A kind of for pinus massoniana seedling 13 Apparatus and method for isotopic labeling

Abstract

The application discloses a kind of for pinus massoniana seedlings 13 The application discloses a device and method for marking C isotope, and relates to the technical field of forest seedling raising.In order to solve the problem that the existing isotope marking device cannot accurately control the key environmental factors affecting the growth of seedlings, the following technical solution is proposed: it comprises a marking chamber, a gas storage chamber and a reaction chamber, the marking chamber comprises a first chamber, a second chamber and a third chamber separated from each other, the side wall of the second chamber is provided with a sensor assembly, a cooler and a humidifier, the inside of the first chamber is in a vacuum state, and CO2 gas is generated in the reaction chamber 13 The CO2 gas is sequentially introduced into the gas storage chamber and the marking chamber for gas storage and marking reaction; the pinus massoniana seedlings to be marked are arranged on the multi-well plate of the second chamber, and the third chamber is provided with an alkaline solution. The sensor assembly, the cooler and the humidifier are respectively communicatively connected with an automatic control system. The application can make the marking environment more controllable and more beneficial to the normal growth of the pinus massoniana seedlings, thereby improving the marking efficiency.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of forest seedling culture, and particularly relates to a device for marking C isotope of Pinus massoniana seedlings 13 Device and method for marking C isotope. BACKGROUND

[0002] Pinus massoniana is a main pioneer tree species for afforestation in mountainous areas in southern China, and has the characteristics of strong adaptability, rapid growth and high productivity. Under the background of global climate change and the demand for afforestation in difficult sites, in-depth research on the distribution and regulation of photosynthetic products of Pinus massoniana seedlings, accurate detection and directional control of the accumulation of photosynthetic products in different organs will be one of the important means to improve the quality of seedlings.

[0003] At present, due to the lack of scientific research experimental devices and methods, it is difficult to accurately analyze the distribution rule of photosynthetic products of Pinus massoniana seedlings, which has become one of the 'bottlenecks' of the basic research and quality regulation of Pinus massoniana seedlings. 13 C stable isotope-labeled plants are widely used in the research of carbon pool circulation in the atmosphere-plant-soil system, and can be used to study the carbon assimilation, transportation, accumulation and reuse of photosynthesis of plants, and the formation of assimilation capacity, but the existing plant 13 The device for marking C isotope has many shortcomings, for example, it cannot control or cannot accurately control the key environmental factors such as temperature, humidity and CO2 concentration that affect the growth of seedlings. And the method for marking C isotope of other plants 13 The method for marking C isotope cannot accurately calculate the mass of expensive chemicals (Na2 13 CO3, Ba2 13 CO3, etc.), which is usually added in excess, causing unnecessary waste. In addition, 13 The marking of C isotope needs to be carried out in a closed environment, and usually needs to be carried out in sunny weather, so the environmental temperature will rise rapidly during the marking process, which greatly affects the photosynthesis of plants, and even causes the death of seedlings in severe cases. The existing technology usually reduces the environmental temperature through water cooling or air cooling, but the effect is limited. SUMMARY

[0004] The purpose of the present application is to provide a device and method for marking C isotope of Pinus massoniana seedlings 13 to solve the problem that the existing isotope labeling device cannot accurately control the key environmental factors affecting the growth of seedlings.

[0005] The technical solution of the present application to solve the above technical problems is as follows:

[0006] A device for marking C isotope of Pinus massoniana seedlings 13A C isotope labeling device includes: a labeling chamber, a gas storage chamber, and a reaction chamber that are interconnected. The labeling chamber includes a first cube and a second cube disposed inside the first cube. A first chamber is formed between the first cube and the second cube. The second cube is divided into an upper second chamber and a lower third chamber by a perforated plate disposed inside. The side wall of the second chamber is provided with a sensor assembly, a cooler, and a humidifier. The tops of the first cube and the second cube are sealed by a sealing cap.

