Application of glycine in resisting low-temperature stress of apple trees

Abstract

The invention relates to the technical field of fruit tree cultivation, in particular to application of glycine in resistance to 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 apple tree seedlings. By increasing the biomass of the apple trees under low-temperature stress, increasing the photosynthetic pigment content of apple tree leaves under low-temperature stress, promoting the growth and development of apple tree seedling roots under low-temperature stress, enhancing the activity of nitrogen metabolism-related enzymes and improving the expression of nitrogen absorption assimilation-related genes, the resistance of the apple trees to low-temperature stress is improved.

Description

Technical Field

[0001] The invention relates to the technical field of fruit tree cultivation, and in particular to application of glycine in resisting low temperature stress of apple trees. Background Art

[0002] Low temperature stress is one of the main hazards that affect the development of fruit trees in cold areas in the north. After a certain period of low temperature stress, the physiological and biochemical indicators and phenotypic characteristics of plants will change to a certain extent. The typical symptoms are slowed growth rate, which leads to disorders in the normal physiological metabolism of plants, making their most basic life activities unable to be guaranteed, and even death. Low temperature stress is divided into chilling injury and freezing injury. The temperature range of chilling injury is generally 0℃~5℃, and the temperature range of freezing injury is generally less than 0℃. Studies have found that after plants are harmed by low temperature, the growth rate of plant height, leaf area, etc. slows down. Sun et al. used corn (Zea mays) as research material and found that low temperature can cause cell dysfunction, and the response of various tissues to low temperature will be different with the change of growth and development stages. At the same time, low temperature can also inhibit the growth of plant roots. The root system is the organ for fruit trees to absorb water and nutrients, and is crucial in the development of fruit trees, such as vegetative growth and reproductive growth. Studies have found that low temperatures in the root zone of fruit trees can cause the growth of apple roots to slow down, which is manifested as a shortening of the total length of the roots, and a reduction in the root surface area and volume. This inhibits the root system's ability to absorb various mineral nutrients, thereby affecting the growth and development of seedlings.

[0003] Photosynthesis is the material and energy basis for plants to produce organic matter and survive. Low temperature can directly cause damage to photosynthetic structures (chloroplasts, etc.), reducing the photosynthetic intensity of plants. Under low temperature conditions, the chlorophyll synthesis process and photosynthetic rate of plants will be significantly inhibited. The decrease in chlorophyll content caused by low temperature is caused by two factors: on the one hand, the enzyme activity of chloroplast pigment synthesis decreases, which in turn inhibits chloroplast synthesis; on the other hand, low temperature conditions destroy the structure of chloroplasts in leaf cells, causing chlorophyll degradation, which in turn affects the capture, conversion and distribution of light energy, resulting in obstruction of photosynthetic electron transfer, damage to photosynthetic machinery, and inhibition of carbon assimilation, which ultimately leads to a decrease in the photosynthetic capacity of leaves. Low temperature destroys the structure of the light system, reduces Pn and chlorophyll content, and has a significant inhibitory effect on plant photosynthesis.

[0004] Plant nutrient absorption is affected by ambient temperature. Low temperature will inhibit the absorption, transformation and utilization of nitrogen by plants. The basic morphological structure of plant roots will be affected by low temperature, and the root system's ability to absorb mineral nutrients will be weakened, causing the aboveground part to grow short. Low temperature inhibits the root system's absorption of nitrogen, phosphorus and potassium. The inhibitory effect on nitrogen absorption is greater than that on 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 nitrogen absorption in sensitive varieties; under long-term low temperature conditions, the nitrogen accumulation of both strong and weak resistant varieties is significantly reduced. Low temperature will also inhibit the activity of enzymes related to nitrogen absorption and metabolism, thereby affecting the nitrogen metabolism process of plants, thereby affecting plant growth.

[0005] Apple is a perennial deciduous fruit tree with a large demand for nitrogen, but low nitrogen utilization is an important problem facing apple production and one of the main factors affecting apple yield and quality. Low temperature is the main environmental stress factor in early spring and late autumn in my country's northern apple-producing areas. Low temperatures in spring affect nitrogen mineralization in the soil and reduce nitrogen absorption by fruit tree roots. It can also directly affect the nutrient concentration of the tree by inhibiting the absorption and transportation of nutrients by the plant's roots, affecting the early spring organ construction of apple trees, manifested as delayed leaf development and reduced fruit set, which in turn affects apple yield and quality. Therefore, alleviating low temperature stress in apples is an issue that needs to be addressed urgently. Summary of the invention

[0006] In order to alleviate the low temperature stress of apples, the present invention provides the application of glycine in the resistance to low temperature stress of apple trees. The present invention improves the resistance of apple trees to low temperature stress by increasing the biomass of apple trees under low temperature stress, increasing the content of photosynthetic pigments in apple tree leaves under low temperature stress, increasing photosynthetic parameters of apple tree leaves under low temperature stress, promoting the growth and development of the root system of apple tree seedlings under low temperature stress, enhancing the activity of nitrogen metabolism-related enzymes and increasing the expression of nitrogen absorption and assimilation-related genes.

[0007] The invention provides an application of glycine in resisting low temperature stress of apple trees, and improves the resistance of apple trees to low temperature stress by applying a glycine aqueous solution to apple tree seedlings. The low temperature stress conditions of the invention are 8°C to 10°C during the day and 0°C to 2°C at night.

[0008] The invention improves the biomass of apple trees under low temperature stress, improves the content of photosynthetic pigments in leaves of apple trees under low temperature stress, improves photosynthetic parameters of leaves of apple trees under low temperature stress, promotes the growth and development of the root system of apple tree seedlings under low temperature stress, enhances the activity of nitrogen metabolism-related enzymes and improves the expression of nitrogen absorption and assimilation-related genes by irrigating the apple trees with a glycine aqueous solution during the seedling stage of the apple trees, thereby improving the resistance of the apple trees to low temperature stress.

