Application of glycine in low temperature stress resistance of apple tree

Abstract

The application relates to the field of fruit tree cultivation technology, and particularly relates to application of glycine in resisting low-temperature stress of apple trees, and the resistance of the apple trees to the low-temperature stress is improved by applying a glycine aqueous solution to the apple tree seedlings. The application improves the biomass of the apple trees under the low-temperature stress, improves the photosynthetic pigment content of the apple tree leaves under the low-temperature stress, promotes the growth and development of the root system of the apple tree seedlings under the low-temperature stress, enhances the enzyme activity related to nitrogen metabolism, and improves the expression of the genes related to nitrogen absorption and assimilation, so that the resistance of the apple trees to the low-temperature stress is improved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of fruit tree cultivation, and particularly relates to application of glycine in resisting low-temperature stress of apple trees. BACKGROUND

[0002] Low-temperature stress is one of the main hazards affecting the development of fruit trees in cold northern regions. After being subjected to low-temperature stress for a certain period, the physiological and biochemical indexes and phenotypic characteristics in the body of a plant will change to a certain extent. Typical symptoms are a decrease in growth rate, which further leads to disorder of normal physiological metabolism, so that the most basic life activities of the plant cannot be guaranteed, and the plant may even die. Low-temperature stress is divided into cold damage and freezing damage. The temperature range of cold damage is generally 0-5°C, and the temperature range of freezing damage is generally less than 0°C. It has been found that after being subjected to low-temperature damage, the growth rate of a plant, such as plant height and leaf area, decreases. Sun et al. found that low temperature can cause cell function disorder, and with the change of growth and development stages, the response of each tissue to low temperature will also be different. At the same time, low temperature can also inhibit the growth of the root system of a plant. The root system is an organ for absorbing water and nutrients, and is crucial in the development processes of fruit trees, such as vegetative growth and reproductive growth. It has been found that low temperature in the root zone of a fruit tree can cause the growth of the root system of an apple tree to slow down, which is manifested in a decrease in total root length, root surface area and volume, so that the ability of the root system to absorb various mineral nutrients is inhibited, and the growth and development of seedlings are affected.

[0003] Photosynthesis is the basis for the generation of organic matter and the material and energy for the survival of a plant. Low temperature can directly cause damage to photosynthetic structures (such as chloroplasts), so that the photosynthetic intensity of a plant decreases. Under low-temperature conditions, the synthesis of chlorophyll and the photosynthetic rate of a plant are both significantly inhibited. The decrease in chlorophyll content caused by low temperature is caused by two factors: on the one hand, the enzyme activity for the synthesis of chloroplast pigments decreases, so that the synthesis of chloroplasts is inhibited; on the other hand, the structure of chloroplasts in leaf cells is destroyed under low-temperature conditions, which causes chlorophyll to degrade, and thus affects the capture, conversion and distribution of light energy, causes the photosynthetic electron transport to be blocked, the photosynthetic mechanism to be damaged, and carbon assimilation to be inhibited, and finally causes the photosynthetic capacity of leaves to decrease. Low temperature destroys the structure of the photosystem, decreases Pn and chlorophyll content, and has a significant inhibitory effect on photosynthesis of a plant.

[0004] Plant nutrient absorption is affected by environmental temperature. Low temperature can inhibit plant nitrogen absorption, transformation and utilization. The basic morphological structure of plant root system can be affected by low temperature, and the mineral nutrient absorption capacity of root system is weakened, so that the aboveground growth is short. Low temperature can inhibit the absorption of nitrogen, phosphorus and potassium elements by root system. The inhibition of nitrogen absorption is greater than that of other elements. The effect of low temperature on plant nitrogen absorption is also related to the duration of low temperature. Short-term low temperature can inhibit the nitrogen absorption of sensitive varieties. Under long-term low temperature conditions, the nitrogen accumulation of both resistant and weak varieties is significantly reduced. Low temperature can also inhibit the activity of enzymes related to nitrogen absorption and metabolism, thereby affecting the nitrogen metabolism process of plants and causing influence on plant growth.

[0005] Apple is a perennial deciduous fruit tree, and the demand for nitrogen is large. However, the low utilization rate of nitrogen is an important problem in current apple production, and is also one of the main factors affecting the yield and quality of apples. Low temperature is the main environmental stress factor in early spring and late autumn in apple production areas in northern China. Spring low temperature can affect soil nitrogen mineralization and reduce the nitrogen absorption of fruit tree root system. Low temperature can also directly affect the nutrient concentration of tree body by inhibiting the absorption and transportation of plant root system, thereby affecting the early spring organ construction of apple tree, showing that the leaf development is slow and the fruit setting rate is low, and further affecting the yield and quality of apples. Therefore, it is urgent to solve the problem of relieving the low temperature stress of apple. SUMMARY

[0006] In order to relieve the low temperature stress of apple, the application provides the application of glycine in resisting low temperature stress of apple tree. The application can improve the biomass of apple tree under low temperature stress, improve the photosynthetic pigment content of apple tree leaf under low temperature stress, improve the photosynthetic parameter of apple tree leaf under low temperature stress, promote the growth and development of root system of apple tree seedling under low temperature stress, enhance the activity of nitrogen metabolism related enzyme and improve the expression of nitrogen absorption and assimilation related gene, so as to improve the resistance of apple tree to low temperature stress.

[0007] The application provides the application of glycine in resisting low temperature stress of apple tree. The resistance of apple tree to low temperature stress is improved by applying glycine aqueous solution to apple tree seedling. The low temperature stress condition of the application is that the day is 8-10 DEG C and the night is 0-2 DEG C.

[0008] The application can improve the biomass of apple tree under low temperature stress, improve the photosynthetic pigment content of apple tree leaf under low temperature stress, improve the photosynthetic parameter of apple tree leaf under low temperature stress, promote the growth and development of root system of apple tree seedling under low temperature stress, enhance the activity of nitrogen metabolism related enzyme and improve the expression of nitrogen absorption and assimilation related gene, so as to improve the resistance of apple tree to low temperature stress.

[0009] Compared with the prior art, the application has the beneficial effects that:

[0010] The application improves the resistance of apple trees to low temperature stress in the following ways:

[0011] The biomass of the apple trees under low temperature stress, such as the above-ground biomass of seedlings, total biomass and crown root ratio, is improved; the photosynthetic pigment content of the leaves of the apple trees under low temperature stress, such as the chlorophyll a content, chlorophyll b content and carotenoid content, is improved; the photosynthetic parameters of the leaves of the apple trees under low temperature stress, such as the net photosynthetic rate, transpiration rate, stomatal conductance and water use efficiency, are improved; the growth and development of the root system of the seedlings of the apple trees under low temperature stress is promoted, such as the total root length, total surface area of the seedlings of the apple trees under low temperature stress is increased and the root system activity of the seedlings is improved; the activities of nitrogen metabolism related enzymes, such as the nitrate reductase activity, glutamine synthetase activity, glutamate synthetase activity and glutamate dehydrogenase activity, are improved; and the expression of nitrogen absorption and assimilation related genes NRT1.1, NRT2.4, NRT2.7, AMT1.2, AMT1.5 and NADH-GOGAT is improved. BRIEF DESCRIPTION OF DRAWINGS

[0012] In order to more clearly illustrate the technical solutions of the embodiments of the present application or the prior art, the following will briefly introduce the drawings needed to be used in the embodiments or prior art description. Obviously, the drawings in the following description only show some embodiments of the present application, and other drawings can also be obtained by those skilled in the art without any creative effort based on these drawings.

