Hydro-fluctuation belt repairing method

Abstract

The invention provides a hydro-fluctuation belt restoration method, and belongs to the technical field of ecological restoration. The method comprises the following steps: planting mulberry leaf grape special No.1 in a hydro-fluctuation belt; the mulberry leaf grape specific No. 1 is preserved in China General Microbiological Culture Collection Center, the address is No.3, No.1 yard, Beichen West Road, Chaoyang District, Beijing, the preservation date is February 19, 2025, and the preservation number is CGMCC No.32201. The mulberry leaf grape specific No. 1 has the advantages that the mulberry leaf grape specific No. 1 can be used for preparing the mulberry leaf grape specific No. 1; when the water level rises, the mulberry leaf grape special No.1 can tolerate water logging and keep a dormant state; after the water level drops, growth can be rapidly recovered, stable vegetation coverage is formed, water and soil loss is effectively reduced, the ecological environment of the hydro-fluctuation belt is improved, and the key problem of ecological restoration and reconstruction of the hydro-fluctuation belt is solved.

Description

Technical Field

[0001] The present invention relates to the technical field of ecological restoration, and particularly relates to a method for restoring the water-level-fluctuation zone. Background Art

[0002] The water-level-fluctuation zone is a special area formed during the water-level rise and fall of rivers, lakes, reservoirs and other water bodies. The frequent water-level rise and fall makes the ecological environment in the water-level-fluctuation zone fragile and special. Many plants are difficult to adapt to this periodic waterlogging and drought environment. Therefore, the vegetation coverage rate in the water-level-fluctuation zone is low. For the water-level-fluctuation zone lacking vegetation protection, soil erosion is serious and the difficulty of ecological restoration is great. In current research or projects on the restoration of the water-level-fluctuation zone, common woody plants applied to the restoration of the water-level-fluctuation zone include Taxodium distichum var. imbricatum, Betula nigra, and Pterocarya stenoptera, etc., and herbaceous plants such as Cynodon dactylon, Xanthium sibiricum, and Cyperus rotundus, etc. The selection of plants has limitations, and there is a lack of plant germplasm of woody vines with fast growth rate and whole-plant waterlogging tolerance applied to the restoration of the water-level-fluctuation zone.

[0003] Vitis, a woody vine plant of the genus Vitis in the Vitaceae family, is one of the oldest fruit tree species in the world. Wild grapes usually have characteristics such as fast growth, strong stress resistance, and wide adaptability, and can expand the important gene pool of the genetic basis of cultivated grapes. Wild grapes can be hybridized with the required cultivated varieties to cultivate various plant germplasm materials with resistance. This is beneficial to agricultural production and the development of local economic crops. However, in recent years, due to various factors such as pollution, overexploitation of resources, and destruction of the living environment, a large number of wild plant populations have died, and the wild species of grapes are no exception. Many species are facing endangerment.

[0004] The prior art points out that the responses of plants under waterlogging stress include: changes in root morphology and structure, improvement of antioxidant enzyme activity, accumulation of osmoregulatory substances, regulation of photosynthesis and respiration, etc. Most plants will have intuitive changes in their morphological and anatomical structures after being waterlogged, and will also initiate a series of physiological and biochemical responses under waterlogging stress to adapt to the adverse environment. Research shows that after water submerges plants, adventitious roots are generated through the roots so that they can grow normally in the harsh waterlogged environment. When the primary roots die in large numbers due to waterlogging, the roots of the plants will quickly grow adventitious roots with waterlogging adaptability. The adventitious roots can grow normally due to their unique structure. The root tip cells have high cell division ability and physiological activity, and well-developed aerenchyma is formed in the elongation zone, which greatly increases the porosity of the internal tissues of the root. Through these structures, the plant's absorption of oxygen can be enhanced, and the aerenchyma can enable gases to diffuse rapidly over long distances in the body, so that the plant's photosynthesis can be restored, and the oxygen transported to the roots is also sufficient to maintain its life activities.

[0005] The physiological and biochemical response of plants under flooding conditions is a complex and sensitive process. First of all, flooding stress has a direct impact on plant roots. Due to the reduced oxygen supply in the soil, the roots cannot carry out normal respiratory metabolism. This hypoxic state not only inhibits the growth and development of the roots, but also may cause oxidative stress in the roots. The accumulation of reactive oxygen species (ROS) causes oxidative damage to biological macromolecules such as cell membranes, proteins and DNA, which may cause lipid peroxidation of cell membranes, protein oxidation and DNA breakage. In response to this oxidation, plants may increase the activity of antioxidant enzymes, such as superoxide dismutase (SOD) and catalase, to remove reactive oxygen species and protect cells from damage. However, as flooding stress continues, plants may gradually lose their ability to adapt to the environment, resulting in a decrease in antioxidant enzyme activity after reaching a peak. This may be due to the collapse of the antioxidant system in the plant or resource limitations, which prevents the plant from continuing to maintain a high level of antioxidant enzyme activity. At this point, plants may face more severe oxidative stress and cell damage. In addition, some transcription factors and aquaporins are also involved in regulating plant stress responses.