[0009] Compared with the prior art, the present invention has the following beneficial effects:

[0010] The present invention improves the resistance of apple trees to low temperature stress by the following methods:

[0011] Increase the biomass of apple trees under low temperature stress, such as the aboveground biomass, total biomass and crown-root ratio of seedlings; increase the content of photosynthetic pigments in apple tree leaves under low temperature stress, such as chlorophyll a, chlorophyll b and carotenoid content; improve the photosynthetic parameters of apple tree leaves under low temperature stress, such as net photosynthetic rate, transpiration rate, stomatal conductance and water use efficiency; promote the growth and development of the root system of apple seedlings under low temperature stress, such as increasing the total root length and total surface area of ​​apple seedlings under low temperature stress and improving the root activity of seedlings; enhance the activity of nitrogen metabolism-related enzymes, such as increasing the activity of nitrate reductase, glutamine synthetase, glutamate synthetase and glutamate dehydrogenase; improve the expression of nitrogen absorption and assimilation-related genes NRT1.1, NRT2.4, NRT2.7, AMT1.2, AMT1.5 and NADH-GOGAT. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

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

[0014] Figure 2 Figure 1 is the effect of glycine on the content of photosynthetic pigments in the leaves of mountain stator under low temperature; in the figure, A is the effect of glycine on the content of chlorophyll a in the leaves of mountain stator under low temperature; B is the effect of glycine on the content of chlorophyll b in the leaves of mountain stator under low temperature; C is the effect of glycine on the content of carotenoids in the leaves of mountain stator under low temperature; D is the effect of glycine on the total chlorophyll content in the leaves of mountain stator under low temperature.

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

[0016] Figure 4This is the effect of glycine treatment on the root morphology of Aralia dasyphylla seedlings under low temperature.

[0017] Figure 5 The effect of glycine on the root activity of Scutellaria baicalensis under low temperature.

[0018] Figure 6 Figure 3 shows the effect of glycine treatment on the contents of nitrate nitrogen, nitrite nitrogen and ammonium nitrogen in the roots and leaves of Psoralea corylifolia under low temperature. In the figure, A shows the effect of glycine treatment on nitrate nitrogen in the roots of Psoralea corylifolia under low temperature. B shows the effect of glycine treatment on nitrite nitrogen in the roots of Psoralea corylifolia under low temperature. C shows the effect of glycine treatment on ammonium nitrogen in the roots of Psoralea corylifolia under low temperature. D shows the effect of glycine on nitrate nitrogen in the leaves of Psoralea corylifolia seedlings. E shows the effect of glycine on nitrite nitrogen in the leaves of Psoralea corylifolia seedlings. F shows the effect of glycine on ammonium nitrogen in the leaves of Psoralea corylifolia seedlings.

[0019] Figure 7 Figure 3 shows the effect of glycine treatment on the total nitrogen content in the roots and leaves of Psoralea corylifolia under low temperature. Figure 3 shows the effect of glycine treatment on the total nitrogen content in the roots ... leaves of Psoralea corylifolia under low temperature.

[0020] Figure 8 Figure 1 shows the effect of glycine treatment on the activity of nitrogen absorption and assimilation enzymes in the roots of Scutellaria baicalensis under low temperature; in the figure, A shows the effect of glycine treatment on the activity of nitrate reductase in the roots of Scutellaria baicalensis under low temperature; B shows the effect of glycine treatment on the activity of glutamine synthetase in the roots of Scutellaria baicalensis under low temperature; C shows the effect of glycine treatment on the activity of glutamate synthetase in the roots of Scutellaria baicalensis under low temperature; D shows the effect of glycine treatment on the activity of glutamate dehydrogenase in the roots of Scutellaria baicalensis under low temperature.

[0021] Fig. 9 Figure 1 shows the effect of glycine treatment on the activity of nitrogen absorption and assimilation enzymes in the leaves of mountain stator under low temperature; In the figure, A shows the effect of glycine treatment on the activity of nitrate reductase in the leaves of mountain stator under low temperature; B shows the effect of glycine treatment on the activity of glutamine synthetase in the leaves of mountain stator under low temperature; C shows the effect of glycine treatment on the activity of glutamate synthetase in the leaves of mountain stator under low temperature; D shows the effect of glycine treatment on the activity of glutamate dehydrogenase in the leaves of mountain stator under low temperature.

[0022] Fig.10 Figure 3 shows the effect of glycine treatment on the activities of aspartate aminotransferase (GOT) and alanine aminotransferase (GPT) in the roots and leaves of Scutellaria baicalensis under low temperature; in the figure, A shows the effect of glycine treatment on the aspartate aminotransferase activity in the roots of Scutellaria baicalensis under low temperature; B shows the effect of glycine treatment on the alanine aminotransferase activity in the roots of Scutellaria baicalensis under low temperature; C shows the effect of glycine treatment on the aspartate aminotransferase activity in the leaves of Scutellaria baicalensis under low temperature; D shows the effect of glycine treatment on the alanine aminotransferase activity in the leaves of Scutellaria baicalensis under low temperature.

[0023] Fig.11 Figure 1 shows the effect of glycine treatment on the composition and content of free amino acids in the roots of Psoralea corylifolia under low temperature. In the figure, A shows the effect of glycine treatment on the alanine content in the roots of Psoralea corylifolia under low temperature. B shows the effect of glycine treatment on the methionine content in the roots of Psoralea corylifolia under low temperature. C shows the effect of glycine treatment on the glycine content in the roots of Psoralea corylifolia under low temperature. D shows the effect of glycine treatment on the arginine content in the roots of Psoralea corylifolia under low temperature. E shows the effect of glycine treatment on the tyrosine content in the roots of Psoralea corylifolia under low temperature. F shows the effect of glycine treatment on the proline content in the roots of Psoralea corylifolia under low temperature. G shows the effect of glycine treatment on the threonine content in the roots of Psoralea corylifolia under low temperature. H shows the effect of glycine treatment on the histidine content in the roots of Psoralea corylifolia under low temperature. I shows the effect of glycine treatment on the glutamate content in the roots of Psoralea corylifolia under low temperature. J shows the effect of glycine treatment on the glutamine content in the roots of Psoralea corylifolia under low temperature. K shows the effect of glycine treatment on the aspartic acid content in the roots of Psoralea corylifolia under low temperature.

