium were added, the carotenoid contents in Sedum plumbizincicola were increased by 7.50% and 2.50% respectively under the treatment of 50 μM cadmium. This indicates that adding 5 μM selenium has a significant promoting effect on the photosynthesis of Sedum plumbizincicola within the above concentration range.

[0026] Determination of antioxidant enzyme activity 0.30 g of fresh plant leaves were weighed and placed in a porcelain mortar pre-cooled at 4°C, and 3 mL of potassium phosphate buffer (pH 7.8) was added and ground into a homogenate, which was then completely transferred to a 10 mL clean centrifuge tube. The homogenate was centrifuged at 4°C and 8000g for 10 min, and the supernatant was taken as the crude extract for the determination of enzyme activity.

[0027] (1) Determination of catalase (CAT) activity 100 μL of the crude extract was taken and added to 2.8 mL of phosphate buffer containing EDTA (pH 7.0) and 100 μL of hydrogen peroxide solution. The initial absorbance value A1 was measured at a wavelength of 240 nm using a UV-visible spectrophotometer. After the reaction for 1 min, the final absorbance value A2 was recorded, and the CAT activity was calculated based on the change in absorbance before and after the reaction.

[0028] It can be seen from Figure 5As can be seen from A, when no exogenous selenium was added, the CAT activity of Sedum plumbizincicola decreased significantly by 36.19% - 71.52% under different cadmium concentrations; under the interaction of selenium and cadmium, the CAT activity of Sedum plumbizincicola showed a trend of first increasing and then decreasing. Among them, under the treatments of 5 μM selenium + 100 μM cadmium and 50 μM selenium + 50 μM cadmium, the CAT activity of the plants increased significantly, by 190.95% and 161.29% respectively. Adding 50 or 100 μM selenium decreased the CAT activity.

[0029] (2) Determination of superoxide dismutase (SOD) activity The SOD activity was determined by the nitroblue tetrazolium chloride (NBT) colorimetric method. Add 2.725 mL of a mixed solution composed of phosphate buffer (pH 7.8), EDTA-Na2 solution, methionine solution, nitroblue tetrazolium solution and riboflavin solution to the sample tube, and then add 25 mL of the crude extract and 25 mL of hydrogen peroxide solution. Add an equal amount of the mixed solution and hydrogen peroxide solution to the blank tube, without adding the crude extract. Place the sample tube and the blank tube under sunlight with an illumination intensity of 4000 Lx for reaction for 20 min. At the same time, set up a control tube composed of 2.725 mL of the mixed solution, 25 mL of deionized water and 25 mL of hydrogen peroxide solution, and place the control tube in the dark. Using the control tube as a reference, measure the absorbance at 560 nm with an enzyme-labeling instrument.

[0030] As can be seen from Figure 5 B, when no exogenous selenium was added, under the treatments of 100 and 150 μM cadmium, the SOD activity of the plants decreased by 31.24% and 27.27% respectively; adding 5 and 25 μM selenium increased the SOD activity of Sedum plumbizincicola under different cadmium concentrations. Among them, under the treatment of 5 μM selenium + 150 μM cadmium, the SOD activity of the plants increased most significantly, by 133.31%. Adding high-concentration selenium (50 or 100 μM) significantly decreased the SOD activity of Sedum plumbizincicola.

[0031] (3) Determination of peroxidase (POD) activity The POD activity was determined by the guaiacol colorimetric method. Add 5 μL of the crude extract to the sample tube, and then add 120 μL of guaiacol solution, 30 μL of hydrogen peroxide solution (300 mM), 30 μL of phosphate buffer containing EDTA (pH 7.0) and 60 μL of deionized water respectively. Mix quickly and transfer to a micro quartz cuvette. Use an ultraviolet-visible spectrophotometer to measure the absorbance A1 at 30 s at a wavelength of 470 nm. After the reaction for 1 min, record the final absorbance A2, and calculate the POD activity according to the change in absorbance before and after the reaction.

[0032] As can be seen from Figure 5It can be seen that without exogenous selenium addition, the POD activity of Sedum plumbizincicola showed a decreasing trend under different cadmium concentration treatments. Under the interaction of selenium and cadmium, the addition of 5-50 μM selenium increased the POD activity of the plants to varying degrees. Among them, the treatment with 25 μM selenium and 150 μM cadmium had the most significant increase in the POD activity of the plants, with an increase of 344.40%. The addition of 100 μM selenium decreased the POD activity of Sedum plumbizincicola.