[0007] The first chamber is under vacuum, and the reaction chamber generates... 13 CO2 gas and 13 CO2 gas is stored in a gas storage chamber, and the gas storage chamber stores... 13 CO2 gas and 13 CO2 gas is released into the second chamber; pine seedlings to be labeled are placed on the porous plate; an alkaline solution is placed in the third chamber; and the sensor assembly, cooler, and humidifier are respectively connected to the automatic control system.

[0008] Preferably, multiple support columns are provided between the first cube and the second cube.

[0009] Preferably, a vacuum pump interface is provided on the side end of the first cube.

[0010] Preferably, the second chamber has a liquid inlet channel at its side end, which is connected to the third chamber, and the bottom of the third chamber has a liquid drain channel.

[0011] Preferably, the sealing cover includes hinges and fasteners respectively disposed on the side ends, and the bottom of the sealing cover is provided with multiple silicone gaskets.

[0012] Preferably, a solenoid valve is installed on the pipe connecting the marking chamber and the gas storage chamber.

[0013] An improvement for the above-mentioned use in Masson pine seedlings 13 A method for improving the labeling efficiency of a C isotope labeling device includes the following steps:

[0014] S1. Determine and optimize the volume of the labeling chamber: First, measure the space required for the pine seedlings to be labeled, then design and adjust the size of the labeling chamber to ensure that the volume of the labeling chamber is minimized under the given concentration requirements, while not affecting the normal photosynthesis of the plant;

[0015] S2. Starvation treatment and preparation of the labeling environment: Place the pine seedlings to be labeled in a low-concentration CO2 environment to improve the labeling efficiency; fix the plants in the second chamber of the labeling chamber, add NaOH solution to the appropriate level to completely absorb the original CO2 in the chamber, and use a vacuum pump to perform a vacuuming operation on the first chamber to further ensure the presence of no interfering CO2.

[0016] S3, accurately calculating and generating 13 CO2 gas: based on the volume of the labeling chamber, the initial CO2 concentration requirement, the labeling duration and the CO2 consumption rate of the Pinus massoniana seedlings, accurately calculating the required Na 13 CO3 mass; subsequently, the calculated amount of Na 13 CO3 is added into the reaction chamber with excess sulfuric acid H2SO4, and the reaction generates CO2 and is guided into the gas storage chamber for temporary storage for subsequent use; 13

[0017] S4, setting and starting the automatic control system: configuring the automatic control system parameters, including the upper and lower threshold values of the CO2 concentration, temperature and humidity; after connecting the power supply, the system automatically controls according to the preset conditions, that is, when the CO2 concentration, temperature or humidity deviates from the preset range, the operations of opening or closing the electromagnetic valve, the cooler or the humidifier are respectively performed to maintain the stability of the environment in the labeling chamber;

[0018] S5, completing labeling and detection 13 C abundance: after the labeling experiment is completed, the labeling chamber is opened, the labeled Pinus massoniana seedlings are transferred and whole-plant sampling is performed; after the samples are separated and treated, the content and proportion of carbon isotopes in the plant components are accurately measured using an isotope mass spectrometer, and the labeling efficiency of the labeling experiment is calculated according to the measurement results. 13

[0019] The mass of Na 13 CO3 added in step S3 is calculated according to the following formula:

[0020]

[0021] The formula for calculating the labeling efficiency of the labeling experiment in step S5 is:

[0022]

[0023] The present application has the following beneficial effects:

[0024] 1. The labeling chamber with a heat insulation chamber (first chamber) provided by the present application, through the vacuum pump connected to the first chamber, the air inside the first chamber is extracted, so that the pressure inside the first chamber is low, the number of molecules moving actively inside is reduced, and the state is in a relative vacuum. Thus, the overall heat conduction coefficient of the labeling chamber is reduced, the first chamber can isolate the internal and external environments in sunny weather, reduce heat conduction, and slow down the temperature rise inside the labeling chamber. The environmental temperature is easier to control, which is more beneficial to the normal growth of the Pinus massoniana seedlings. At the same time, through the relative vacuum environment of the first chamber, the sealing performance of the labeling chamber is further improved.