[0024] Fig.12 This is the effect of glycine treatment on the expression levels of genes related to nitrogen absorption and assimilation in the roots of Psoralea corylifolia under low temperature.

[0025] Fig.13 Figure 1 shows the effect of glycine treatment on soil nutrient content under low temperature. In the figure, A shows the effect of glycine treatment on soil available nitrogen content under low temperature. B shows the effect of glycine treatment on soil available phosphorus content under low temperature. C shows the effect of glycine treatment on soil available potassium content under low temperature. D shows the effect of glycine treatment on soil organic matter content under low temperature.

[0026] Fig.14 Figure 3 shows the effect of glycine treatment on the content of different forms of soil nitrogen under low temperature. In the figure, A shows the effect of glycine treatment on the content of nitrate nitrogen in soil under low temperature. B shows the effect of glycine treatment on the content of ammonium nitrogen in soil under low temperature. C shows the effect of glycine treatment on the content of microbial biomass nitrogen in soil under low temperature.

[0027] Fig.15 Figure 3 shows the effect of glycine treatment on soil organic nitrogen components under low temperature. In the figure, A shows the effect of glycine treatment on the acid-hydrolyzable total nitrogen content in soil under low temperature. B shows the effect of glycine treatment on the acid-hydrolyzable amino acid nitrogen content in soil under low temperature. C shows the effect of glycine treatment on the acid-hydrolyzable amino acid nitrogen content in soil under low temperature. D shows the effect of glycine treatment on the acid-hydrolyzable amino sugar nitrogen content in soil under low temperature.

[0028] Fig.16Figure 2 shows the effect of glycine treatment on soil enzyme activities under low temperature. In the figure, A shows the effect of glycine treatment on soil urease activity under low temperature. B shows the effect of glycine treatment on soil asparaginase activity under low temperature. C shows the effect of glycine treatment on soil glutaminase activity under low temperature. D shows the effect of glycine treatment on soil nitrate reductase activity under low temperature. DETAILED DESCRIPTION

[0029] The specific embodiments of the present invention are described in detail below, but it should be understood that the protection scope of the present invention is not limited by the specific embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without creative work belong to the scope of protection of the present invention. The experimental methods described in the embodiments of the present invention are conventional methods unless otherwise specified, and the materials, reagents, etc. used in the following embodiments can be obtained from commercial channels unless otherwise specified.

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

[0031] 1. Experimental Materials and Methods

[0032] 1. Experimental Materials and Treatment

[0033] One-year-old Malus baccata Borkh. seedlings were used as test materials. The Malus baccata seeds were sown after stratification and whitening. When the seedlings grew to about 4 to 5 true leaves, they were transplanted into nutrient pots containing mixed substrates (orchard soil, seedling substrate and river sand were mixed in a ratio of 2:1:1). One Malus baccata seedling was planted in each nutrient pot. Water was applied in a timely and appropriate amount every day, and attention was paid to the prevention of diseases and insect pests during the period. All test materials were moved to the cold-proof shed for wintering before the leaves fell in winter. In April of the second year, when the seedlings began to sprout, the test treatment was carried out. The seedlings with consistent growth, no diseases and insect pests, and strong were selected. The seedlings were divided into two groups and placed in two artificial climate boxes with different temperatures for cultivation. The cultivation temperatures were normal temperature: 24℃ (day) / 14℃ (night) and low temperature: 10℃ (day) / 2℃ (night), and the light intensity was set to 400μmol·m -2 ·s -1 , 12 hours of light per day, average relative humidity 75%, and the experimental treatment began after the seedlings had been pre-cultured for 1 day.

[0034] Three treatments were set up in the experiment, namely: (1) control (CK): room 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 always kept at 24℃ (day) / 14℃ (night) were used as the control.

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

[0036] 2. Determination of biomass

[0037] On the 12th day of treatment, 5 plants were taken from each treatment, the seedlings of the yam tree were washed with clean water, the water attached to the surface was dried with absorbent paper, the seedlings were cut from the rhizome with scissors and weighed separately with a balance, and this was recorded as fresh weight; then the weighed tissue materials were placed in an oven, fixed at 121℃ for 20min, dried at 60℃ to constant weight, and after cooling, the dry matter mass of each part was weighed with an electronic balance, and recorded as dry weight. The dry weight ratio of the aboveground part to the underground part represents the root-crown ratio.

[0038] 3. Determination of morphological indicators

[0039] The plant height and stem diameter of the schisandra chinensis were measured with a tape measure and a vernier caliper (accuracy 0.02) after 12 days of treatment.

[0040] Root morphology indicators: When sampling was carried out on the 12th day of treatment, the root morphology indicators of S. salsa were quantitatively analyzed using a digital scanner and WinRhizo PRO2016 analysis software.

[0041] 4. Determination of photosynthetic parameters of seedlings

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

[0043] 5. Determination of chlorophyll content

[0044] The method for determining the chlorophyll content of seedling leaves is as follows: after taking the leaves of Sterculia sibiricum, weigh 40 mg and place them in a centrifuge tube, add 0.25 mL of pure acetone and 8.75 mL of 80% acetone to each tube, cover the lid and shake violently once, then extract in a dark room at room temperature overnight. To speed up the extraction effect, shake four to five times during this period until the sample turns completely white. On the next day, add 1 mL of 80% acetone and make up the volume, and use a UV spectrophotometer to measure the wavelengths at 470 nm, 646 nm and 663 nm and calculate.

[0045] Calculation formula: C Chla =12.21×OD 663 -2.81×OD 646 Where, C Chla Indicates chlorophyll a content; OD 663 Indicates the wavelength of 663nm of UV spectrophotometer; OD 646 Indicates the wavelength of 646nm in UV spectrophotometer.

[0046] C Chlb =20.13×OD 646 -5.03×OD 663 ; C Chlb Indicates chlorophyll b content; OD 663 Indicates the wavelength of the UV spectrophotometer is 663nm; OD 646 Indicates the wavelength of 646nm in UV spectrophotometer.