[0006] The Three Gorges drawdown zone is located in the Three Gorges area of ​​the Yangtze River. It is a very special ecological environment that is not only affected by water level fluctuations, but also by seasonal climate changes, soil types, vegetation coverage and other factors. Providing a woody vine that can repair the drawdown zone, especially the Three Gorges drawdown zone, can not only provide plant germplasm resources for vegetation restoration and reconstruction in the Three Gorges Dam drawdown zone environment, but also help to gain a deeper understanding of the adaptation mechanism and survival strategy of plants under flooding conditions, and provide new ideas and methods for scientific research in related fields. Summary of the invention

[0007] The purpose of the present invention is to provide a method for repairing a drawdown zone.

[0008] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0009] The invention provides a method for repairing a water-fluctuation zone, and a mulberry leaf grape specific 1 is planted in the water-fluctuation zone; the mulberry leaf grape specific 1 is deposited in the General Microbiological Center of China Microbiological Culture Collection Committee, the address of which is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, the preservation date is: February 19, 2025, and the preservation number is: CGMCC No.32201.

[0010] Preferably, the drawdown zone is the drawdown zone of the Three Gorges Reservoir area.

[0011] Preferably, before the planting, the Mulberry Leaf Grape Specific No. 1 is also propagated, and the propagation is carried out in a nursery or a nutrient pot, and the propagation method is a hard branch cutting method or a soft branch full-light spray cutting method.

[0012] Preferably, the time for propagation by the hardwood cutting method is from February to May, from June to September, or from October to November, and the time for propagation by the softwood full-light spray cutting method is from July to September.

[0013] Preferably, the method for hardwood cutting is as follows: After cutting the hardwood branches of Morus alba var. tomentosa cv. 'Teiyihao' into cuttings with 2-3 buds, insert the morphological lower end of the cuttings into the cutting substrate in the nursery or nutrient pot.

[0014] Preferably, the hardwood branches are one-year-old or perennial branches;

[0015] The morphological lower end of the cutting is obliquely cut, and the morphological upper end is horizontally cut;

[0016] The nutrient pot is a non-woven fabric nutrient pot with a diameter of 12-30 cm;

[0017] The depth of insertion into the cutting substrate is 1 / 3-1 / 2 of the length of the cutting; the cutting substrate is yellow brown soil.

[0018] Preferably, the method for softwood full-light spray cutting is as follows: After cutting the softwood branches of Morus alba var. tomentosa cv. 'Teiyihao' into cuttings with 2-3 buds, insert the morphological lower end of the cuttings into the river sand bed, and use a full-light spray watering device for watering management. After 40-80 days, transplant the rooted seedlings into nutrient pots for cultivation.

[0019] Preferably, the morphological upper end of the cutting has leaves, and 1 / 3-1 / 2 of the leaf area is left;

[0020] The nutrient pot is a non-woven fabric nutrient pot with a diameter of 12-30 cm.

[0021] Preferably, when planting, use the asexual clone nutrient pot seedlings of Morus alba var. tomentosa cv. 'Teiyihao' over two years old; the spacing during planting is 2-3 m×3-5 m; the length×width×depth of the planting hole is 15-35 cm×15-35 cm×25-35 cm.

[0022] Preferably, after planting Morus alba var. tomentosa cv. 'Teiyihao', compact and fix the rootstock of the seedlings, cover the planting hole with a weed-proof cloth and fix the weed-proof cloth around.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] The present invention provides a method for repairing the water-level-fluctuation zone by using Vitis heyneana Roem. et Schult. cv. Tiyi, and the Vitis heyneana Roem. et Schult. cv. Tiyi is planted in the water-level-fluctuation zone area. When the water level rises, the Vitis heyneana Roem. et Schult. cv. Tiyi can tolerate waterlogging and maintain a dormant state; after the water level drops, it can quickly resume growth, form a stable vegetation cover, effectively reduce soil erosion, improve the ecological environment of the water-level-fluctuation zone, and solve the key problems of ecological restoration and reconstruction of the water-level-fluctuation zone. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the drawings in the following description are only the embodiments of the present invention, and those of ordinary skill in the art can also obtain other drawings according to the provided drawings without creative efforts.