[0025] ​​2. The marking chamber with an automatic control system provided by this invention, through the connection of components such as sensors, microcontrollers, solenoid valves, coolers, humidifiers, and power supplies, can achieve automated control of the environmental values ​​(temperature, humidity, and CO2 concentration) within the marking chamber. Regarding temperature control, although the insulated chamber effectively isolates heat from conduction and convection, some heat still occurs through thermal radiation, and the insulated chamber cannot affect the heat generated by thermal radiation. Through the dual cooling mechanism of the insulated chamber and the cooler, a better temperature control effect can be achieved.

[0026] 3. The Na required for the calculation reaction provided by this invention 13 The formula for CO3 mass is derived by considering the volume of the labeled chamber, the rate at which the Masson pine seedlings consume carbon dioxide, and Na+. 13 The Na required for calculating CO3 molecular weight, gas molar volume, and other data. 13 The quality of CO3 was improved by eliminating the need for adding excessive Na in traditional methods. 13 Using CO3 (typically 2-5 times the required amount) ensures accurate experimentation while avoiding unnecessary waste. Furthermore, the formula for calculating labeling efficiency provided by this invention calculates the consumption of CO3 by pine seedlings. 13 C and fixed in the form of photosynthetic products 13 The ratio of C can accurately assess the labeling efficiency of the experiment, providing a basis for subsequent analysis. Attached Figure Description

[0027] Fig. 1 This invention is for use in Masson pine seedlings 13 A schematic diagram of the structure of a C isotope labeling device;

[0028] Fig. 2 It refers to the composition and connection methods of an automatic control system;

[0029] Fig. 3 It is a flowchart of the logic decision-making process of an automatic control system;

[0030] Figs. 1-3 The reference numerals in the accompanying drawings are as follows: 1-marking chamber, 11-first chamber, 111-support column, 112-vacuum pump interface, 12-second chamber, 121-perforated plate, 122-liquid inlet channel, 13-third chamber, 131-liquid outlet channel, 14-sealing cover, 141-hinge, 142-fastener, 143-silicone gasket; 2-gas storage chamber; 3-reaction chamber; 4-sensor assembly; 5-solenoid valve; 6-cooler; 7-humidifier. Detailed Implementation

[0031] The technical solutions of the present application are clearly and completely described below with reference to the drawings. Obviously, the described embodiments are only some of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative effort belong to the scope of the present application.

[0032] Please refer to Figs. 1-3 , the embodiment provides a device for C isotope labeling of Pinus massoniana seedlings 13 , and a method for C isotope labeling of Pinus massoniana seedlings 13 . In order to achieve the above-mentioned purpose, according to the first aspect of the present application, a device for C isotope labeling of Pinus massoniana seedlings is provided 13 , comprising a labeling chamber 1, a gas storage chamber 2, a reaction chamber 3, a sensor assembly 4, a solenoid valve 5, a cooler 6, a humidifier 7 and an automatic control system.

[0033] In the embodiment, the labeling chamber 1 is made of high-density polyethylene (HDPE) material with heat insulation, corrosion resistance and light transmittance greater than 95%, and has a shape of a cube or a cuboid, and is divided into a first cube of an outer layer and a second cube of an inner layer. The labeling chamber is composed of three cavities, wherein the first cavity 11 is composed of the cavity between the inner and outer cubes, and the inner and outer cubes are provided with cylindrical support columns 111 on five central surfaces except the top to maintain the stability of the labeling chamber. The first cavity is provided with a vacuum pump interface 112 for connecting a vacuum pump to exhaust the air in the first cavity 11, form a vacuum layer and isolate the heat exchange between the inner and outer environments. The second cavity 12 in the upper layer and the third cavity 13 in the lower layer of the second cube are internal cavities separated by a porous plate 121, the second cavity 12 is used for placing Pinus massoniana seedlings to be labeled, and the third cavity 13 is used for placing an alkaline solution for absorbing carbon dioxide. The second cavity 12 is provided with a liquid inlet 122 leading from the top to the bottom at the corner, for adding the alkaline solution to the third cavity 13. The third cavity 13 is provided with a liquid outlet 131 at the bottom for discharging the alkaline solution in the third cavity 13. The top of the inner and outer layers of the labeling chamber 1 is designed with a sealable cover 14, one side of the cover 14 is connected with the labeling chamber 1 through a hinge 141, and the other three sides are provided with buckles. The top of the labeling chamber 1 is provided with a clamping groove matched with the buckle, and the buckle and the clamping groove constitute a fastener 142. The position where the cover 14 contacts with the edge of the top of the labeling chamber 1 is provided with a silica gel gasket 143, to ensure the air tightness. In addition, the side of the labeling chamber 1 is provided with a connecting port leading to the second cavity 12.