[0047] C Chl(a+b) =Chla+Chlb; where 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×OD 470 -3.27×Ca-104×Cb) / 229; where C Car Indicates carotenoid content; OD 470 represents the wavelength of 646nm of the ultraviolet spectrophotometer; Ca represents the chlorophyll a content; Cb represents the chlorophyll b content.

[0049] Chlorophyll content Where, C: pigment concentration; V: extraction volume; m: sample mass.

[0050] 6. Determination of chlorophyll fluorescence parameters

[0051] After the leaves were dark-adapted for 20 min, the basic fluorescence parameters of the leaves of P. truncatula were measured using a Pocket PEA (Hansatech, UK) fluorometer.

[0052] 7. Determination of root activity

[0053] After the test materials were treated, the root activity of the seedlings of the yam tree was measured by the triphenyltetrazolium chloride reduction method (TTC method) on the 12th day. The operation was as follows: the white young roots were cut into 2 cm long segments and about 0.5 g was weighed, and 1 / 15 mol·L -1Phosphate buffer and TTC were kept at 37°C for 4 hours. 1M H2SO4 was added to terminate the reaction and allowed to stand for 15 minutes. The roots were then taken out and the liquid attached to their surface was dried. 10 mL of ethanol was added and the roots were extracted at 25°C for 24 hours. The absorbance at 485 nm was measured using a UV spectrophotometer.

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

[0055] Nitrate nitrogen (NO3 - The salicylic acid-sulfuric acid method described by Patterson et al. in "Distinct signalling pathways and transcriptome response signatures differentiate ammonium- and nitrate-supplied plants" was used to determine the ammonium-N content.

[0056] Nitrite (NO2 - The α-naphthylamine method described by Ogawa et al. in "Relationships between nitrite reductase activity and genotype-dependent callus growth in rice cell cultures" was used to determine the nitrite reductase content.

[0057] Ammonium nitrogen (NH4 + -N) content was determined using et al. recorded the ninhydrin method in "High-throughput colorimetric method for the parallel assay of glyoxylic acid and ammonium in a single extract".

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

[0059] Add 1.4 mL of 0.1 mmol·L -1 KNO3 and 0.2 mL NADH were mixed thoroughly, incubated at room temperature for 0.5 h, and then the colorimetric agent was added immediately. After centrifugation, the absorbance at 540 nm was measured. The activity of NADH-GOGAT was measured according to the method described by Zheng Chaofeng et al. in "Rapid Determination of Glutamate Synthase Activity". After 0.3 mL of enzyme solution was drawn, 100 mmol·L - 1 Tris-HCl (pH=7.5), 0.1mmol·L -1NADH, 10mmol·L -1 α-ketoglutaric acid and 10mmol·L -1 The reaction was started with glutamine buffer, and the absorbance at 340 nm was measured 10 times in succession. The value at a stable decrease in density was used to measure the enzyme activity. The activity of glutamate dehydrogenase (GDH) was determined according to the method recorded by 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". The activity of GOT and GPT in the roots and leaves of Vitis vinifera was determined according to the method recorded by Wu Lianghuan et al. in "Progress in the absorption and utilization of amino acid nitrogen by higher plants". After aspirating the crude enzyme solution and enzyme substrate, place them in a 37°C water bath for 1.5 hours, then take them out, add 2,4-dinitrophenylhydrazine, and bathe again in water for 20 minutes. Finally, add 0.4 mol·L -1 Mix NaOH thoroughly, let it stand for 10 min, adjust to zero with distilled water and measure the absorbance at 500 nm.

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

[0061] Weigh 0.3g of fresh plant tissue sample, add 1mL of mixed solution (deionized water: chloroform: methanol = 3:5:12, v / v / v), shake thoroughly (4℃, 30min), centrifuge (4℃, 12000×g, 15min), and collect the supernatant in a new centrifuge tube. Add 375μL of chloroform and 625μL of deionized water to wash away impurities in the solution, blow the supernatant to near dryness with a low-temperature rotary freeze dryer, add 1mL of acetonitrile, pass through an organic filter membrane (0.22μm) and then put it on the machine. The mobile phase I of the liquid chromatography is ammonium acetate and acetonitrile, and the mobile phase II is a mixed aqueous solution of ammonium acetate and formic acid. The mobile phase flow rate is set to 0.3μL·min -1 , injection volume 5 μL.

[0062] 11. Determination of gene expression

[0063] The CTAB method was used to extract RNA from plant roots. Specifically, 150 mg of plant tissue samples were taken, and 700 μL of CTAB (preheated at 65°C to make the turbid and viscous reagent transparent) and 50 μL of β-mercaptoethanol were added to the tube. This process was carried out in a fume hood. After mixing, the tube was placed in a water bath at 65°C for 30 minutes. During this period, the centrifuge tube was inverted and mixed several times. After the water bath, an equal volume of chloroform:isoamyl alcohol (volume ratio of 24:1) was added. After sufficient mixing, the tube was centrifuged. The supernatant was recovered for the second extraction. A new centrifuge tube was taken, and the supernatant and 50 μL of lithium chloride were added. The tube was placed in a -20°C refrigerator overnight. After standing, the precipitate was centrifuged at 4°C to retain the precipitate, and 75% ethanol was added to remove impurities and centrifuged again. The precipitate was washed once with anhydrous ethanol and centrifuged, then placed in a clean bench, blown dry, and DEPC water was added to preserve the RNA. cDNA synthesis was performed using a reverse transcription kit (Takara, Japan) according to the instructions. The synthesized cDNA was stored in a -20°C refrigerator for future use. Real-time fluorescence quantitative (RT-qPCR) amplification was performed using a 384-well plate. Primers were synthesized by Sangon Biotech (Shanghai) Co., Ltd. The forward and reverse primers are shown in Table 1. The cDNA was diluted 10 times for qRT-PCR and 2 -ΔΔCt Methods The relative gene expression was calculated.

[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 Shidan in "A Review of Nitrogen Utilization in Plants".

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

[0069] The soil microbial biomass nitrogen (SMBN) content was determined using the chloroform fumigation-K2SO4 extraction method described by Lin Xiangui in Principles and Methods of Soil Microbial Research.