[0026] Figure 1 It is the measurement result of the MDA content in the roots of each Vitis heyneana Roem. et Schult. cv. Tiyi after different waterlogging time treatments in Example 2;

[0027] Figure 2 It is the measurement result of the free proline content in the roots of each Vitis heyneana Roem. et Schult. cv. Tiyi after different waterlogging time treatments in Example 2;

[0028] Figure 3 It is the measurement result of the soluble sugar content in the roots of each Vitis heyneana Roem. et Schult. cv. Tiyi after different waterlogging time treatments in Example 2;

[0029] Figure 4 It is the measurement result of the soluble protein content in the roots of each Vitis heyneana Roem. et Schult. cv. Tiyi after different waterlogging time treatments in Example 2;

[0030] Figure 5 It is the measurement result of the peroxidase content in the roots of each Vitis heyneana Roem. et Schult. cv. Tiyi after different waterlogging time treatments in Example 2;

[0031] Figure 6 It is the measurement result of the superoxide dismutase content in the roots of each Vitis heyneana Roem. et Schult. cv. Tiyi after different waterlogging time treatments in Example 2;

[0032] Figure 7 It is the measurement result of the catalase content in the roots of each Vitis heyneana Roem. et Schult. cv. Tiyi after different waterlogging time treatments in Example 2;

[0033] Figure 8Morphological characteristics of each Muscadinia rotundifolia after different flooding time treatments in Example 2; where A in the figure is the Muscadinia rotundifolia plant without flooding treatment, B is the Muscadinia rotundifolia plant after 3 days of flooding treatment, C is the Muscadinia rotundifolia plant after 6 days of flooding treatment, D is the Muscadinia rotundifolia plant after 9 days of flooding treatment, E is the above-ground part of the Muscadinia rotundifolia plant after 12 days of flooding treatment, F is the underground part of the Muscadinia rotundifolia plant after 12 days of flooding treatment, G is the Muscadinia rotundifolia plant after 15 days of flooding treatment, and H is the Muscadinia rotundifolia plant after 30 days of flooding treatment;

[0034] Figure 9 Microscopic structures of the root tips of each Muscadinia rotundifolia after different flooding time treatments in Example 2; where A is the microscopic structure of the root tip cross-section of Muscadinia rotundifolia without flooding treatment, B is the microscopic structure of the root tip cross-section of Muscadinia rotundifolia after 3 days of flooding treatment, C is the microscopic structure of the root tip cross-section of Muscadinia rotundifolia after 6 days of flooding treatment, D is the microscopic structure of the root tip cross-section of Muscadinia rotundifolia after 9 days of flooding treatment, E is the microscopic structure of the root tip cross-section of Muscadinia rotundifolia after 12 days of flooding treatment, F is the microscopic structure of the root tip cross-section of Muscadinia rotundifolia after 15 days of flooding treatment, and G and H are the microscopic structures of the root tip cross-section of Muscadinia rotundifolia after 30 days of flooding treatment;

[0035] Figure 10 Local enlarged views of the microscopic structures of the root tip cross-sections of Muscadinia rotundifolia after 3 days (A), 6 days (B), and 9 days (C) of flooding treatment in Example 2; where a is the periderm, b is the phloem, c is the phloem ray, d is the xylem ray, e is the secondary xylem, f is the primary xylem, g is a large amount of accumulated suberin and lignin, and h is the lysigenous aerenchyma formed by programmed cell death and dissolution of living cells;

[0036] Figure 11 Effect picture of Muscadinia rotundifolia specific No. 1 planted in the water-level-fluctuating zone of the Three Gorges Reservoir Area in Example 3;

[0037] Figure 12 Effect picture of Muscadinia rotundifolia specific No. 1 planted in the water-level-fluctuating zone of the Three Gorges Reservoir Area in Example 3;

[0038] Figure 13 Effect picture of Muscadinia rotundifolia specific No. 1 planted in the water-level-fluctuating zone of the Three Gorges Reservoir Area in Example 3. Detailed implementation manners

[0039] The technical solutions provided by the present invention will be described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0040] Example 1 Obtaining of Muscadinia rotundifolia specific No. 1

[0041] The inventor discovered a wild grape plant in the water-level-fluctuation zone of the Three Gorges. It was identified as Vitis ficifolia and preserved in the General Microbiology Center of the China Microbial Culture Collection Center, located at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing. The preservation date was February 19, 2025, and the preservation number was CGMCC No. 32201.