[0013] Figure 1 Figure 1 is the effect of glycine treatment on the growth of seedlings of Shan Dingzuo under low temperature; in the figure, A is before treatment; B is after 12 days of treatment.

[0014] Figure 2 Figure 2 is the effect of glycine on the photosynthetic pigment content of the leaves of Shan Dingzuo under low temperature; in the figure, A is the effect of glycine on the chlorophyll a content of the leaves of Shan Dingzuo under low temperature; B is the effect of glycine on the chlorophyll b content of the leaves of Shan Dingzuo under low temperature; C is the effect of glycine on the carotenoid content of the leaves of Shan Dingzuo under low temperature; D is the effect of glycine on the total chlorophyll content of the leaves of Shan Dingzuo under low temperature.

[0015] Figure 3 Figure 3 is the effect of glycine on the fluorescence parameters of the leaves of seedlings of Shan Dingzuo under low temperature; in the figure, A is the effect of glycine on the initial fluorescence of the leaves of Shan Dingzuo under low temperature; B is the effect of glycine on the potential activity of PSII of the leaves of Shan Dingzuo under low temperature; C is the effect of glycine on the maximum photochemical efficiency of PSII of the leaves of Shan Dingzuo under low temperature; D is the effect of glycine on the photochemical performance index of the leaves of Shan Dingzuo under low temperature; E is the effect of glycine on the light energy absorbed per unit area of the leaves of Shan Dingzuo under low temperature; F is the effect of glycine on the quantum yield of electron transport per unit area of the leaves of Shan Dingzuo under low temperature.

[0016] Figure 4Effect of glycine treatment on root morphology of P. taxifolia seedlings under low temperature.

[0017] Figure 5 Effect of glycine treatment on root activity of P. taxifolia under low temperature.

[0018] Figure 6 Effect of glycine treatment on nitrate nitrogen, nitrite nitrogen and ammonium nitrogen content in roots and leaves of P. taxifolia under low temperature; in the figure, A is the effect of glycine treatment on nitrate nitrogen in roots of P. taxifolia under low temperature; B is the effect of glycine treatment on nitrite nitrogen in roots of P. taxifolia under low temperature; C is the effect of glycine treatment on ammonium nitrogen in roots of P. taxifolia under low temperature; D is the effect of glycine on nitrate nitrogen in leaves of P. taxifolia seedlings; E is the effect of glycine on nitrite nitrogen in leaves of P. taxifolia seedlings; F is the effect of glycine on ammonium nitrogen in leaves of P. taxifolia seedlings.

[0019] Figure 7 Effect of glycine treatment on total nitrogen content in roots and leaves of P. taxifolia under low temperature; in the figure, A is the effect of glycine treatment on total nitrogen content in roots of P. taxifolia under low temperature; B is the effect of glycine treatment on total nitrogen content in leaves of P. taxifolia under low temperature.

[0020] Figure 8 Effect of glycine treatment on nitrogen assimilation enzyme activity in roots of P. taxifolia under low temperature; in the figure, A is the effect of glycine treatment on nitrate reductase activity in roots of P. taxifolia under low temperature; B is the effect of glycine treatment on glutamine synthetase activity in roots of P. taxifolia under low temperature; C is the effect of glycine treatment on glutamate synthetase activity in roots of P. taxifolia under low temperature; D is the effect of glycine treatment on glutamate dehydrogenase activity in roots of P. taxifolia under low temperature.

[0021] Figure 9 Effect of glycine treatment on nitrogen assimilation enzyme activity in leaves of P. taxifolia under low temperature; in the figure, A is the effect of glycine treatment on nitrate reductase activity in leaves of P. taxifolia under low temperature; B is the effect of glycine treatment on glutamine synthetase activity in leaves of P. taxifolia under low temperature; C is the effect of glycine treatment on glutamate synthetase activity in leaves of P. taxifolia under low temperature; D is the effect of glycine treatment on glutamate dehydrogenase activity in leaves of P. taxifolia under low temperature.

[0022] Figure 10 Effect of glycine treatment on glutamate-oxaloacetate transaminase (GOT) and glutamate-pyruvate transaminase (GPT) activities in roots and leaves of P. taxifolia under low temperature; in the figure, A is the effect of glycine treatment on glutamate-oxaloacetate transaminase activity in roots of P. taxifolia under low temperature; B is the effect of glycine treatment on glutamate-pyruvate transaminase activity in roots of P. taxifolia under low temperature; C is the effect of glycine treatment on glutamate-oxaloacetate transaminase activity in leaves of P. taxifolia under low temperature; D is the effect of glycine treatment on glutamate-pyruvate transaminase activity in leaves of P. taxifolia under low temperature.

[0023] Figure 11 Effects of glycine treatment on the composition and content of free amino acids in the roots of S. rosthomii under low temperature; in the figure, A is the effect of glycine treatment on the content of alanine in the roots of S. rosthomii under low temperature; B is the effect of glycine treatment on the content of methionine in the roots of S. rosthomii under low temperature; C is the effect of glycine treatment on the content of glycine in the roots of S. rosthomii under low temperature; D is the effect of glycine treatment on the content of arginine in the roots of S. rosthomii under low temperature; E is the effect of glycine treatment on the content of tyrosine in the roots of S. rosthomii under low temperature; F is the effect of glycine treatment on the content of proline in the roots of S. rosthomii under low temperature; G is the effect of glycine treatment on the content of threonine in the roots of S. rosthomii under low temperature; H is the effect of glycine treatment on the content of histidine in the roots of S. rosthomii under low temperature; I is the effect of glycine treatment on the content of glutamic acid in the roots of S. rosthomii under low temperature; J is the effect of glycine treatment on the content of glutamine in the roots of S. rosthomii under low temperature; K is the effect of glycine treatment on the content of aspartic acid in the roots of S. rosthomii under low temperature.

[0024] Figure 12 Effects of glycine treatment on the expression amount of genes related to nitrogen absorption and assimilation in the roots of S. rosthomii under low temperature.