[0070] NO3 in soil - The method for determining the -N content is as follows: accurately weigh 10 g of air-dried soil that has passed through a 20-mesh sieve and add 0.01 mol·L -1 Calcium chloride (CaCl2) extract, shake well. Shake for 30 minutes and filter. After absorbing the filtrate, immediately add 1mL sulfuric acid solution (1:9) to acidify the test solution and shake well. Zero with the acidified CaCl2 extract and measure the absorbance at 210nm and 275nm.

[0071] NH4 in soil +The method for determining the -N content is as follows: weigh 5 g of fresh soil sample that has passed through a 20-mesh sieve and accurately add 2 mol L -1 After adding 25 mL of potassium chloride (KCl) solution, shake it thoroughly and oscillate for 1 hour. After taking out the suspension to be clarified, filter the upper clear liquid of the suspension, which is the soil leachate to be tested. Add phenol solution and 2 mol·L -1 5 mL of KCl and sodium hypochlorite alkaline solution were incubated at 27 °C for 1 h, and a masking agent (400 g L -1 Potassium sodium tartrate, 100g·L -1 EDTA disodium salt, 10 mol·L -1 The precipitate was dissolved with NaOH and the absorbance at 625 nm was measured.

[0072] 14. Determination of soil organic nitrogen components

[0073] The determination of soil organic nitrogen components was based on the method described by Lu Rukun in "Soil Agricultural Chemical Analysis Methods".

[0074] 15. Determination of soil enzyme activity

[0075] Soil urease activity was measured using the sodium phenol-sodium hypochlorite colorimetric method with a slight modification. Specifically, 10 g of sample was weighed, 1 mL of toluene was added and shaken, reacted for 15 min, citric acid buffer with a pH of 6.7 and urea were added, mixed, and cultured at 37 °C for 1 day. The next day, distilled water and 1.53 mol·L -1 Sodium phenolate and sodium hypochlorite solutions were fully mixed and shaken until the solution turned into indigo blue. The absorbance at 578 nm was measured within 1 hour.

[0076] The activities of soil nitrate reductase (NR), soil glutaminase (GLNS) and soil asparaginase (ASP) were measured according to the instructions of the corresponding kits (Jiangsu Enzyme-Free Industry Co., Ltd.). The measurement wavelength was 450 nm. The standard curve was based on the instructions provided by the kits.

[0077] 16. Data statistics and analysis

[0078] Microsoft Excel was used to input and organize the data, and SPSS27.0 software was used for statistical analysis. The data were tested for significance (p < 0.05) by single factor and least significant difference method (LSD method). Origin 2023 was used to draw the graph. The data in the charts are expressed as the mean ± standard error of three experimental repetitions.

[0079] 2. Experimental Results

[0080] 1. Effects of exogenous glycine on the growth of Psoralea corylifolia seedlings under low temperature

[0081] (1) Effects of glycine treatment on biomass of Psoralea corylifolia under low temperature

[0082] As shown in Table 2, compared with CK, low temperature treatment for 12 days significantly inhibited the normal growth of S. cerana seedlings. The plant height, aboveground biomass, root biomass, total biomass and crown-root ratio of S. cerana seedlings were reduced by 16.5%, 37.9%, 22.5%, 32.9% and 20.7% respectively compared with CK, with significant differences. Compared with the L treatment, the aboveground biomass, total biomass and crown-root ratio of the seedlings under the LG treatment increased to varying degrees, increasing by 28.0%, 2.6%, and 21.5%, respectively, with significant differences. This shows that exogenous application of glycine can effectively alleviate the inhibitory effect of low temperature on the growth of S. cerana seedlings ( Figure 1 ).

[0083] Table 2 Effects of glycine on growth parameters of Scutellaria baicalensis plants under low temperature

[0084] deal with Plant height (cm) Stem diameter (mm) Aboveground biomass (g DW) Root biomass (g DW) Total biomass (g DW) Crown-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 significant differences between different treatments in the same column, p < 0.05. CK: control, room temperature + distilled water; L: low temperature treatment; LG: low temperature + glycine treatment.

[0086] 2. Effects of glycine treatment on photosynthetic and fluorescence characteristics of leaves of P. truncatula under low temperature

[0087] (1) Effects of glycine treatment on the content of photosynthetic pigments in leaves of Sterculia serrata under low temperature

[0088] Depend on Figure 2 It can be seen that compared with the L treatment, the chlorophyll a (Chla), chlorophyll b (Chlb) and carotenoid contents in the LG treatment increased significantly, by 93.3%, 28.4% and 21.0%, respectively.

[0089] (2) Effects of glycine treatment on photosynthetic parameters of steller leaves under low temperature

[0090] Table 3 Effects of different treatments on photosynthetic parameters of Psoralea corylifolia seedlings

[0091]

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

[0093] (3) Effects of glycine treatment on fluorescence parameters of steller leaves under low temperature

[0094] Depend on Figure 3 A showed that the F0 of the leaves of the mountain stator seedlings increased by 16.3% after 12 days of L treatment, which was a significant difference. Compared with the L treatment, the F0 under the LG treatment decreased by 7.2%, which was not a significant difference.

[0095] Fv / F0 reflects the potential activity of PSⅡ. Figure 3 From B, we can see that the Fv / F0 of the leaves of the schizonepeta tenuifolia seedlings under L treatment for 12 days was significantly reduced by 25.9%. Compared with the L treatment, the Fv / F0 under LG treatment increased by 33.0%, which was a significant difference.

[0096] Fv / Fm indicates the conversion efficiency of light energy in the PSⅡ reaction center to a certain extent. Figure 3 It can be seen from the C that low temperature caused a significant decrease in Fv / Fm of leaves of S. cerana seedlings, which decreased by 28.4%. Compared with the L treatment, the Fv / Fm in the LG treatment increased by 25.2%, which was a significant difference.

[0097] The photosynthetic performance index (PI abs) can reflect the overall photosynthetic capacity of the plant. Figure 3 As shown in D, PIabs decreased significantly under L treatment, which was 37.6% lower than that of CK. Compared with L treatment, PI abs of leaves under LG treatment increased significantly by 66.3%.