[0033] Thus, it can be seen that within the above concentration range, the addition of 5 μM selenium improved the antioxidant enzyme activity of the plants and alleviated the toxicity caused by high-concentration cadmium to Sedum plumbizincicola under different cadmium concentration treatments.

[0034] Determination of malondialdehyde content: The thiobarbituric acid (TBA) colorimetric method was used to determine the malondialdehyde content. Weighed 0.10 g of fresh plant leaves and placed them in a porcelain mortar pre-cooled at 4°C, added 10% trichloroacetic acid solution, ground them into a homogenate and transferred all of it to a 10 mL centrifuge tube. Centrifuged the homogenate at 4°C and 12,000 g for 10 min, took 2 mL of the supernatant and put it into a 10 mL test tube, then added 2 mL of 0.67% thiobarbituric acid solution. Reacted the mixed solution in a boiling water bath for 10 min, quickly cooled it in an ice bath and then centrifuged it. Subsequently, took an appropriate amount of the supernatant and placed it on a 96-well microplate, and used a microplate reader to measure the absorbance values at 450 nm, 532 nm and 600 nm respectively, and calculated the MDA content. The results are as Figure 5 shown.

[0035] As can be Figure 6 seen, without exogenous selenium addition, the malondialdehyde content in the leaves of Sedum plumbizincicola showed an increasing trend under different cadmium concentration treatments. When 5 μM selenium was added, the malondialdehyde content decreased by 31.49 - 49.21% under different cadmium concentration treatments. When high-concentration selenium (50 or 100 μM) was added, the malondialdehyde content increased by 4.53% - 240.26% under different cadmium concentration treatments. This indicates that within the above concentration range, the addition of 5 μM selenium reduced the degree of membrane lipid peroxidation of plant cells and alleviated the oxidative damage induced by cadmium.

[0036] Ultrastructure of root tip cells: Figure 7It shows the changes in the ultrastructure of the root tip cells of Sedum plumbizincicola. The cell walls of the root tip cells in the control group were smooth and continuous, the cell nuclei were clear, the nucleoli were round, and oval mitochondria and Golgi apparatuses were clearly visible; under the treatment of 100 μM cadmium, a series of morphological changes occurred in the root tip cells of Sedum plumbizincicola, such as cell deformation and rupture, thickening of the cell wall, disappearance of some organelles, etc.; the addition of 5 μM selenium improved the structures of the cell membranes and vacuoles of the root tip cells, maintained the integrity of the cells, and presented clear cell walls and obviously differentiated organelles. This indicates that under high-concentration cadmium stress, the addition of 5 μM selenium improved the ultrastructure of the root tip cells and alleviated the cadmium-induced membrane damage.

[0037] Root transcriptome: To determine the regulatory mechanism of selenium treatment on Sedum plumbizincicola seedlings under cadmium stress, the present invention prepared root samples of Sedum plumbizincicola seedlings with different treatments for RNA-seq experiments, analyzed the transcriptome data, and screened the differentially expressed genes (DEGs) between CK and other treatments. The present invention respectively collected samples of 3 treatments, namely CK (blank), Cd (100 μM cadmium), and Cd / Se5 (100 μM cadmium + 5 μM selenium), and each treatment had 3 replicated samples, totaling 9 samples, for RNA-seq transcriptome analysis. The RNA-seq was commissioned to Shanghai Majorbio Bio-Pharm Technology Co., Ltd. to complete. After the library construction was completed, sequencing was performed using the Illumina platform. After quality control and assembly of the sequencing data, the expression analysis of differentially expressed genes was carried out.

[0038] The DESeq2 software was used to perform differential expression analysis between sample groups with different treatments. The fold change (FC) ≥ 2 and the false discovery rate (FDR) < 0.05 were set as the screening conditions. As Figure 8 shown, a total of 12,626 differentially expressed genes were identified in 2 comparison groups, including 9,288 up-regulated genes and 3,338 down-regulated genes. In the CK vs Cd comparison group, there were 2,264 differentially expressed genes, including 1,623 up-regulated genes and 641 down-regulated genes; in the Cd vs Cd / Se5 comparison group, there were 10,362 differentially expressed genes, including 7,665 up-regulated genes and 2,697 down-regulated genes.