[0034] The gas storage chamber 2 is made of polyethylene material and has a shape of a cylinder, and is used for storing the 13 CO2 gas generated by the reaction. The gas storage chamber 2 is provided with connecting ports on two sides, one side of which is connected with the connecting port of the labeling chamber 1 through a rubber hose.

[0035] Reaction chamber 3 is made of glass and is shaped like a wide-mouthed bottle, used for chemical reactions to produce the necessary components. 13 CO2 gas. A connection port is located on the side of reaction chamber 3, which connects to another connection port of gas storage chamber 2 via a rubber hose. A rubber stopper is installed on the top of reaction chamber 3; after adding chemicals, the stopper is inserted to ensure airtightness.

[0036] The sensor assembly 4 includes a temperature sensor, a humidity sensor, and a CO2 concentration sensor, and is installed inside the second chamber 12 of the labeling chamber 1 to detect the temperature, humidity, and CO2 concentration inside the labeling chamber.

[0037] Solenoid valve 5 is a one-way solenoid valve, installed between the connecting hoses of gas storage chamber 2 and marking chamber 1. The arrow of the one-way solenoid valve points towards the marking chamber, ensuring that gas can only flow from gas storage chamber 2 to marking chamber 1.

[0038] Cooler 6 is a semiconductor device based on the Peltier effect, installed on the inner wall of the second chamber 12 of the marking chamber 1, and is used to control the temperature inside the marking chamber 1.

[0039] The humidifier 7 is installed inside the second chamber 12 of the marking chamber 1 to control the humidity inside the marking chamber 1.

[0040] The automatic control system employs a microcontroller-based control circuit, encompassing both hardware connectivity and software programming. Hardware connectivity involves connecting temperature, humidity, and CO2 concentration sensors to the microcontroller's analog input ports to read these values; and connecting solenoid valve 5, humidifier 7, and cooler 6 to the microcontroller's digital output ports to control their opening and closing. Software programming involves writing logic to read sensor data and control the opening and closing of solenoid valve 5, humidifier 7, and cooler 6 based on preset values. Integrating the microcontroller into the entire automatic control system ensures its seamless integration with components such as solenoid valve 5, humidifier 7, sensor assembly 4, and cooler 6.

[0041] This invention also provides a method for improving the growth of Masson pine seedlings. 13 The method for improving C isotope labeling efficiency includes the following steps:

[0042] S1. Determine the size of the labeling chamber: Under the same concentration requirement, the smaller the volume of the labeling chamber, the less space is needed. 13 The lower the CO2 content, the better. Therefore, while meeting the experimental requirements and not affecting the normal photosynthesis of the pine seedlings to be labeled, the volume of the labeling chamber should be minimized as much as possible.

[0043] S2. "Starvation treatment" for the pine seedlings to be marked: Placing the plants to be marked in a low-concentration CO2 environment for a certain period of time helps to improve...13 The labeling efficiency of C. Place the labeling chamber on a flat surface, open the inner and outer sealing covers, place the plant to be labeled in the second chamber and fix it; add NaOH solution to the liquid inlet to absorb CO2, and the amount of NaOH solution added should not exceed the height of the multi-well plate; close the inner and outer sealing covers, and use a vacuum pump to draw the first chamber to a vacuum state. After 30 minutes, open the liquid outlet to discharge the NaOH solution in the labeling chamber, and close the liquid outlet after the NaOH solution is completely discharged.