[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 The results of E and F of 3 showed that the ABS / CSm and ETo / CSm of the leaves of the seedlings of the mountain stator in the L treatment were 47.3% and 38.6% lower than those of the CK, respectively, with significant differences. Compared with the L treatment, the LG treatment significantly increased the ABS / CSm and ETo / CSm of the leaves of the mountain stator seedlings, increasing by 111.3% and 61.2%, respectively.

[0099] 3. Effects of glycine treatment on root growth and development of Psoralea corylifolia seedlings under low temperature

[0100] (1) Effects of glycine treatment on the basic morphological structure of the root system of Aralia serrata seedlings under low temperature

[0101] Table 4 Effects of glycine treatment on root morphological parameters of Dictyophora serrata seedlings under low temperature

[0102] deal with Total root length (cm) <![CDATA[Root surface area (cm 2 )]]> <![CDATA[Total volume (cm 3 )]]> Average root 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] After 12 days of low temperature treatment, the changes in root morphology of the seedlings of Psoralea corylifolia were analyzed ( Figure 4). As shown in Table 4, the L treatment significantly inhibited the total root length, total surface area and total volume of the root system of the Achyranthes bidentata, which were reduced by 34.9%, 36.9% and 31.5% respectively compared with the CK. Compared with the L treatment, the LG treatment increased the total root length and total surface area of ​​the Achyranthes bidentata seedlings by 25.5% and 23.2% respectively.

[0104] (2) Effects of glycine treatment on root activity of Psoralea corylifolia seedlings under low temperature

[0105] like Figure 5 As shown, the root activity of S. truncatum was significantly reduced under L treatment, which was 31.7% lower than that of CK; the root activity of S. truncatum was significantly increased under LG treatment, which was 28.8% higher than that of L treatment.

[0106] 4. Effects of glycine treatment on nitrogen metabolism of Psoralea corylifolia seedlings under low temperature

[0107] (1) Effects of glycine treatment on the contents of nitrate, nitrite, and ammonium in roots and leaves of P. truncatula under low temperature

[0108] Depend on Figure 6 A and Figure 6 From B, we can see that under L treatment, NO3 - -N and NO2 - -N content showed a decreasing trend with the extension of treatment time, and was significantly lower than that of CK at each treatment time. Compared with the L treatment, the NO3 - -N and NO2 - -N content increased to varying degrees. Figure 6 It can be seen from the C that under the L treatment, the NH4 + -N content showed a trend of decreasing first and then increasing. Compared with the L treatment, the LG treatment significantly reduced the NH4 + -N content decreased by 23.4%, 35.5%, 32.4% and 38.7% respectively.

[0109] Effect of exogenous application of glycine on NO3 in leaves of Psoralea corylifolia seedlings - -N, NO2 - -N and NH4 + -N content also has different degrees of influence. Figure 6 D. Figure 6 E and Figure 6 As shown in F, NO3 in the stator blades - -N content showed a decreasing trend with the extension of low temperature treatment time, and was significantly lower than that of CK, decreasing by 10.8%, 13.8%, 22.5% and 21.4% at each treatment time, respectively. - -N content changes and NO3- -N content changes were similar, with significant differences compared with CK at the 9th and 12th days of treatment, decreasing by 20.9% and 24.1% respectively. + -N content in L treatment was significantly lower than that in CK after 6 days. Compared with L treatment, LG treatment significantly increased NO3 - -N content. It increased significantly by 29.1% and 22.4% on the 6th and 9th day of treatment, with significant differences. + -N content was significantly lower than that of L treatment at other time points except the 12th day.

[0110] 5. Effect of glycine treatment on total nitrogen content in roots and leaves of P. truncatula under low temperature

[0111] like Figure 7 As shown in the figure, the TN content in the roots of the shandingzi gradually decreased with the extension of the low temperature treatment time, and compared with CK, it was significantly reduced by 20.9%, 30.9%, 44.4% and 46.9%, respectively, while the TN content in the leaves decreased significantly after the 6th day of treatment and was lower than that of CK, decreasing by 34.4%, 16.4% and 60.1%, respectively. Compared with the L treatment, the TN content in the roots of the LG treatment increased significantly after the 6th day 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 at the 6th and 12th days of treatment, increasing by 32.6% and 97.7%.

[0112] 6. Effects of glycine treatment on the activities of nitrogen absorption and assimilation enzymes in roots and leaves of Psoralea corylifolia under low temperature

[0113] like Figure 8 As shown in the figure, the activities of NR and NADH-GOGAT in the roots of D. truncatum gradually decreased under the L treatment and were significantly lower than those of CK. Both enzyme activities reached the lowest value at the 12th day of treatment. The activity of GS showed a trend of increasing first and then decreasing under the L treatment. It was significantly higher than CK at the 6th day of treatment, with a significant increase of 18.9%, and was significantly lower than CK at the 9th and 12th days of treatment. The activity of NADH-GDH showed a trend of decreasing first and then increasing under the L treatment, reaching the lowest value at the 6th day of treatment, which was significantly different from that of CK. Compared with the L treatment, the LG treatment promoted the activities of NR, GS, NADH-GOGAT and NADH-GDH in the roots. Among them, the activity of NADH-GDH reached a significant level at each treatment time point. There was no significant difference between the activity of NR and the L treatment at 3d, and it was significantly higher than that of the L treatment at other treatment time points. There was no significant difference between the activity of NADH-GOGAT and the L treatment at 6d, and it was significantly higher than that of the L treatment at other treatment time points. The activity of GS increased significantly at the 9th and 12th days of treatment.