[0042] Example 2: Study on the waterlogging tolerance mechanism of Vitis ficifolia Special No. 1

[0043] Artificial waterlogging treatment was carried out in the Three Gorges Botanical Garden. Potted Vitis ficifolia Special No. 1 was cultivated. In early October, three-year-old two-year-old cuttings of Vitis ficifolia Special No. 1 with the same size, good growth, and the same development period were selected, with every 3 pots as a group. Among them, 3 pots were selected for open-field cultivation management as the control group. The remaining pots were subjected to whole-plant complete submergence tests for 3d, 6d, 9d, 12d, 15d, and 30d in the pond respectively. After each group reached the corresponding waterlogging time, the grape pots were immediately removed from the pond.

[0044] 1. Detection of physiological and biochemical indexes

[0045] After different waterlogging time treatments, the contents of MDA, osmotic regulation factors (free proline, soluble sugar, soluble protein), and antioxidant enzymes (peroxidase, superoxide dismutase, catalase) in the roots of each Vitis ficifolia were detected. The measurement methods were as follows:

[0046] The content of malondialdehyde (MDA) was determined by the TBA spectrophotometric method. The basic principle is that malondialdehyde and TBA undergo a condensation reaction to produce a colored compound with a maximum absorption peak at 532nm. The amount of the colored compound produced is proportional to the MDA content. Therefore, the content of malondialdehyde can be indirectly determined by measuring the absorbance at 532nm.

[0047] The content of proline was determined by the acidic ninhydrin colorimetric method. The principle is based on the characteristic that free amino acids react with ninhydrin reagent to form a colored compound. During the reaction, amino acids react with acidic ninhydrin to form a blue-violet compound, which has the maximum light absorption at a wavelength of 520nm. The content of the compound represents the level of free proline content.

[0048] The soluble sugar content was determined by anthrone colorimetry, with 100 μg / mL sucrose solution as the standard solution, and six test tubes were added with 0 mL, 0.2 mL, 0.4 mL, 0.6 mL, 0.8 mL, and 1.0 mL of standard sucrose solution, and distilled water was added to make the volume 2 mL, and then 0.5 mL of anthrone-ethyl acetate and 5 mL of concentrated sulfuric acid were added in sequence. After sufficient shaking, the test tubes were immediately placed in a boiling water bath and heated for 1 min. After cooling to room temperature, the blank was used as a reference, and the OD value was measured at a wavelength of 630 nm. The standard curve of soluble sugar was drawn with the OD value as the ordinate and the concentration of each standard solution (μg / mL) as the abscissa. Then the soluble sugar content in the sample was calculated.

[0049] The soluble protein content was determined using the Coomassie Brilliant Blue method. Coomassie Brilliant Blue G-250 can bond with the hydrophobic region of the protein in an acidic environment, causing its maximum absorption peak to change from 465nm to 595nm, and its color development degree is positively correlated with the protein concentration.

[0050] The peroxidase activity was determined using the guaiacol method, which is based on the fact that peroxidase can oxidize guaiacol into a dark brown product under the action of hydrogen peroxide, and its absorption peak wavelength is 470nm. By measuring the change in absorbance (OD) at each moment, the size of POD activity can be obtained.

[0051] The activity of superoxide dismutase (SOD) was determined by the nitroblue tetrazolium (NBT) method. This method is based on the ability of SOD to scavenge superoxide anion free radicals (O 2- ), thereby inhibiting the photoreduction reaction of NBT. By measuring the reduction degree of NBT in the reaction system, the activity of SOD can be calculated.

[0052] The activity of catalase was determined by potassium permanganate titration. The basic principle of this method is that catalase decomposes hydrogen peroxide into oxygen, and potassium permanganate oxidizes hydrogen peroxide. Therefore, the activity of catalase can be measured by titrating potassium permanganate solution.

[0053] In this experiment, Excel was used to calculate the mean and standard deviation; SPSS27 was used for one-way analysis of variance, and when P<0.05, the difference was significant; origin2021 was used for drawing.

[0054] The results of the determination of the contents of MDA, free proline, soluble sugar, soluble protein, peroxidase, superoxide dismutase and catalase in the roots of each mulberry leaf grape after different flooding time treatments are as follows Figures 1 to 7 shown.

[0055] Malondialdehyde is the end product of cell membrane lipid peroxidation, and its content can reflect the degree of damage to plants under stress. In adversity, plants mainly use inorganic ions to resist adversity, but due to Na+ The enrichment of elements such as... leads to ionic toxicity in plants. Therefore, the content of malondialdehyde will increase. As can be seen from Figure 1 it, the content of malondialdehyde shows a trend of first increasing, then decreasing, and then increasing with the prolongation of waterlogging stress time, reaching the highest at 9 days of waterlogging treatment. At this time, the MDA content is 1.85 times that of the control (treatment for 0 days).