[0044] S3. Calculate the amount of Na 13 CO3 required for labeling and generate 13 CO2 gas: Na 13 CO3 or Ba 13 CO3 is expensive, so its amount needs to be accurately calculated to avoid waste. According to the volume of the labeling chamber, which is x (m 3 ), the required initial carbon dioxide concentration a (ppm) for the labeling test, the labeling duration t (h), and the carbon dioxide consumption rate b (ppm / h) of the Pinus massoniana seedlings to be labeled, the amount of Na 13 CO3 added is:

[0045]

[0046] According to the calculation, the corresponding amount of Na 13 CO3 and excess 1 mol / L -1 H2SO4 solution is added to the reaction chamber to generate 13 CO2 gas, which is guided to the gas storage chamber through the connected rubber hose for temporary storage.

[0047] S4. Set the parameters of the automatic control system and start labeling: turn on the automatic control system and connect the power supply, when the CO2 concentration is lower than the preset range, the control system automatically opens the electromagnetic valve, when the CO2 concentration is higher than the preset range, the control system automatically closes the electromagnetic valve; when the temperature is higher than the preset range, the control system automatically opens the cooler, when the temperature is lower than the preset range, the control system automatically closes the cooler; when the humidity is lower than the preset range, the control system automatically opens the humidifier, when the humidity is higher than the preset range, the control system automatically closes the humidifier.

[0048] S5. Sampling and detection of Pinus massoniana seedlings to be labeled 13 C abundance: after the labeling test is completed, move the labeled seedlings away from the labeling chamber for sampling; sample the whole labeled seedlings, separate the samples according to the test requirements, weigh and dry them, grind them, and use an isotope mass spectrometer to measure the content and proportion of carbon isotopes in plant components. According to the actual loss of 13 C and 13 C absorbed by the labeled Pinus massoniana seedlings, calculate the labeling efficiency of the labeling test

[0049]

[0050] 1. Marker chamber size

[0051] The present application is described in detail by applying it to a carbon isotope labeling test of Pinus massoniana seedlings. In this case, the test material comes from the seed orchard of the national Pinus massoniana breeding base in Ma'anshan Forest Farm, Duyun City, Guizhou Province. Container seedlings were carried out at the end of March 2021, and seedling raising and management were carried out according to the conventional method. In September 2021, Pinus massoniana seedlings with consistent growth vigor were selected for the test. A total of 3 groups of seedlings with different plant numbers were set up for the test, including 9 seedlings in group 1, 18 seedlings in group 2, and 27 seedlings in group 3. The test cultivation pots are 15 cm in diameter and 18 cm in height. According to the diameter of the cultivation pot, the area occupied by each Pinus massoniana seedling is calculated to be 0.0225 m 2 , and the inner layer size of 0.5x0.5x0.5m, 0.7x0.7x0.5m and 0.8x0.8x0.5m, and the outer layer size of 0.6x0.6x0.6m, 0.8x0.8x0.6m and 0.9x0.9x0.6m are set up for the test, respectively. The inner layer bottom area is 0.25, 0.49 and 0.64 m 2 , respectively, which is slightly larger than the total area occupied by the Pinus massoniana seedlings to be labeled (see Table 1), so that the seedlings to be labeled do not block each other while minimizing the volume of the labeling chamber. In addition, the height of the test seedlings is between 18.73-26.71 cm, so the height of the three labeling chambers is 0.5 m, of which the lower layer is 0.1 m high and the upper layer is 0.4 m high. The two layers are separated by a multi-well plate.