[0114] like Fig. 9As shown in the results, the NR activity in the leaves of D. shandingzi seedlings under L treatment was significantly reduced, by 38.5%, 35.1%, 48.4% and 58.5%, respectively. Compared with the L treatment, the LG treatment significantly increased the NR activity in the leaves of D. shandingzi seedlings, by 30.5%, 12.2%, 40.1% and 86.9%, respectively. Compared with the L treatment, the GS activity in the leaves of D. shandingzi seedlings under LG treatment was significantly increased. After 6 days of L treatment, the NADH-GOGAT activity in the leaves of D. shandingzi seedlings decreased significantly, by 45.6%, 40.6% and 80.3%, respectively. Compared with the L treatment, the LG treatment significantly increased the NADH-GOGAT activity in the leaves at the 9th and 12th days of treatment. The GDH activity in the leaves of D. shandingzi seedlings under L treatment was significantly lower than that under CK. Compared with the L treatment, the LG treatment increased the GDH activity in the leaves of the Achyranthes bidentata seedlings by 24.8%, 9.1%, 19.5% and 41.0%, respectively, reaching a significant level except for the 6th day of treatment.

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

[0116] GOT and GPT activities can reflect the activity of amino acid metabolism and protein synthesis and decomposition. Fig.10 A and Fig.10 As shown in B, compared with CK, the activities of GOT and GPT in the roots of M. truncatum were significantly reduced under L treatment. The activities of GOT and GPT in the roots of M. truncatum increased first and then decreased under LG treatment. The activity of GOT reached the maximum value on the 6th day of treatment, which was significantly increased by 62.7% compared with the L treatment; while the activity of GPT reached the maximum value on the 9th day of treatment, which was significantly increased by 52.8%.

[0117] like Fig.10 C and Fig.10 As shown in D, the activities of GOT and GPT in the leaves of the seedlings of the mountain stator under the L treatment were significantly lower than those of the CK. The activity of GOT in the leaves showed a trend of increasing first and then decreasing under the L treatment, while the activity of GPT gradually decreased. Compared with the L treatment, the activity of GOT in the leaves of the mountain stator under the LG treatment increased significantly by 23.6% and 39.1% on the 9th and 12th days of treatment, respectively. On the 6th, 9th and 12th days of treatment, the activity of GPT in the leaves increased significantly 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 Psoralea corylifolia under low temperature

[0119] The present invention uses liquid chromatography-mass spectrometry to analyze and detect 11 kinds of free amino acids, namely alanine, proline, methionine, glycine, glutamic acid, glutamine, arginine, tyrosine, aspartic acid, histidine and threonine. Fig.11 It can be seen that on the third day of treatment, the L treatment had no significant effect on the content of amino acids in the roots of P. truncatum. Compared with the L treatment, only the proline content increased significantly under the LG treatment. Compared with the CK treatment, the threonine and histidine contents increased significantly, by 33.4% and 26.8%, respectively.

[0120] On the 6th day of treatment, the contents of alanine, tyrosine, proline and aspartic acid in the roots of S. salsa subsp. tinctorius treated with L significantly increased. Compared with the L treatment, the content of threonine in the roots of S. salsa subsp. tinctorius treated with LG significantly increased.

[0121] On the 9th day of treatment, the contents of alanine, methionine, glycine, arginine, aspartic acid tyrosine, proline and glutamic acid in the roots of the L-treated S. cerevisiae were significantly increased, and the content of histidine was decreased, but there was no significant difference. Compared with the L-treated, the LG-treated significantly increased the contents of alanine, tyrosine, threonine, glutamic acid and glutamine, which increased by 17.9%, 17.6%, 24.3%, 5.8% and 18.0%, respectively.

[0122] On the 12th day of treatment, the L treatment significantly increased the contents of alanine, arginine, tyrosine, proline, threonine and glutamine in the roots of C. salsa. 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 of genes related to nitrogen absorption and assimilation in roots of P. truncatula under low temperature

[0124] Depend on Fig.12 It can be seen that on the third day of treatment, the expression levels of AMT1.5 and NADH-GOGAT in the roots of C. truncatula 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 in LG treatment were significantly upregulated.

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

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

[0127] On the 12th day of treatment, the expression levels of NRT2.5, AMT2.7, AMT2.1, NR and NiR were significantly downregulated, while the expression level of GS was upregulated after L treatment. Compared with L treatment, after LG treatment, except for the downregulation of the expression levels of AMT2.1 and GS genes, the expression of other genes was upregulated to varying degrees.

[0128] 10. Effect of glycine treatment on soil nutrient content under low temperature

[0129] Depend on Fig.13 It can be seen that low temperature caused a decrease in the content of available nitrogen, available phosphorus and organic matter in the soil, and had no significant effect on the content of available potassium. The content of available nitrogen decreased significantly with the extension of low temperature treatment time, decreasing by 4.8%, 7.4%, 8.8% and 8.4% respectively. The content of available phosphorus in the soil was significantly lower than that in CK at the 3rd, 6th and 12th day of treatment. The content of soil organic matter was significantly different only at the 12th day of treatment, which decreased significantly by 13.2%. Compared with the L treatment, the content of available nitrogen in the soil of LG treatment increased significantly at each treatment time, increasing by 3.7%, 4.8%, 4.8% and 6.1% respectively, while the content of available potassium decreased significantly, decreasing by 10.7%, 17.8%, 8.8% and 15.5% respectively. The content of available phosphorus decreased significantly at the 3rd, 9th and 12th day of treatment, decreasing by 10.2%, 14.8% and 16.1% respectively.

[0130] 11. Effects of glycine treatment on the content of different forms of soil nitrogen under low temperature

[0131] Depend on Fig.14 It can be seen from A that NO3 - -N content decreased significantly with the extension of low temperature treatment time. Compared with the L treatment, the NO3 - -N content increased significantly, by 6.3% and 10.3% respectively. Fig.14 It can be seen from B that compared with CK, the soil NH4 + -N content decreased significantly. Compared with the L treatment, the LG treatment had soil NH4 + -N content further decreased by 6.2%, 6.5%, 20.2% and 15.9% respectively. Fig.14 From the C, we can see that the soil microbial biomass nitrogen (SMBN) content of L treatment was significantly different from that of CK only at the 12th day of treatment, which was significantly reduced by 38.2%. Compared with L treatment, the SMBN content of LG treatment was significantly higher than that of L treatment at each treatment time point.