[0056] Proline, as an important substance to maintain the osmotic balance in plants, in the initial stage of waterlogging stress, in order to maintain the stability of cell structure, the content of free proline in plants may increase, thus increasing the water retention capacity of cells. As can be seen from Figure 2 it, the content of free proline increases significantly with the prolongation of waterlogging time, reaching the maximum at 12 days, which is 2.29 times that of the control group.

[0057] Under waterlogging conditions, to maintain the intracellular osmotic pressure balance and energy supply, the synthesis and accumulation of soluble sugars in grape roots may be affected. These soluble sugars can act as an osmotic adjustment substance, assisting cells to maintain the water balance in the body and avoid excessive water loss when flooded. At the same time, soluble sugars are also an important energy source for plants to cope with environmental stress, providing an important material basis for their survival and repair in adversity. As can be seen from Figure 3 it, with the increase of waterlogging time, the content of soluble sugars increases significantly, reaching the highest at 12 days, which is 4.71 times that of the control group; long-term waterlogging stress may lead to hypoxia in the roots of Vitis bryoniifolia, affecting the absorption and utilization of nutrients by the roots, and further restricting the synthesis and accumulation of soluble sugars. Therefore, the content of soluble sugars decreases at 12 - 30 days.

[0058] Soluble proteins have important physiological functions in plant cells, and their accumulation can increase the osmotic pressure of plants and reduce the damage of waterlogging stress to plants. As can be seen from Figure 4 it, the content of soluble proteins in the roots of Vitis bryoniifolia increases with the prolongation of waterlogging treatment time, reaching the highest at 15 days, which is 10.6 times that of the control; but after 15 days, the content of soluble proteins decreases significantly.

[0059] Peroxidase is a very valuable antioxidant enzyme in plants, and its main function is to protect cells from oxidative damage by hydrolyzing peroxides. As can be seen from Figure 5 it, after waterlogging treatment, the activity of peroxidase shows a change rule of first increasing and then decreasing, reaching the maximum at 15 days of waterlogging, which is 2.14 times that of the control; with the prolongation of waterlogging time, the activity of peroxidase decreases again at 30 days of waterlogging stress.

[0060] Superoxide dismutase is a key antioxidant enzyme, whose main function is to catalyze the dismutation reaction of superoxide free radicals, converting them into hydrogen peroxide (H2O2) and oxygen (O2), thereby protecting cells from oxidative damage. As can be seen from Figure 6 it, under waterlogging stress, the activity of superoxide dismutase in the roots of Vitis bryoniifolia Bunge first increased significantly and then decreased. Among them, the activity of superoxide dismutase reached the maximum value at 3 days of waterlogging, which was 1.19 times that of the control group, and the activity of superoxide dismutase continued to decline after 6 days.

[0061] Catalase is a class of antioxidant enzymes with important physiological activities. Its main role is to degrade hydrogen peroxide into water and oxygen to maintain the normal growth and metabolism of plants and reduce the oxidative damage of cells. As can be seen from Figure 7 it, the activity of catalase was relatively low at 0 days. When waterlogging stress was carried out, the activity of catalase first increased significantly and reached the highest at 3 days, which was 2.1 times that of the control group. After 3 days, with the extension of time, the activity of catalase gradually decreased.

[0062] From the above results, it can be seen that within a certain period of waterlogging treatment time, all physiological indicators showed an upward trend, but beyond this time, all physiological indicators showed a downward trend. This may be due to the fact that long-term waterlogging stress leads to metabolic imbalance in plants and they are unable to synthesize and accumulate organic substances normally. In addition, long-term waterlogging stress may also lead to changes in other metabolic pathways in plants, resulting in an increase in the decomposition and consumption of proline and soluble proteins. The above results indicate that the Vitis bryoniifolia Bunge specific No. 1 adapts to adversity by regulating the content of osmoregulatory substances and the activity of antioxidant enzymes under waterlogging stress, but its adaptability is limited, and long-term waterlogging stress will lead to the inhibition of its growth.

[0063] 2. Root tissue structure analysis

[0064] The plant morphology of Vitis bryoniifolia Bunge after different waterlogging time treatments was observed and photographed. At the same time, the thinnest surviving roots of each Vitis bryoniifolia Bunge plant were taken, and sections were made by the paraffin section method to observe and analyze the anatomical structure of the root tissues of Vitis bryoniifolia Bunge after different waterlogging time treatments. The specific section-making method is as follows:

[0065] (1) Sampling and fixation: Select the root tip of Vitis bryoniifolia Bunge (about 1 cm) as the section-making material, fix it with FAA (70% alcohol: formaldehyde: glacial acetic acid = 90:5:5) and remove the air, and place it for more than 24 hours.