[0052] Table 1 Selection of labeling box scheme

[0053]

[0054] 2. Carbon isotope labeling

[0055] On the labeling day, the weather was sunny, the highest temperature was 27 degrees, and the lowest temperature was 16 degrees. The labeling time period was 10:00-14:00. The labeling chamber was placed on a stable surface, the inner and outer layer sealing covers were opened, and the Pinus massoniana seedlings to be labeled were evenly placed on the multi-well plate in the second cavity. 1 mol·L -1 of NaOH solution was added to the lower layer of the labeling chamber, and the solution surface was below the multi-well plate. Close the inner and outer layer sealing covers, use a vacuum pump to remove the air in the first cavity to make it a relative vacuum state. Start timing and make the Pinus massoniana seedlings to be labeled "hunger reaction" for 30 min. After the hunger reaction is over, open the drain port to drain all the NaOH solution, and close the drain port.

[0056] Generated13 The chemical reagent for CO2 is Na2. 13 CO3 13 C abundance > 98% and 1 mol·L⁻¹ -1 The chemical reaction equation for the H2SO4 solution is Na2 13 CO3 + H2SO4 → Na2SO4 + H2O 13 CO2. Calculate the required Na2 for the three labeling chambers using the following formula. 13 CO3:

[0057]

[0058] In the formula, m is the required Na2 13 CO3 mass; a (ppm) is the starting point of the marker test. 13 CO2 concentration; x(m 3 ) represents the volume of the labeling box; b (ppm / h) represents the rate at which the plant consumes carbon dioxide; t (h) represents the labeling time; the coefficient 22.4 represents the molar volume of the gas under standard conditions (24℃ and 1 atm) (22.4 L / mol); the coefficient 106.99 represents Na2 13 The molecular weight of CO3 is 106.99 g / mol. This example includes three sets of experiments. 13 The initial CO2 concentration was set to 1000 ppm; the volumes of the inner labeling chambers in groups 1, 2, and 3 were 0.125 m³ and 0.125 m³, respectively. 3 0.245m 3 and 0.32m 3 According to the data displayed by the CO2 concentration sensor, the CO2 consumption rates in the three groups of experiments were 97.3, 91.7, and 89.1 ppm / h, respectively; the labeling time for all three groups of experiments was 4 hours. Table 2 shows the required Na2 in groups 1, 2, and 3. 13 The CO3 concentrations were 0.755, 1.599, and 2.073 g, respectively (see Table 2). Therefore, 0.755, 1.599, and 2.073 g of Na2 were added to the reaction chambers of groups 1, 2, and 3, respectively. 13 CO3 and an excess of 1 mol.L -1 H2SO4.

[0059] Ensure all equipment (sensors, microcontroller, solenoid valves, cooler, humidifier) ​​is connected to power. Start the microcontroller and load the pre-written control program. Set the preset range for CO2 concentration to 900-1000 ppm; the preset range for temperature to 22-27 degrees Celsius; and the preset range for humidity to 45%-55%. When the CO2 concentration is below 900 ppm, the control system automatically opens the solenoid valve to replenish CO2 into the marking chamber. When the CO2 concentration is above 1000 ppm, the control system automatically closes the solenoid valve to stop replenishing CO2. When the temperature is above 27 degrees Celsius, the control system automatically opens the cooler to lower the temperature. When the temperature is below 22 degrees Celsius, the control system automatically closes the cooler to stop cooling. When the humidity is below 45%, the control system automatically opens the humidifier to increase humidity. When the humidity is above 55%, the control system automatically closes the humidifier to stop humidifying. The control system continuously monitors CO2 concentration, temperature, and humidity, and automatically adjusts according to actual conditions. Record changes in CO2 concentration, temperature, and humidity for subsequent analysis. After marking is complete, manually or via program command stop the control system.