[0132] 12. Effects of glycine treatment on soil organic nitrogen components under low temperature

[0133] like Fig.15It can be seen that the L treatment significantly reduced the content of soil acid-hydrolyzable total nitrogen on the 12th day; on the 9th day of treatment, the content of soil acid-hydrolyzable amino acid nitrogen was significantly reduced compared with CK, by 59.2%; the L treatment did not have a significant effect on the content of soil acid-hydrolyzable amino acid nitrogen and acid-hydrolyzed amino sugar nitrogen. Compared with the L treatment, the content of each component of soil organic nitrogen in the LG treatment increased.

[0134] 13. Effect of glycine treatment on soil enzyme activity under low temperature

[0135] like Fig.16 As shown in A, low temperature significantly inhibited soil urease activity and showed a downward trend as the low temperature treatment time prolonged. Compared with the L treatment, the LG treatment significantly increased soil urease activity in each treatment period, with increases of 14.0%, 51.7%, and 25.3%, respectively, except for the third day of treatment. Fig.16 As shown in Figure B, compared with CK, L treatment significantly inhibited soil asparaginase activity at 3d and 12d, and its activity decreased the most at 12d, significantly decreasing by 17.4%. Compared with L treatment, LG treatment increased soil asparaginase activity at 3d and 12d, significantly increasing by 15.6% and 17.0%, respectively. Fig.16 As shown in Figure C, the soil glutaminase activity under the L treatment was significantly lower than that under the CK treatment at the 12th day, decreasing by 19.2%. Compared with the L treatment, the soil glutaminase activity under the LG treatment increased significantly at the 12th day, increasing by 25.0%. Fig.16 As shown in D, the soil nitrate reductase activity under L treatment was significantly lower than that under CK at 6d and 12d, decreasing by 17.1% and 13.9% respectively. Compared with L treatment, the soil nitrate reductase activity under LG treatment increased significantly at 12d, increasing by 22.2%.

[0136] In summary, low temperature reduces the photosynthetic pigment content of the leaves of the seedlings of the mountain yam, inhibits the photosynthetic and fluorescence properties of the leaves, inhibits the growth of the root system and the aboveground part of the plant, and reduces the absorption and assimilation of nitrogen. Low temperature also reduces the content of available nitrogen and available phosphorus in the soil, and reduces the activity of soil nitrogen conversion-related enzymes, which leads to varying degrees of decrease in the content of nitrate nitrogen, ammonium nitrogen and microbial biomass nitrogen in the soil. The present invention can alleviate the inhibitory effect of low temperature on the growth of the mountain yam seedlings by applying 6mM glycine, which is manifested as increasing the total root length, root surface area and root volume of the seedlings, promoting the accumulation of plant dry matter, improving the photosynthetic fluorescence index of the mountain yam seedlings, significantly increasing the pigment content in the leaves, and promoting its photosynthesis. It enhances the activity of nitrogen metabolism-related enzymes, increases the expression of nitrogen absorption and assimilation-related genes, and significantly increases the content of glutamate and glutamine in the root system. At the same time, the activity of soil nutrient conversion-related enzymes increases, which improves the conversion and supply capacity of nitrogen in the soil. Therefore, the present invention improves the resistance of the mountain yam seedlings to low temperature stress by applying glycine.

[0137] Although preferred embodiments of the present invention have been described, additional changes and modifications may occur to these embodiments once those skilled in the art are aware of the basic inventive concepts.

[0138] Obviously, those skilled in the art may make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. The application of glycine in resisting low temperature stress of apple trees is characterized by: The resistance of apple trees to low temperature stress was improved by applying glycine aqueous solution to apple seedlings.

2. The use of glycine in resisting low temperature stress of apple trees according to claim 1, characterized in that: The method of improving the resistance of apple trees to low temperature stress is manifested in: increasing the biomass of apple trees under low temperature stress, increasing the content of photosynthetic pigments in apple tree leaves under low temperature stress, increasing the photosynthetic parameters of apple tree leaves under low temperature stress, promoting the growth and development of the root system of apple tree seedlings under low temperature stress, enhancing the activity of nitrogen metabolism-related enzymes and improving the expression of nitrogen absorption and assimilation-related genes.

3. The use of glycine in resisting low temperature stress of apple trees according to claim 2, characterized in that: Improving the biomass of apple trees under low temperature stress is manifested in: increasing the aboveground biomass, total biomass and crown-root ratio of apple tree seedlings.

4. The use of glycine in resisting low temperature stress of apple trees according to claim 2, characterized in that: Improving the content of photosynthetic pigments in apple tree leaves under low temperature stress is manifested as: increasing the content of chlorophyll a, chlorophyll b and carotenoids in apple tree leaves under low temperature stress.

5. The use of glycine in resisting low temperature stress of apple trees according to claim 2, characterized in that: Improving the photosynthetic parameters of apple tree leaves under low temperature stress is manifested as: improving the net photosynthetic rate, transpiration rate, stomatal conductance and water use efficiency of apple tree leaves under low temperature stress.

6. The use of glycine in resisting low temperature stress of apple trees according to claim 2, characterized in that: Promoting the root growth and development of apple seedlings under low temperature stress is manifested in: increasing the total root length and total surface area of ​​apple seedlings under low temperature stress, and improving the root activity of seedlings.

7. The use of glycine in resisting low temperature stress of apple trees according to claim 2, characterized in that: The enhancement of nitrogen metabolism-related enzyme activities is manifested by increasing nitrate reductase activity, glutamine synthetase activity, glutamate synthetase activity and glutamate dehydrogenase activity.

8. The use of glycine in resisting low temperature stress of apple trees according to claim 2, characterized in that: Assimilation-related genes include NRT1.1, NRT2.4, NRT2.7, AMT1.2, AMT1.5 and NADH-GOGAT.

9. The use of glycine in resisting low temperature stress of apple trees according to claim 1, characterized in that: The concentration of the glycine aqueous solution is 4 mM to 8 mM, and the application amount is 150 mL to 250 mL per plant.

10. The use of glycine in resisting low temperature stress of apple trees according to claim 9, characterized in that: The concentration of the glycine aqueous solution was 6 mM, and the amount applied was 200 mL per plant.

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