[0039] To further understand the metabolic pathways involved in the differentially expressed genes in response to the interaction between selenium and cadmium in Sedum plumbizincicola, the KEGG database was used to classify and enrich the differentially expressed genes in different comparison groups. In the CK vs Cd comparison group, from Figure 9 the KEGG classification analysis of the CK vs Cd comparison group, it can be seen that the treatment with 100 μM cadmium up-regulated the number of genes in pathways such as environmental adaptation, translation, and carbohydrate metabolism, and increased the number of genes in the replication and repair pathways compared with the down-regulated pathways; fromFigure 10 From the KEGG enrichment analysis of the CK vs Cd comparison group, it can be seen that the differentially expressed genes down-regulated by 100 μM cadmium treatment were mainly enriched in pathways such as isoflavone synthesis, taurine and hypotaurine metabolism, and butyric acid metabolism. The differentially expressed genes induced to be up-regulated were mainly significantly enriched in pathways such as photosynthesis, ribosome, and carbon fixation in photosynthetic organisms. In the Cd vs Cd / Se5 comparison group, Figure 11 From the KEGG classification analysis of the Cd vs Cd / Se5 comparison group, it can be seen that the treatment with 5 μM selenium and 100 μM cadmium increased the number of genes in pathways such as translation, carbohydrate metabolism, folding, sorting and degradation, transport and catabolism, and energy metabolism; Figure 12 From the KEGG enrichment analysis of the Cd vs Cd / Se5 comparison group, it can be seen that the differentially expressed genes up-regulated by the treatment with 5 μM selenium and 100 μM cadmium were mainly significantly enriched in pathways such as photosynthesis, photosynthetic-antenna proteins, biosynthesis of cutin, suberin and wax, and glycerolipid metabolism. The differentially expressed genes down-regulated were mainly significantly enriched in pathways such as zeatin biosynthesis, exopolysaccharide biosynthesis, plant hormone signal transduction, and sesquiterpenoid and triterpenoid biosynthesis. Thus, it can be seen that adding 5 μM selenium can alleviate the toxicity of cadmium to plants by inhibiting the accumulation of reactive oxygen species, protecting photosynthetic organs, and enhancing the life metabolic activities of plants, and thereby promoting the growth of plants.

[0040] The method of the present invention improved the biomass, cadmium content in plants and cadmium transport capacity of Sedum plumbizincicola by adding 5 μM selenium, while adding high concentrations of selenium (50 or 100 μM) had an obvious inhibitory effect on the cadmium transport capacity of Sedum plumbizincicola. In the phytoremediation of cadmium-polluted environments, applying an appropriate concentration of selenium can improve the phytoremediation efficiency of Sedum plumbizincicola for cadmium-polluted environments.

Claims

1. A method for improving the cadmium accumulation and transport ability of Sedum plumbizincicola, characterized in that, It includes the following steps: adding a selenium solution with a selenium ion concentration of 5 μM to the cadmium-polluted water environment, planting Sedum plumbizincicola seedlings in the cadmium-polluted water environment, and carrying out cultivation management; in the selenium solution, the selenium ion concentration is added in the form of Na2SeO3 and calculated as Se. (1) Plant pretreatment: After taking Sedum plumbizincicola seedlings and culturing them in non-polluted soil for at least three generations, select Sedum plumbizincicola plants with uniform size and healthy growth and pre-culture them in Hoaglands nutrient solution for two weeks to take root. (2) Hydroponic treatment: Select healthy plants that have been pre-cultured for two weeks and have the same growth trend, transfer them to Hoagland nutrient solution containing 0 - 100 μM selenium solution for cultivation, and harvest the plants after 28 days. The Sedum plumbizincicola plants of the healthy plants are: plants with 4 - 8 true leaves and a plant length of 6 - 7 cm. The culture environment for the plant pretreatment and hydroponic treatment is as follows: a 16-hour light cycle, with day and night temperatures of 26°C and 20°C respectively, a relative humidity of 70%, a light intensity of 300 µmol / m 2 / s, continuous aeration is maintained for 24 hours, the Hoagland nutrient solution is changed every 3 days, and the pH of the Hoagland nutrient solution is adjusted to 5.8 with 0.1 mol / L NaOH solution or 0.1 mol / L HCl solution.