[0025] Figure 13 Effects of glycine treatment on the content of soil nutrients under low temperature; in the figure, A is the effect of glycine treatment on the content of available nitrogen in the soil under low temperature; B is the effect of glycine treatment on the content of available phosphorus in the soil under low temperature; C is the effect of glycine treatment on the content of available potassium in the soil under low temperature; D is the effect of glycine treatment on the content of organic matter in the soil under low temperature.

[0026] Figure 14 Effects of glycine treatment on the content of different forms of nitrogen in the soil under low temperature; in the figure, A is the effect of glycine treatment on the content of nitrate nitrogen in the soil under low temperature; B is the effect of glycine treatment on the content of ammonium nitrogen in the soil under low temperature; C is the effect of glycine treatment on the content of microbial biomass nitrogen in the soil under low temperature.

[0027] Figure 15 Effects of glycine treatment on the composition of organic nitrogen in the soil under low temperature; in the figure, A is the effect of glycine treatment on the content of total acid-soluble nitrogen in the soil under low temperature; B is the effect of glycine treatment on the content of amino acid nitrogen in the soil under low temperature; C is the effect of glycine treatment on the content of amino acid nitrogen in the soil under low temperature; D is the effect of glycine treatment on the content of amino sugar nitrogen in the soil under low temperature.

[0028] Figure 16Effects of glycine treatment on soil enzyme activities under low temperature; in the figure, A is the effect of glycine treatment on soil urease activity under low temperature; B is the effect of glycine treatment on soil asparaginase activity under low temperature; C is the effect of glycine treatment on soil glutamine activity under low temperature; D is the effect of glycine treatment on soil nitrate reductase activity under low temperature. DETAILED DESCRIPTION

[0029] The specific embodiments of the present application are described in detail below, but it should be understood that the scope of protection of the present application is not limited by the specific embodiments. Based on the examples in the present application, all other examples obtained by those of ordinary skill in the art without creative labor are within the scope of protection of the present application. The experimental methods described in the embodiments of the present application are conventional methods, and the materials and reagents used in the following examples are commercially available unless otherwise specified.

[0030] Example 1: Application of glycine in resisting low temperature stress of apple trees.

[0031] I. Experimental materials and methods

[0032] 1. Experimental materials and treatment

[0033] One-year-old Malus baccata Borkh. seedlings were used as test materials. The seeds were sown after being stratified and sprouted. After the seedlings grew to about 4-5 true leaves, they were transplanted into nutrient pots containing mixed substrate (orchard soil, nursery substrate and river sand mixed at a ratio of 2:1:1). One seedling was planted in each nutrient pot. The seedlings were watered appropriately and timely every day, and the prevention of diseases and pests was paid attention to. Before leaf fall in winter, all the test materials were moved to the cold-proof shed for wintering. In the following spring in April, when the seedling buds began to sprout, the seedlings that were uniform in growth, free of diseases and pests, and healthy were selected and placed in two different temperature artificial climate chambers for cultivation. The cultivation temperatures were normal temperature: 24℃ (day) / 14℃ (night) and low temperature: 10℃ (day) / 2℃ (night). The light intensity was set to 400 μmol·m -2 ·s -1 , and the average relative humidity was 75%. After the seedlings were pre-cultured for 1 day, the test treatment was started.

[0034] Three treatments were set in the test, which were: (1) control (CK): normal temperature + distilled water; (2) low temperature (L): low temperature + distilled water; (3) low temperature + glycine (LG): low temperature + 6mM glycine. The low temperature stress treatment temperature was 10℃ (day) / 2℃ (night), and the seedlings under 24℃ (day) / 14℃ (night) were used as the control.

[0035] Treatment group LG was irrigated with 200 mL of glycine solution per pot, and the control group was irrigated with an equal amount of distilled water per pot. Samples were taken at 3 d, 6 d, 9 d, and 12 d after treatment, with 5 plants per repetition, 3 biological repetitions, and a total of 15 plants per treatment. When sampling, the rhizosphere soil was first collected in a plastic bag using the root shaking method for soil testing. Then the plants were washed clean with water, and root and leaf samples were collected, wrapped in tin foil, and labeled clearly for each treatment, and frozen in liquid nitrogen. On the 12th day of sampling, biomass was measured, with 5 plants per treatment and 3 repetitions.

[0036] 2. Biomass measurement

[0037] On the 12th day of treatment, 5 plants were taken from each treatment, washed with clean water, and the surface water was absorbed with absorbent paper. The seedlings were cut off at the rhizome with scissors and weighed on a balance, which was recorded as fresh weight at this time. Then the weighed tissue materials were placed in an oven, killed at 121°C for 20 min, dried at 60°C to constant weight, and cooled before weighing on an electronic balance to measure the dry weight of each part. The ratio of aboveground and underground dry weight represents the root-shoot ratio.

[0038] 3. Morphological index determination

[0039] Plant height and stem diameter of S. wilsoniana: measured on the 12th day of treatment using a tape measure and vernier caliper (accuracy 0.02).

[0040] Root morphological index: on the 12th day of sampling, the root morphological index of S. wilsoniana was quantitatively analyzed by digital scanner and WinRhizo PRO2016 analysis software.

[0041] 4. Measurement of seedling photosynthetic parameters

[0042] Before sampling on the 12th day of treatment, 3 mature leaves (LPI = 7) were selected from each S. wilsoniana seedling, and the photosynthetic parameters of each treatment were measured and analyzed using a CIRAS-2 photosynthetic instrument (PP Systems, USA).

[0043] 5. Measurement of chlorophyll content

[0044] The method for measuring chlorophyll content in seedling leaves is as follows: After taking the leaves of S. wilsoniana, 40 mg was weighed into a centrifuge tube, 0.25 mL of pure acetone and 8.75 mL of 80% acetone were added to each tube, the lid was closed and shaken vigorously once, then soaked overnight in a dark room at room temperature. To speed up the extraction effect, shake four to five times during this period until the sample is completely white. The next day, add 1 mL of 80% acetone and dilute to volume, and measure the wavelengths at 470 nm, 646 nm and 663 nm using a UV spectrophotometer and calculate.

[0045] Calculation formula: C Chla = 12.21 x OD 663 - 2.81 x OD 646 ; wherein C Chla represents the chlorophyll a content; OD 663 represents the wavelength of the ultraviolet spectrophotometer at 663 nm; OD 646 represents the wavelength of the ultraviolet spectrophotometer at 646 nm.

[0046] C Chlb = 20.13 x OD 646 - 5.03 x OD 663 ; C Chlb represents the chlorophyll b content; OD 663 represents the wavelength of the ultraviolet spectrophotometer at 663 nm; OD 646 represents the wavelength of the ultraviolet spectrophotometer at 646 nm.

[0047] C Chl(a+b) = Chla + Chlb; wherein C Chl(a+b) represents the total chlorophyll content; Chla represents the chlorophyll a content; Chlb represents the chlorophyll b content.