[0066] (2) Softening: Put it into a mixed solution of glycerol and 95% alcohol at a ratio of 1:1 for 7 days.

[0067] (3) Dehydration and clearing: Immerse the sample successively in gradient ethanol solutions of 50%, 70%, 85%, 95%, 100%, and 100% for 1 hour each. Then, transfer it to a mixed solution of xylene: absolute alcohol = 1:1 and wait for 2 hours. Finally, immerse it in pure xylene for 2 hours.

[0068] (4) Impregnation with paraffin and embedding: Mix the sample with a small amount of xylene and paraffin, pour the mixture into molten paraffin, and place it in a constant-temperature oven at 60°C. Replace the paraffin completely every 3 hours for a total of 3 times. Then, use a paraffin embedding machine for embedding.

[0069] (5) Sectioning and mounting: Trim the paraffin block and place it on a microtome. Set the section thickness to 8 μm and cut sections. Float the sections on a water bath at 40°C and then transfer them to a constant-temperature drying oven at 40°C.

[0070] (6) Staining, coverslipping, and microscopic examination: Use safranin-fast green counterstaining for observation. Determine the staining time through preliminary experiments: 35 minutes for safranin and 30 seconds for fast green.

[0071] (7) Deparaffinization: Immerse the sample in pure xylene 1 for 5 minutes, then transfer it to pure xylene 2 for another 5 minutes, and finally transfer it to pure xylene 3 and wait for 5 minutes.

[0072] (8) Rehydration: Immerse the sample in absolute alcohol 1 for 5 minutes, take it out and transfer it to absolute alcohol 2. After 10 minutes, take it out and successively immerse it in alcohol solutions with volume fractions of 95%, 95%, 85%, and 70% for 3 minutes, 2 minutes, 3 minutes, and 3 minutes respectively. Finally, immerse it in filtered water for 2 minutes.

[0073] (9) Safranin staining: Immerse the sample in a 1% safranin solution for 35 minutes. Then, immerse it in filtered water for 5 minutes and then for 2 minutes.

[0074] (10) Dehydration: Immerse the sample successively in alcohol solutions with gradients of 70%, 85%, and 95% for 1 minute each.

[0075] (11) Fast green counterstaining: Immerse the sample in 1% fast green for 30 seconds.

[0076] (12) Dehydration: Immerse the sample successively in staining jars containing 95% alcohol and 95% alcohol for 1 minute each, and then in staining jars containing absolute alcohol and absolute alcohol for 2 minutes each.

[0077] (13) Clearing: Then, immerse the sample in two staining jars containing pure xylene for 2 minutes each.

[0078] (14) Coverslipping: Drop enough mounting medium on the paraffin section, cover the sample with a coverslip, and place it indoors to wait for the mounting medium to dry.

[0079] (15) Microscopic examination: Use a microscope to observe and analyze the material.

[0080] The morphological observation results of Vitis bryoniifolia Bunge plants after different flooding time treatments are as Figure 8 shown in and Table 1. It can be seen that the absorbing roots of the plants without flooding treatment are complete and dense, the root system color is yellow, and the leaves are normal ( Figure 8 in A). After 3 days of flooding stress, most of the secondary roots of the Vitis bryoniifolia Bunge plant root system survive, and the root system color is yellowish-brown; almost all the leaves on the branches are retained, and some leaves turn yellow ( Figure 8 in B). After 6 days of flooding stress, a small amount of the absorbing roots of Vitis bryoniifolia Bunge survive, most of them have blackened, and there are roots with a length of 1 mm and above surviving, and the root system shows yellow; most of the leaves on the branches are retained, and some are yellowed ( Figure 8 in C). After 9 days of flooding stress, all the absorbing roots of Vitis bryoniifolia Bunge have died and turned black, and some of the remaining secondary roots still survive; there are more remaining leaves on the branches ( Figure 8 in D). After 12 days of flooding, all the fine absorbing roots of Vitis bryoniifolia Bunge have died and turned black, and some of the remaining secondary roots have died; the lateral branches survive, the color is green, and there are sporadic remaining leaves on them ( Figure 8 in E and F). After 15 days of flooding treatment, all the absorbing roots of the Vitis bryoniifolia Bunge plant root system have died and turned black, only the secondary roots and the main roots survive, the color is yellowish-brown, and all the leaves on its branches wither and fall off ( Figure 8 in G). After 30 days of flooding treatment, all the absorbing roots of Vitis bryoniifolia Bunge have necrosed, and some remain on them; most of the secondary roots have died, only the main roots survive; all the lateral branches have died and fallen off, there are no remaining leaves, and the branches are smelly and sticky ( Figure 8 in H).