[0060] Table 2. Na2 required for carbon isotope labeling experiments. 13 CO3 mass

[0061]

[0062] 3. In various organs of Masson pine seedlings 13 C's proportion and labeling efficiency

[0063] The marked Masson pine seedlings were removed and allowed to continue growing under the original light conditions. On the fourth day after marking, the seedlings were dug up and sampled as a whole, including roots, stems, and leaves. The samples were rinsed with a mixture of water, deionized water, water, 1% hydrochloric acid, and three times with deionized water. The samples were then blanched at 105℃ for 15 minutes, dried at 80℃ to constant weight, and weighed. After pulverizing, the samples were passed through a 100-mesh sieve and mixed. The concentrations of various compounds in the samples were then measured using an isotope mass spectrometer. 13 C abundance (δ) 13 C) and 13 C content. The calculation formula is as follows:

[0064]

[0065] C i =T i ×W i

[0066]

[0067] In the formula: R S for 13 C / 12The ratio of the absolute values ​​of C; R C (Carbon isotope standard ratio) = 0.0111802; F i (%)for 13 C atomic percentage; C i (g) represents the total carbon content of each organ; W i (g) represents the biomass of each organ; T i The percentage of carbon in each organ; i represents different organs; F n For unlabeled samples 13 C atomic percentage; P i (%) represents the percentages of each organ 13 C allocation ratio; 13 C F Accumulate for all parts 13 The sum of C. Table 3 shows that in the three groups of experiments, the roots, stems, and leaves of *Pinus massoniana* seedlings... 13 The concentrations of C were between 268.39–282.31‰, 192.78–212.68‰, and 305.33–350.10‰, respectively, indicating that all three labeling schemes had high labeling efficiency. 13 The proportions of carbon in roots, stems, and leaves ranged from 14.70–15.38%, 16.93–17.45%, and 66.53–67.71%, respectively. These results indicate that implementing different sized labeling chamber schemes does not affect the labeling effect on Masson pine seedlings.

[0068] In addition, it can also be based on the absorption of marker seedlings. 13 C and 13 The ratio of C to actual loss is used for evaluation. 13 The labeling efficiency of C. The calculation formula is as follows:

[0069]

[0070] In the formula, P represents the labeling efficiency; 13 C F (g) represents the absorption in all marked seedlings. 13 Ctotal; y(h) is the duration of the labeling experiment; b(ppm / h) is the carbon dioxide consumption of the labeled seedlings; the coefficient 12.99 is... 13 The mass fraction of C is 12.99 g / mol. In this embodiment, the labeling time for all schemes is 4 hours. Substituting the above data into the formula, the labeling efficiencies of the labeling experiments in groups 1, 2, and 3 can be calculated to be 35.09%, 32.04%, and 31.17%, respectively, which are all improved compared with existing labeling methods.

[0071] Table 3 δ in different schemes 13 C concentration and 13 Distribution ratio of C in various organs of Masson pine seedlings

[0072]

[0073] Note: Values represent mean ± standard error.

[0074] The above description is only the preferred embodiment of the application, and is not intended to limit the application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the application shall be included in the protection scope of the application.