[0048] C Car = (1000 x OD 470 - 3.27 x Ca - 104 x Cb) / 229; wherein C Car represents the carotenoid content; OD 470 represents the wavelength of the ultraviolet spectrophotometer at 646 nm; Ca represents the chlorophyll a content; Cb represents the chlorophyll b content.

[0049] Chlorophyll pigment content Wherein C: pigment concentration; V: volume of extraction liquid; m: sample mass.

[0050] 6. Determination of chlorophyll fluorescence parameters

[0051] After dark adaptation for 20 min, the basic fluorescence parameters of the leaves of the mountain ardisia were determined using a Pocket PEA (Hansatech, UK) fluorometer.

[0052] 7. Determination of root activity

[0053] After treatment of the test material, the root activity of the mountain ardisia seedlings was determined on the 12th day using the triphenyltetrazolium chloride reduction method (TTC method). The operation is as follows: the white tender roots were cut into small pieces of 2 cm long and weighed about 0.5 g, 1 / 15 mol / L -1Phosphate buffer and TTC, 37℃ constant temperature for 4h, after adding 1M H2SO4 to terminate the reaction and placed for 15min, the roots were removed and the surface attached liquid was absorbed, then 10mL of ethanol was added, the roots were extracted at 25℃ for 24h, and the absorbance value at 485nm was measured by UV spectrophotometer.

[0054] 8. Determination of nitrogen content in different forms of plants

[0055] Determination of nitrate nitrogen (NO3 - -N) content used salicylic acid-sulfuric acid method recorded in Distinct signalling pathways and transcriptome response signatures differentiate ammonium-and nitrate-supplied plants by Patterson et al.

[0056] Determination of nitrite nitrogen (NO2 - -N) content used α-naphthylamine method recorded in Relationships between nitrite reductase activity and genotype-dependent callus growth in rice cell cultures by Ogawa et al.

[0057] Determination of ammonium nitrogen (NH4 + -N) content used ninhydrin method recorded in High-throughput colorimetric method for the parallel assay of glyoxylic acid and ammonium in a single extract by Zheng et al.

[0058] 9. Determination of key enzyme activity in plant nitrogen metabolism

[0059] 1.4mL 0.1mmol·L -1 KNO3 and 0.2mL NADH were added to 0.4mL enzyme extract, mixed thoroughly, and then color developing agent was added immediately after incubation at room temperature for 0.5h. After centrifugation, the absorbance value at 540nm was measured. NADH-GOGAT activity was determined according to the method recorded in Rapid determination of glutamate synthase activity by Zheng Chaofeng et al. - 1 Tris-HCl (pH=7.5), 0.1mmol·L -1 ​NADH, 10 mmol·L -1 Alpha-ketoglutarate and 10 mmol·L -1 Glutamine buffer, start reaction, continuously measure 340 nm absorbance value for 10 times, take a section of density stable reduction value to measure enzyme activity. According to the method recorded in Loulakakis et al. in Roubelakis-Angelakis K A. Intracellular localization and properties of NADH-glutamate dehydrogenase from Vitis vinifera L.: purification and characterization of the major leaf isoenzyme, glutamate dehydrogenase (GDH) activity was measured. According to the method recorded in Wu Lianghuan et al. in Research Progress of Amino Acid Nitrogen Absorption and Utilization in Higher Plants, GOT and GPT activities in grape root and leaf were measured. After taking the crude enzyme solution and enzyme substrate, it was placed in a 37℃ water bath for 1.5h, then 2,4-dinitrophenylhydrazine was added, and it was water bathed for 20min again. Finally, 0.4mol·L -1 NaOH was fully mixed, and the absorbance value at 500nm was measured after standing for 10min and adjusting to zero with distilled water.

[0060] 10. Determination of free amino acid content in plants

[0061] 0.3g of fresh plant tissue sample was weighed, 1mL of mixed solution (deionized water: chloroform: methanol = 3:5:12, v / v / v) was added, and it was fully shaken (4℃, 30min), then centrifuged (4℃, 12000xg, 15min), and the supernatant was collected in a new centrifuge tube. 375μL of chloroform and 625μL of deionized water were added to wash away the impurities in the solution, and the supernatant was blown to near dryness with a low-temperature rotary evaporator, then 1mL of acetonitrile was added, and it was filtered through an organic filter membrane (0.22μm) and then loaded onto the machine. The mobile phase I was ammonium acetate and acetonitrile, and the mobile phase II was ammonium acetate and formic acid mixed aqueous solution. The flow rate of the mobile phase was set to 0.3μL·min -1 , and the injection volume was 5μL.

[0062] 11. Determination of gene expression

[0063] The CTAB method is used to extract RNA from plant roots, and the specific process is as follows: take 150 mg of plant tissue sample, add 700 μL of CTAB (65°C preheating, make the turbid viscous reagent transparent) and 50 μL of β-mercaptoethanol into the tube, and the process is carried out in a fume hood. Mix well and place in a 65°C water bath for 30 min, during which time the centrifuge tube is inverted and mixed several times. After the water bath is over, add an equal volume of chloroform:isopropyl alcohol (volume ratio 24:1) mixture, mix thoroughly, and then centrifuge. Recover the supernatant for the second extraction. Take a new centrifuge tube, add the supernatant and 50 μL of lithium chloride, and place it in a -20°C refrigerator overnight. After standing, centrifuge at 4°C to retain the precipitate, add 75% ethanol to remove impurities, and centrifuge again. After washing the precipitate with anhydrous ethanol once and centrifuging, place it in a clean bench, dry it by blowing, add DEPC water, and store the RNA. The cDNA synthesis is performed using a reverse transcription kit (Takara, Japan) according to the instructions. The synthesized cDNA is stored in a -20°C refrigerator for future use. Use a 384-well plate for real-time fluorescence quantitative (RT-qPCR) amplification, and the primers are synthesized by Shenguo Bioengineering (Shanghai) Co., Ltd. The forward and reverse primers are shown in Table 1. Dilute the cDNA 10 times and perform qRT-PCR. Use 2 -ΔΔCt Method for calculating the relative expression of genes.

[0064] Table 1 qPT-PCR primer sequences

[0065]

[0066] 12. Determination of soil nutrient indicators

[0067] The determination of soil organic matter, available nitrogen, phosphorus and potassium content refers to the method recorded by Bao Shidang in "Nitrogen Utilization in Plants".

[0068] 13. Determination of different forms of nitrogen content in soil

[0069] The content of soil microbial biomass nitrogen (SMBN) is determined by the chloroform fumigation-K2SO4 extraction method recorded by Lin Xianggui in "Principles and Methods of Soil Microbial Research".