2. The method for improving the cadmium accumulation and translocation ability of Sedum plumbizincicola according to claim 1, characterized in that, In order to keep the volume of the nutrient solution constant, replenish the volume lost due to transpiration with Hoagland nutrient solution every day.

3. The method for improving the cadmium accumulation and transport ability of Sedum plumbizincicola according to claim 1, wherein, The basic composition of Hoagland nutrient solution is as follows: Ca(NO3)2·4H2O 2.00 mM, CuSO4·5H2O 0.20 μM, KH2PO4 0.10 mM, MgSO4·7H2O 0.50 mM, KCI 0.10 mM, K2SO4 0.70 mM, H3BO3 10.00 μM, MnSO4·H2O 0.50 μM, ZnSO4·7H2O 0.50 μM, (NH4)6Mo7O 24 0.01 μM.

4. The method for improving the cadmium accumulation and transport ability of Sedum plumbizincicola according to claim 1, characterized in that, Prepare a selenium solution with a concentration of 5 μM by dissolving Na2SeO3 in deionized water. Similarly, set up four concentrations of cadmium solutions of 0, 50, 100, and 150 μM and mix them with it. The cadmium solution is added in the form of CdCl2 and calculated as Cd. After culturing for 28 days, harvest and process the samples. Before collecting and processing the samples, soak the plant roots in 20 mM EDTA-Na2 solution for 15 min to remove the metal ions adsorbed on the root surface; then rinse the plants with deionized water multiple times and dry the residual moisture on the surface. Separate the samples into roots, stems, and leaves; among them, part of the samples are quickly frozen in liquid nitrogen and stored in a -80°C ultra-low temperature refrigerator for physiological index analysis; another part of the samples are placed in an oven at 105°C for 30 min to deactivate enzymes, and then dried at 60°C to a constant weight for standby, and record the dry weight (DW) for biomass determination; after grinding the dried samples, they are used to determine the cadmium content in the roots, stems, and leaves of the plants; adding 5 μM selenium has the best promoting effect on the growth of Sedum plumbizincicola under different cadmium concentration treatments.

5. The method for improving the cadmium accumulation and transportation ability of Sedum plumbizincicola according to claim 1, wherein Determine the cadmium content in Sedum plumbizincicola. Weigh the plant samples and digest them with a nitric acid-perchloric acid mixed solution (v:v = 4:1). After the sample digestion solution is fixed in volume, use a flame atomic absorption spectrometer to measure the absorbance value of cadmium in the test solution. Through the standard curve method, quantitatively analyze the cadmium content and calculate the cadmium transport coefficient. Adding 5 μM selenium under different cadmium concentration treatments increases the cadmium content in the roots, stems, and leaves of Sedum plumbizincicola and improves the cadmium transport ability of the plants.

6. The method for improving the cadmium accumulation and translocation ability of Sedum plumbizincicola according to claim 1, characterized in that, Under high-concentration cadmium stress, adding 5 μM selenium increases the antioxidant enzyme activity of the plants and alleviates the toxicity caused by high-concentration cadmium to Sedum plumbizincicola.

7. The method for improving the cadmium accumulation and transportation ability of Sedum plumbizincicola according to claim 1, characterized in that, Use the 80% acetone extraction method to measure the photosynthetic pigment content; under high-concentration cadmium stress, adding 5 μM selenium significantly increases the photosynthetic pigment content of Sedum plumbizincicola.

8. The method for improving the cadmium accumulation and transport capacity of Sedum plumbizincicola according to claim 1, wherein Use the thiobarbituric acid (TBA) colorimetric method to measure the malondialdehyde content; under high-concentration cadmium stress, adding 5 μM selenium reduces the malondialdehyde content in plant cells and reduces the degree of membrane lipid peroxidation.

9. The method for improving the cadmium accumulation and transportation ability of Sedum plumbizincicola according to claim 1, wherein Under high-concentration cadmium stress, adding 5 μM selenium improves the ultrastructure of root tip cells and alleviates cadmium-induced membrane damage.

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