[0081] Table 1 Morphological changes of the stems, leaves and root systems of Vitis bryoniifolia Bunge under different flooding time treatments

[0082]

[0083] The microscopic structure of the root tip cross-section of Vitis bryoniifolia Bunge plants after different flooding time treatments is as Figure 9 shown. Among them, A is the microscopic structure of the root tip cross-section of Vitis bryoniifolia Bunge without flooding treatment, B is the microscopic structure of the root tip cross-section of Vitis bryoniifolia Bunge after 3 days of flooding treatment, C is the microscopic structure of the root tip cross-section of Vitis bryoniifolia Bunge after 6 days of flooding treatment, D is the microscopic structure of the root tip cross-section of Vitis bryoniifolia Bunge after 9 days of flooding treatment, E is the microscopic structure of the root tip cross-section of Vitis bryoniifolia Bunge after 12 days of flooding treatment, F is the microscopic structure of the root tip cross-section of Vitis bryoniifolia Bunge after 15 days of flooding treatment, and G and H are the microscopic structures of the root tip cross-section of Vitis bryoniifolia Bunge after 30 days of flooding treatment. The partial enlarged views of the microscopic structures of the root tip cross-sections of Vitis bryoniifolia Bunge after 3 days (A), 6 days (B) and 9 days (C) of flooding treatment are as Figure 10As shown in the figure, where a is the periderm, b is the phloem, c is the phloem ray, d is the xylem ray, e is the secondary xylem, f is the primary xylem, g is the suberin and lignin that have accumulated in large amounts, and h is the lysigenous aerenchyma formed due to the programmed death and dissolution of living cells.

[0084] It can be seen from Figure 9 and Figure 10 that the primary structure of the roots of normal Vitis bryoniifolia Bunge without flooding treatment includes the stele, endodermis, cortex, exodermis, lignified 'Φ'-shaped thickening, aerenchyma, and epidermis; the secondary structure is composed of secondary xylem, secondary phloem, aerenchyma, and cork cambium. After flooding stress, multiple cells in the cortex show 'Φ'-shaped lignified thickening, a large amount of suberin and lignin are deposited in the exodermis, a cork cambium is produced, and the aerenchyma is more developed. In Vitis bryoniifolia Bunge after 3 days of flooding stress, a small number of smaller voids are generated in the cortex of its roots. This is mainly due to the separation of cells from each other. When the flooding stress reaches 6 days, these voids become denser in the cortex. After 9 days of flooding stress, lysigenous air cavities produced due to cell programmed death or dissolution begin to appear in the cortex. When the flooding stress reaches 12 days, a large number of lysigenous air cavities appear in its cortex, and the voids generated previously also become wider. These changes are distributed on the cortex of the wild grape root system, making its parenchyma look wide. After 15 days and 30 days of flooding stress, basically no significant changes occur.

[0085] Example 3

[0086] A method for using the specific No. 1 of Vitis bryoniifolia Bunge to repair the water-level-fluctuating zone of the Three Gorges Reservoir area is as follows:

[0087] (1) Propagation of the specific No. 1 of Vitis bryoniifolia Bunge: The propagation is carried out in a nursery or a nutrient pot, and the propagation method is hardwood cutting or softwood full-light spray cutting;

[0088] Among them, the time for propagation using the hardwood cutting method is from February to May, from June to September, or from October to November (the branches used for propagation from June to September are refrigerated and preserved branches). When propagating, the one-year-old or perennial hardwood branches of the specific No. 1 of Vitis bryoniifolia Bunge are cut into cuttings with 2 - 3 buds. The morphological lower end of the cutting is cut obliquely, and the morphological upper end is cut flat. The morphological lower end of the cutting is inserted into the cutting substrate (yellow-brown soil) of the nursery or a non-woven fabric nutrient pot with a diameter of 12 - 30 cm, and the insertion depth into the cutting substrate is 1 / 3 - 1 / 2 of the cutting length.