[0070] The determination method of NO3 - content in soil is as follows: accurately weigh 10 g of air-dried soil passing through a 20-mesh sieve, add 0.01 mol·L -1 of calcium chloride (CaCl2) extraction solution, and shake well. Shake for 30 min and then filter. Immediately after the filtrate is taken, add 1 mL of sulfuric acid solution (1:9) to acidify the test solution, and shake well. Adjust the zero with the acidified CaCl2 extraction solution, and measure the absorbance values at 210 nm and 275 nm.

[0071] The determination method of NH4 +-N content determination method: take 5 g of fresh soil sample passed through a 20 mesh sieve, accurately add 2 mol L -1 KCl) solution 25 mL, shake well, and oscillate for 1 h. After clarification, filter the upper clear liquid of the suspension, which is the soil leachate to be tested. Add phenol solution, 2 mol L -1 KCl and sodium hypochlorite alkaline solution 5 mL each at 27°C for 1 h, add masking agent (400 g L -1 Potassium sodium tartrate, 100 g L -1 EDTA disodium salt, 10 mol L -1 NaOH) after dissolving the precipitate produced, determine the absorbance at 625 nm.

[0072] 14. Determination of soil organic nitrogen components

[0073] The determination of soil organic nitrogen components refers to the method recorded by Lu Rukun in Soil Agricultural Chemistry Analysis Method.

[0074] 15. Determination of soil enzyme activity

[0075] The soil urease activity is determined by the phenol sodium-sodium hypochlorite colorimetric method with slight modifications. Specifically, take 10 g of sample, add 1 mL of toluene, shake well, and react for 15 min. Then add citric acid buffer with pH 6.7 and urea, mix well, and incubate at 37°C for 1 d. The next day, add distilled water, 1.53 mol L -1 Mix the phenol sodium and sodium hypochlorite solution well, shake well until the solution turns indigo blue, and determine the absorbance at 578 nm within 1 h.

[0076] The activities of soil nitrate reductase (NR), soil glutamine synthetase (GLNS), and soil asparagine (ASP) are determined according to the instructions of the corresponding kit (Jiangsu Enzyme Free Industry Co., Ltd.). The determination wavelength is 450 nm. The standard curve is based on the instructions provided by the kit.

[0077] 16. Data statistics and analysis

[0078] Microsoft Excel is used for data entry and arrangement, and SPSS 27.0 software is used for data statistical analysis. Single factor and least significant difference method (LSD method) are used for significance test (p < 0.05). Origin 2023 is used for plotting. The data in the chart are represented as mean ± standard error of three repeated tests.

[0079] II. Experimental results

[0080] 1. Effect of exogenous glycine on the growth of Shandingsu seedlings at low temperature

[0081] (1) Effect of glycine treatment on the biomass of P. montana seedlings under low temperature

[0082] As shown in Table 2, compared with CK, the normal growth of P. montana seedlings was significantly inhibited under low temperature treatment for 12 days, and the plant height, aboveground biomass, root biomass, total biomass and crown / root ratio of P. montana seedlings were reduced by 16.5%, 37.9%, 22.5%, 32.9% and 20.7% respectively, and the differences were significant. Compared with L treatment, the aboveground biomass, total biomass and crown / root ratio of seedlings under LG treatment increased by 28.0%, 2.6% and 21.5% respectively, and the differences were significant. It was shown that exogenous application of glycine could effectively alleviate the inhibition of low temperature on the growth of P. montana seedlings Figure 1 ).

[0083] Table 2 Effect of glycine on the growth parameters of P. montana seedlings under low temperature

[0084] Treatment Plant height (cm) Stem diameter (mm) Shoot biomass (g DW) Root biomass (g DW) Total biomass (g DW) Shoot to root ratio CK 46.27±1.21a 4.62±0.07a 5.23±0.15a 2.80±0.09a 8.03±0.13a 1.88±0.1a L 38.63±0.72b 4.58±0.05a 3.22±0.13c 2.17±0.08b 5.39±0.10c 1.49±0.11b LG 39.93±0.78b 4.61±0.09a 4.12±0.07b 2.30±0.15b 6.42±0.21b 1.81±0.09a

[0085] Note: Lowercase letters represent the significant difference of the same column among different treatments, p<0.05. CK: control, normal temperature + distilled water; L: low temperature treatment; LG: low temperature + glycine treatment.

[0086] 2、Effect of glycine treatment on the photosynthetic and fluorescence characteristics of P. montana leaves under low temperature

[0087] (1) Effect of glycine treatment on the photosynthetic pigment content of P. montana leaves under low temperature

[0088] As shown in Table 2, compared with CK, the normal growth of P. montana seedlings was significantly inhibited under low temperature treatment for 12 days, and the plant height, aboveground biomass, root biomass, total biomass and crown / root ratio of P. montana seedlings were reduced by 16.5%, 37.9%, 22.5%, 32.9% and 20.7% respectively, and the differences were significant. Compared with L treatment, the aboveground biomass, total biomass and crown / root ratio of seedlings under LG treatment increased by 28.0%, 2.6% and 21.5% respectively, and the differences were significant. It was shown that exogenous application of glycine could effectively alleviate the inhibition of low temperature on the growth of P. montana seedlings Figure 2

[0089] (2) Effect of glycine treatment on the photosynthetic parameters of P. montana leaves under low temperature

[0090] Table 3 Effect of different treatments on the photosynthetic parameters of P. montana seedlings

[0091]

[0092] As shown in Table 3, low temperature treatment significantly inhibited the Pn, Tr, Gs and WUE of P. montana seedling leaves, which were reduced by 48.0%, 31.8%, 23.7% and 15.1% respectively compared with CK. Compared with L treatment, the Pn, Tr, Gs and WUE of P. montana seedling leaves under LG treatment increased by 40.1%, 11.3%, 14.9% and 15.3% respectively.

[0093] (3) Effect of glycine treatment on the fluorescence parameters of P. montana leaves under low temperature​

[0094] Depend on Figure 3 As shown in A, the F0 value of leaves of *Stachys chinensis* seedlings treated with L increased by 16.3% after 12 days, a significant difference. Compared with the L treatment, the F0 value of the LG treatment decreased by 7.2%, with no significant difference.

[0095] Fv / F0 reflects the potential activity of PSⅡ. Figure 3 As shown in B, the Fv / F0 ratio of leaves in 12-day-old *Sedum morganianum* seedlings treated with L was significantly reduced by 25.9%. Compared with the L treatment, the Fv / F0 ratio in the LG treatment was significantly increased by 33.0%.

[0096] Fv / Fm, to some extent, represents the light energy conversion efficiency within the PSⅡ reaction center. Figure 3 As can be seen from the C values, low temperature significantly reduced the Fv / Fm ratio in the leaves of *Symplocos edulis* seedlings, decreasing by 28.4%. Compared with the L treatment, the Fv / Fm ratio increased by 25.2% under the LG treatment, showing a significant difference.