Claims

1. A method for improving the survival of Pinus massoniana seedlings 13 A method for the determination of the isotopic labeling efficiency, characterized in that, The utility model relates to a kind of marking chamber, gas storage chamber and reaction chamber, including: marking chamber (1), gas storage chamber (2) and reaction chamber (3) are communicated with each other, the marking chamber (1) includes first cube and second cube being arranged inside first cube, first chamber (11) is formed between the first cube and the second cube, the second cube is divided into second chamber (12) of upper layer and third chamber (13) of lower layer by porous plate (121) being arranged inside, the side wall of second chamber (12) is equipped with sensor assembly (4), cooler (6) and humidifier (7), the top of the first cube and the second cube is sealed by sealing cover (14); The utility model relates to a kind of marking chamber, gas storage chamber and reaction chamber, including: marking chamber (1), gas storage chamber (2) and reaction chamber (3) are communicated with each other, the marking chamber (1) includes first cube and second cube being arranged inside first cube, first chamber (11) is formed between the first cube and the second cube, the second cube is divided into second chamber (12) of upper layer and third chamber (13) of lower layer by porous plate (121) being arranged inside, the side wall of second chamber (12) is equipped with sensor assembly (4), cooler (6) and humidifier (7), the top of the first cube and the second cube is sealed by sealing cover (14); The first chamber (11) is in a vacuum state, and CO2 gas is generated in the reaction chamber (3) 13 CO2 gas is stored in the gas storage chamber (2) 13 CO2 gas is stored in the gas storage chamber (2) 13 CO2 gas is stored in the gas storage chamber (2) 13 CO2 gas is released into the second chamber (12); the porous plate (121) is provided with the seedlings of Pinus massoniana to be marked, the third chamber (13) is provided with an alkaline solution, and the sensor assembly (4), the cooler (6) and the humidifier (7) are respectively connected with an automatic control system in communication; an electromagnetic valve (5) is arranged on a pipeline in communication between the marking chamber (1) and the gas storage chamber (2). S1, determine and optimize the volume of marking chamber (1): first measure the space required for the marked pinus massoniana seedlings, then design and adjust the size of the marking chamber (1) to ensure that the volume of the marking chamber (1) is minimized under the given concentration requirement without affecting the normal photosynthesis of the plants; S2, starvation treatment and preparation of the marking environment: the pinus massoniana seedlings to be marked are cultured in a low-concentration CO2 environment to improve the marking efficiency; the plants are fixed in the second chamber (12) of the marking chamber (1), NaOH solution is added to the appropriate liquid level to completely absorb the original CO2 in the chamber, and a vacuum pump is used to perform vacuum pumping on the first chamber (11) to further ensure the presence of undisturbed CO2; S4, set and start the automatic control system: configure the parameters of the automatic control system, including the upper and lower threshold values of CO2 concentration, temperature and humidity; after connecting the power supply, the system automatically controls according to the preset conditions, that is, when the CO2 concentration, temperature or humidity deviates from the preset range, the electromagnetic valve (5), cooler (6) or humidifier (7) is opened or closed respectively to maintain the stability of the environment in the marking chamber (1); S3, accurately calculate and generate 13 CO2 gas: based on the volume of the labeling chamber (1), the initial CO2 concentration requirement, the labeling duration, and the CO2 consumption rate of the Pinus massoniana seedlings, accurately calculate the required Na 13 CO3 mass; subsequently, add the calculated amount of Na 13 CO3 and excess sulfuric acid H2SO4 into the reaction chamber (3), react to generate 13 CO2, and guide into the gas storage chamber (2) for temporary storage for subsequent use; A plurality of support columns (111) are arranged between the first cube and the second cube. S5, completion of labeling and detection 13 C abundance: after the end of the labeling experiment, open the labeling chamber (1), transfer the labeled Pinus massoniana seedlings and sample the whole plant; after separating and processing the sample, use the isotope mass spectrometer to accurately measure the content and proportion of carbon isotopes in plant components, and calculate the labeling efficiency of the labeling experiment according to the measurement results 13 C labeling efficiency.

2. The method for improving Pinus massoniana seedlings according to claim 1, wherein the seedlings are Pinus massoniana seedlings. 13 A method for the determination of the isotopic labeling efficiency, characterized in that, A vacuum pump interface (112) is arranged at the side end of the first cube.

3. The method for improving Pinus massoniana seedlings according to claim 1, wherein the seedling is a 1-year-old seedling. 13 A method for the determination of the isotopic labeling efficiency, characterized in that, A liquid inlet channel (122) is arranged at the side end of the second chamber (12), the liquid inlet channel (122) is communicated with the third chamber (13), and a liquid outlet channel (131) is arranged at the bottom of the third chamber (13).

4. The method for improving Pinus massoniana seedlings according to claim 1, wherein the seedling is a 1-year-old seedling. 13 A method for the determination of the isotopic labeling efficiency, characterized in that, The sealing cover (14) includes a hinge (141) and a fastener (142) arranged at the side end respectively, and a plurality of silica gel pads (143) are arranged at the bottom of the sealing cover (14).

5. The method for improving Pinus massoniana seedlings according to claim 1, wherein the seedling is a 1-year-old seedling. 13 A method for the determination of the isotopic labeling efficiency, characterized in that, ​

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