[0089] The time for propagation using the above-mentioned softwood full-light spraying cutting method is from July to September. When propagating, the softwood branches of Morus australis Poir. cv. are cut into cuttings with 2 - 3 buds. The morphological upper end of the cuttings has leaves, and 1 / 3 - 1 / 2 of the leaf area is retained. The morphological lower end of the cuttings is inserted into the river sand bed, and watering management is carried out using a full-light spraying watering device. After 2 months, the rooted seedlings are transplanted into non-woven fabric nursery pots with a diameter of 12 - 30 cm for cultivation.

[0090] (2) Plant the two-year-old or older Morus australis Poir. cv. clone nursery pots (planted with nursery pots) propagated in step (1) in the water-level-fluctuating zone of the Three Gorges Reservoir Area. Plant after the water-level-fluctuating zone is exposed. The spacing during planting is 2.5 m × 4 m, and the length × width × depth of the planting hole is 25 cm × 25 cm × 30 cm. After planting, use the stones around the planting hole in the water-level-fluctuating zone to compact and fix the rootstock of the seedlings, cover the planting hole with a weed-proof cloth and fix the weed-proof cloth around to reduce the scouring of surging waves. Appropriate fertilization can be carried out in the first and second years of planting to promote growth. In the first year of planting, according to the growth of weeds such as Cynodon dactylon around, take the measure of mowing and tending around the planting hole to prevent weeds.

[0091] The effect picture of Morus australis Poir. cv. planted at an altitude of 159 meters in the water-level-fluctuating zone of the Three Gorges Reservoir Area is as Figures 11 to 13 shown. Through experiments, it is found that Morus australis Poir. cv. has the characteristics of good flood tolerance, high propagation rate, strong drought tolerance and strong adaptability. It can adapt to the environment of being completely flooded by water for 7 months during the dormant period, the survival rate of cutting propagation is more than 90%, it can adapt to the environment of drought and less rain, and has low requirements for soil fertility, and can grow in the barren soil of the water-level-fluctuating zone.

[0092] The above is only the preferred embodiment of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A method for repairing a water-fluctuation zone, characterized in that: Mulberry Leaf Grape Specific No. 1 is planted in the drawdown zone; the Mulberry Leaf Grape Specific No. 1 is preserved in the General Microbiology Center of China Microbiological Culture Collection Administration, the address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, the preservation date is: February 19, 2025, and the preservation number is: CGMCC No.32201.

2. The method according to claim 1, characterized in that The drawdown zone is the drawdown zone of the Three Gorges Reservoir area.

3. The method according to claim 1, characterized in that Before the planting, the mulberry leaf grape special No. 1 is propagated in a nursery or a nutrient pot, and the propagation method is a hard branch cutting method or a soft branch full light spray cutting method.

4. The method according to claim 3, characterized in that The time for propagation by using the hard branch cutting method is February to May, June to September or October to November, and the time for propagation by using the soft branch full light spray cutting method is July to September.

5. The method according to claim 4, characterized in that The hard branch cutting method comprises the following steps: cutting the hard branch branches of Mulberry Leaf Grape Specific No. 1 into cuttings with 2 to 3 buds, and inserting the morphological lower ends of the cuttings into a cutting medium of a nursery or a nutrient pot.

6. The method according to claim 5, characterized in that The hardwood branches are annual or perennial branches; The morphological lower end of the cutting is cut obliquely, and the morphological upper end is cut flatly; The nutrient pot is a non-woven nutrient pot, and the diameter of the non-woven nutrient pot is 12 to 30 cm; The depth of inserting into the cutting medium is 1 / 3 to 1 / 2 of the length of the cutting; the cutting medium is yellow-brown soil.

7. The method according to claim 4, characterized in that The method for tender branch full-light spray cuttings is as follows: after cutting tender branches of Mulberry Leaf Grape Specific No. 1 into cuttings with 2 to 3 buds, the morphological lower ends of the cuttings are inserted into a river sand bed, and full-light spray watering equipment is used for watering management. After 40 to 80 days, the rooted seedlings are transplanted into nutrient pots for cultivation.

8. The method according to claim 7, characterized in that The morphological upper end of the cutting has a leaf, and the leaf area is 1 / 3 to 1 / 2; The nutrient pot is a non-woven nutrient pot, and the diameter of the non-woven nutrient pot is 12 to 30 cm.

9. The method according to claim 1, characterized in that During the planting, two-year-old or more mulberry leaf grape specific No. 1 asexual seedlings are selected in nutrient pots; the spacing during the planting is 2-3m×3-5m; the planting hole length×width×depth is 15-35cm×15-35cm×25-35cm.

10. The method according to any one of claims 1 to 9, characterized in that: After planting Mulberry Leaf Grape Special No. 1, compact and fix the roots of the seedlings, cover the planting holes with weed-proof cloth and fix the weed-proof cloth around them.

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