[0097] The photosynthetic performance index (PI abs) reflects the overall photosynthetic capacity of a plant. (The rest of the text appears to be a fragment and requires further context for accurate translation.) Figure 3 As can be seen from the D curve, PIabs were significantly reduced under the L treatment, decreasing by 37.6% compared to the CK. In contrast, PIabs in the leaves were significantly increased by 66.3% under the LG treatment.

[0098] ABS / CSm represents the light energy absorbed per unit area, while ETo / CSm represents the quantum yield of electron transfer per unit area. Figure 3 As shown in E and F of 3, the ABS / CSm and ETo / CSm of *Symplocos sylvestris* seedling leaves under the L treatment were reduced by 47.3% and 38.6% respectively compared with the CK treatment, with significant differences. Compared with the L treatment, the LG treatment significantly increased the ABS / CSm and ETo / CSm of *Symplocos sylvestris* seedling leaves, by 111.3% and 61.2% respectively.

[0099] 3. Effects of glycine treatment on root growth and development of *Stachys pubescens* seedlings under low temperature

[0100] (1) Effects of glycine treatment on the basic morphological structure of roots of *Stachys pubescens* seedlings grown at low temperatures

[0101] Table 4. Effects of glycine treatment on root morphological parameters of *Sedum morganianum* seedlings under low temperature.

[0102] Treatment Total root length (cm) Root surface area (cm 2 )]]> Total volume (cm 3 ) Root average diameter (mm) CK 1461.59±20.86a 176.81±7.84a 1.68±0.15a 0.38±0.02a L 951.71±49.24c 111.65±8.97c 1.15±0.18b 0.37±0.01a LG 1194.80±11.99b 137.53±1.95b 1.29±0.07b 0.37±0.01a

[0103] The changes in root morphology of *Symplocos rubrum* seedlings after 12 days of low-temperature treatment were analyzed. Figure 4As shown in Table 4, the L treatment significantly inhibited the total root length, total surface area, and total volume of *Symplocos sanguisorba* root system, reducing them by 34.9%, 36.9%, and 31.5% respectively compared to the CK treatment. In contrast, the LG treatment increased the total root length and total surface area of ​​*Symplocos sanguisorba* seedlings, increasing them by 25.5% and 23.2% respectively.

[0104] (2) Effect of glycine treatment on root vigor of *Stachys pubescens* seedlings under low temperature

[0105] like Figure 5 As shown, the root activity of *Staphyllum indicum* under the L treatment was significantly reduced, decreasing by 31.7% compared to the control (CK); while the root activity of *Staphyllum indicum* under the LG treatment was significantly increased, increasing by 28.8% compared to the L treatment.

[0106] 4. Effects of glycine treatment on nitrogen metabolism in *Sedum morganianum* seedlings under low temperature

[0107] (1) Effects of glycine treatment on the contents of nitrate nitrogen, nitrite nitrogen and ammonium nitrogen in the roots and leaves of *Symplocos chinensis* under low temperature

[0108] Depend on Figure 6 A and Figure 6 From B, we know that L treats NO3 in the root system of the downhill stator. - -N and NO2 - -N content decreased with increasing treatment time, and was significantly lower than CK at all treatment times. Compared with L treatment, NO3- content in the roots was significantly lower under LG treatment. - -N and NO2 - -N content increased to varying degrees. Figure 6 From C, we can see that L treatment of NH4 in the root system of the mountain stator + -N content showed a trend of first decreasing and then increasing. Compared with the L treatment, the LG treatment significantly reduced NH4+ in the roots at all treatment times. + The -N content decreased by 23.4%, 35.5%, 32.4%, and 38.7%, respectively.

[0109] Exogenous application of glycine to NO3 in the leaves of *Symplocos henryi* seedlings - -N, NO2 - -N and NH4 + The -N content is also affected to varying degrees. Figure 6 D, Figure 6 E and Figure 6 As shown in F, NO3 in the leaves of *Symplocos chinensis* - The NO2 content decreased with increasing low-temperature treatment time and was significantly lower than the control (CK), decreasing by 10.8%, 13.8%, 22.5%, and 21.4% at different treatment times. - Changes in -N content and NO3- The changes in NH4+ content were similar, showing significant differences compared to the control (CK) at days 9 and 12, decreasing by 20.9% and 24.1%, respectively. Meanwhile, NH4+ in the leaves... + -N content was significantly lower than CK after day 6 in the L treatment. Compared with the L treatment, the LG treatment significantly increased NO3 in the leaves. - -N content. It increased significantly by 29.1% and 22.4% on days 6 and 9 of treatment, respectively, with statistically significant differences. NH4 in leaves + The -N content was significantly lower than that of the L treatment at all time points except for day 12.

[0110] 5. Effects of glycine treatment on total nitrogen content in roots and leaves of *Sedum morganianum* under low temperature.

[0111] like Figure 7 As shown, the TN content in the roots of *Symplocos chinensis* gradually decreased with the extension of low-temperature treatment time, significantly decreasing by 20.9%, 30.9%, 44.4%, and 46.9% compared to the control (CK). The TN content in the leaves significantly decreased after day 6 of treatment and was lower than that of the CK, decreasing by 34.4%, 16.4%, and 60.1%, respectively. Compared to the L treatment, the TN content in the roots of the LG treatment significantly increased after day 6 of treatment, increasing by 54.5%, 75.7%, and 93.9%, respectively. In the leaves, the TN content was significantly higher than that of the L treatment only on days 6 and 12, increasing by 32.6% and 97.7%, respectively.

[0112] 6. Effects of glycine treatment on the activity of nitrogen absorption assimilation enzymes in the roots and leaves of *Symplocos chinensis* under low temperature.

[0113] like Figure 8 As shown, the activities of NR and NADH-GOGAT in the roots of *Symplocos chinensis* gradually decreased under the L treatment, significantly lower than the CK, with both enzyme activities reaching their lowest values ​​on day 12. GS activity under the L treatment showed a trend of first increasing and then decreasing, significantly higher than the CK by 18.9% on day 6, and significantly lower than the CK on days 9 and 12. NADH-GDH activity under the L treatment showed a trend of first decreasing and then increasing, reaching its lowest value on day 6, significantly different from the CK. Compared with the L treatment, the LG treatment promoted the activities of NR, GS, NADH-GOGAT, and NADH-GDH in the roots. Specifically, NADH-GDH activity reached significant levels at all treatment time points; NR activity showed no significant difference from the L treatment at day 3, but was significantly higher than the L treatment at other treatment time points; NADH-GOGAT activity showed no significant difference from the L treatment at day 6, but was significantly higher than the L treatment at other treatment time points; GS activity significantly increased on days 9 and 12.

[0114] like Figure 9As shown, the NR activity in the leaves of *Symplocos henryi* seedlings treated with L was significantly reduced, decreasing by 38.5%, 35.1%, 48.4%, and 58.5%, respectively. Compared with L, LG treatment significantly increased the NR activity in the leaves of *Symplocos henryi* seedlings, increasing by 30.5%, 12.2%, 40.1%, and 86.9%, respectively. Compared with L, the GS activity in the leaves of *Symplocos henryi* seedlings treated with LG was significantly increased. After 6 days of L treatment, the NADH-GOGAT activity in the leaves of *Symplocos henryi* seedlings significantly decreased, decreasing by 45.6%, 40.6%, and 80.3%, respectively. Compared with L treatment, LG treatment significantly increased the NADH-GOGAT activity in the leaves at days 9 and 12. The GDH activity in the leaves of *Symplocos henryi* seedlings treated with L was significantly lower than that in the control group (CK). Compared with the L treatment, the LG treatment increased the GDH activity in the leaves of *Syzygium stenoptera* seedlings by 24.8%, 9.1%, 19.5%, and 41.0%, respectively, and all of these increases were significant except for day 6 of treatment.

[0115] 7. Effects of glycine treatment on nitrogen transaminase activity in roots and leaves of *Symplocos chinensis* under low temperature.

[0116] GOT and GPT activities can reflect the activity levels of amino acid metabolism and protein synthesis and breakdown. For example... Figure 10 A and Figure 10 As shown in Figure B, compared with the control (CK), the activities of GOT and GPT in the roots of *Staphyllum oxypetalum* under the L treatment were significantly reduced. Under the LG treatment, the activities of GOT and GPT in the roots first increased and then decreased. The GOT activity reached its maximum on day 6 of treatment, significantly increasing by 62.7% compared with the L treatment; while the GPT activity reached its maximum on day 9 of treatment, significantly increasing by 52.8%.

[0117] like Figure 10 C and Figure 10 As shown in Figure D, the activities of GOT and GPT in the leaves of *Symplocos edulis* seedlings under the L treatment were significantly lower than those under the control (CK). GOT activity in the leaves under the L treatment showed a trend of first increasing and then decreasing, while GPT activity gradually decreased. Compared with the L treatment, GOT activity in *Symplocos edulis* leaves under the LG treatment was significantly increased at days 9 and 12, increasing by 23.6% and 39.1%, respectively. At days 6, 9, and 12, GPT activity in the leaves was significantly increased compared with the L treatment, increasing by 40.4%, 75.6%, and 65.4%, respectively.

[0118] 8. Effects of glycine treatment on the composition and content of free amino acids in the roots of *Stachys chinensis* under low temperature.

[0119] This invention uses liquid chromatography-mass spectrometry (LC-MS) to analyze and determine 11 free amino acids: alanine, proline, methionine, glycine, glutamic acid, glutamine, arginine, tyrosine, aspartic acid, histidine, and threonine.Figure 11 It was found that on day 3, the L treatment had no significant effect on the amino acid content in the roots of *Symplocos edulis*. Compared with the L treatment, only the proline content increased significantly under the LG treatment. Compared with the control (CK), the contents of threonine and histidine increased significantly, by 33.4% and 26.8%, respectively.

[0120] On day 6 of treatment, the contents of alanine, tyrosine, proline and aspartic acid in the roots of *Symplocos edulis* in treatment L were significantly increased, while the contents of threonine in the roots of *Symplocos edulis* in treatment LG were significantly increased compared with treatment L.

[0121] On day 9, the L treatment significantly increased the contents of alanine, methionine, glycine, arginine, aspartic acid, tyrosine, proline, and glutamic acid in the roots of *Symplocos chinensis*, while the histidine content decreased, but the difference was not significant. Compared with the L treatment, the LG treatment significantly increased the contents of alanine, tyrosine, threonine, glutamic acid, and glutamine, by 17.9%, 17.6%, 24.3%, 5.8%, and 18.0%, respectively.

[0122] On day 12, the L treatment significantly increased the contents of alanine, arginine, tyrosine, proline, threonine, and glutamine in the roots of *Symplocos chinensis*. Compared with the L treatment, the LG treatment further increased the contents of proline, glutamic acid, and glutamine in the roots.

[0123] 9. Effects of glycine treatment on the expression levels of genes related to nitrogen absorption and assimilation in the roots of *Sedum morganianum* under low temperature.

[0124] Depend on Figure 12 It was found that on day 3 of treatment, the expression levels of AMT1.5 and NADH-GOGAT in the roots of *Staphyllum indicum* were significantly downregulated after L treatment. Compared with L treatment, the expression levels of NRT1.1, NRT2.4, NRT2.7, AMT1.2, AMT1.5, and NADH-GOGAT were significantly upregulated after LG treatment.

[0125] On day 6 of treatment, the expression levels of NRT2.5, AMT2.1, NR, GS, and NADH-GOGAT were significantly downregulated in the roots of *Staphyllum oxypetalum* after L treatment, while the expression level of NiR was significantly upregulated. Compared with L treatment, the expression levels of all genes except NRT2.5 and NiR were significantly upregulated after LG treatment.

[0126] On day 9 of treatment, L treatment significantly downregulated the expression levels of NRT1.1, AMT2.1, NR, and NADH-GOGAT, while AMT1.5 and GS expression levels were upregulated. Compared to L treatment, LG treatment only significantly downregulated the expression level of the GS gene, while all other genes were upregulated.

[0127] On day 12 of treatment, L treatment significantly downregulated the expression levels of NRT2.5, AMT2.7, AMT2.1, NR, and NiR, while GS expression was upregulated. Compared to L treatment, LG treatment upregulated the expression levels of all genes except AMT2.1 and GS to varying degrees.

[0128] 10. Effects of glycine treatment on soil nutrient content at low temperatures

[0129] Depend on Figure 13 It was found that low temperature caused a decrease in the contents of available nitrogen, available phosphorus, and organic matter in the soil, while the effect on available potassium content was not significantly different. Available nitrogen content decreased significantly with increasing low temperature treatment time, decreasing by 4.8%, 7.4%, 8.8%, and 8.4%, respectively. Soil available phosphorus content was significantly lower than the control (CK) at treatments 3, 6, and 12 days. Soil organic matter content showed a significant difference only at treatment 12 days, decreasing significantly by 13.2%. Compared with treatment L, soil available nitrogen content in treatment LG increased significantly at all treatment times, increasing significantly by 3.7%, 4.8%, 4.8%, and 6.1%, respectively, while available potassium content decreased significantly, decreasing significantly by 10.7%, 17.8%, 8.8%, and 15.5%, respectively. Available phosphorus content decreased significantly at treatments 3, 9, and 12 days, decreasing by 10.2%, 14.8%, and 16.1%, respectively.

[0130] 11. Effects of glycine treatment on the content of different forms of nitrogen in soil at low temperatures

[0131] Depend on Figure 14 From A, we know that the NO3 in the soil treated with L is... - -N content decreased significantly with increasing low-temperature treatment time. Compared with the L treatment, the NO3- content in the soil at days 9 and 12 was significantly lower with increasing low-temperature treatment time. - -N content increased significantly, by 6.3% and 10.3%, respectively. Figure 14 As shown in B, compared with CK, L treatment of soil NH4 + -N content decreased significantly in the LG treatment. Compared with the L treatment, the LG treatment significantly reduced soil NH4+ content. + -N content further decreased, by 6.2%, 6.5%, 20.2%, and 15.9%, respectively. Figure 14 As shown in C, the soil microbial biomass nitrogen (SMBN) content in the L treatment was significantly different from that in the CK treatment only at day 12, showing a significant decrease of 38.2%. Compared with the L treatment, the SMBN content in the LG treatment was significantly higher than that in the L treatment at all treatment time points.

[0132] 12. Effects of glycine treatment on soil organic nitrogen composition at low temperatures

[0133] like Figure 15It was found that the L treatment significantly reduced the content of soil acidified total nitrogen at day 12; at day 9, the content of soil acidified amino acid nitrogen was significantly reduced compared to the control (CK), decreasing by 59.2%; the L treatment did not have a significant effect on the content of soil acidified amino acid nitrogen and acidified amino sugar nitrogen. Compared with the L treatment, the LG treatment increased the content of all components of soil organic nitrogen.

[0134] 13. Effects of glycine treatment on soil enzyme activity at low temperatures

[0135] like Figure 16 As shown in Figure A, low temperature significantly inhibited soil urease activity, and the activity decreased with increasing low temperature treatment time. Compared with the L treatment, the LG treatment significantly increased soil urease activity at all treatment stages, except for day 3, by 14.0%, 51.7%, and 25.3%, respectively. Figure 16 As shown in Figure B, compared with the control (CK), the L treatment significantly inhibited soil asparaginase activity at 3 and 12 days, with the largest decrease of 17.4% on day 12. Compared with the L treatment, the LG treatment increased soil asparaginase activity at 3 and 12 days, significantly increasing it by 15.6% and 17.0%, respectively. Figure 16 As shown in Figure C, soil glutaminase activity in the L treatment was significantly lower than that in the CK treatment on day 12, decreasing by 19.2%. Compared with the L treatment, the LG treatment showed a significant increase in soil glutaminase activity on day 12, increasing by 25.0%. Figure 16 As shown in Figure D, soil nitrate reductase activity under the L treatment was significantly lower than that under the control (CK) at days 6 and 12, decreasing by 17.1% and 13.9%, respectively. Compared with the L treatment, soil nitrate reductase activity under the LG treatment was significantly increased at day 12, increasing by 22.2%.

[0136] In summary, low temperature reduces the content of photosynthetic pigments in leaves of S. wilsoniana seedlings, inhibits photosynthesis and fluorescence performance of leaves, inhibits the growth of roots and aboveground parts of plants, and reduces the absorption and assimilation of nitrogen. Low temperature also reduces the content of available nitrogen and available phosphorus in soil, reduces the activity of soil nitrogen transformation-related enzymes, and further leads to a decrease in the content of nitrate nitrogen, ammonium nitrogen and microbial biomass nitrogen in soil. The application can alleviate the inhibitory effect of low temperature on the growth of S. wilsoniana seedlings by applying 6 mM glycine, which increases the total root length, root surface area and root volume of seedlings, promotes the accumulation of plant dry matter, improves the photosynthetic and fluorescent indicators of S. wilsoniana seedlings, significantly increases the pigment content in leaves, and promotes photosynthesis. The activity of nitrogen metabolism-related enzymes is enhanced, the expression of nitrogen absorption and assimilation-related genes is improved, and the content of glutamic acid and glutamine in the root system is significantly increased. At the same time, the activity of soil nutrient transformation-related enzymes is increased, and the transformation and supply capacity of nitrogen in soil is improved. Therefore, the application improves the resistance of S. wilsoniana seedlings to low temperature stress by applying glycine.

[0137] Although preferred embodiments of the application have been described, those skilled in the art will be able to make additional changes and modifications without departing from the spirit and scope of the application once they have the benefit of the following claims.

[0138] Obviously, those skilled in the art can make various modifications and variations to the application without departing from the spirit and scope of the application. Thus, if these modifications and variations of the application fall within the scope of the claims and their equivalents, the application also intends to include these modifications and variations.

Claims

1. Use of glycine in the fight against low temperature stress in apple trees, characterized in that, The application relates to a method for improving the resistance of apple seedlings to low-temperature stress by applying a glycine aqueous solution to the apple seedlings. The improved resistance of the apple seedlings to the low-temperature stress is: improving the biomass of the apple seedlings under low-temperature stress, improving the photosynthetic pigment content of the leaves of the apple seedlings under low-temperature stress, improving the photosynthetic parameters of the leaves of the apple seedlings under low-temperature stress, promoting the growth and development of the root system of the apple seedlings under low-temperature stress, enhancing the activity of nitrogen metabolism related enzymes and improving the expression of nitrogen absorption and assimilation related genes. The enhanced activity of the nitrogen metabolism related enzymes is: improving the activity of nitrate reductase, glutamine synthetase, glutamate synthetase and glutamate dehydrogenase. Genes related to improving nitrogen uptake assimilation NRT1.1 , NRT2.4 , NRT2.7 , AMT1.2 , AMT1.5 and NADH-GOGAT ; The concentration of the glycine aqueous solution is 4 mM-8 mM, and the application amount is 150 mL-250 mL per plant.

2. The use of glycine according to claim 1 against low temperature stress in apple trees, characterized by the fact that, The improved biomass of the apple seedlings under low-temperature stress is: improving the above-ground biomass, total biomass and crown-root ratio of the apple seedlings.

3. The use of glycine according to claim 1 against cold stress in apple trees, characterized by the fact that, The improved photosynthetic pigment content of the leaves of the apple seedlings under low-temperature stress is: improving the chlorophyll a content, chlorophyll b content and carotenoid content of the leaves of the apple seedlings under low-temperature stress.

4. The use of glycine according to claim 1 against cold stress in apple trees, characterized by the fact that, The improved photosynthetic parameters of the leaves of the apple seedlings under low-temperature stress are: improving the net photosynthetic rate, transpiration rate, stomatal conductance and water use efficiency of the leaves of the apple seedlings under low-temperature stress.

5. The use of glycine according to claim 1 against cold stress in apple trees, characterized by the fact that, The promoted growth and development of the root system of the apple seedlings under low-temperature stress are: increasing the total root length and total surface area of the root system of the apple seedlings under low-temperature stress and improving the root system activity of the seedlings.

6. The use of glycine according to claim 1 against cold stress in apple trees, characterized by the fact that, The concentration of the glycine aqueous solution is 6 mM, and the application amount is 200 mL per plant.

Images