Device and method for labeling 13C isotope of pinus massoniana seedlings

Abstract

The invention discloses a device and a method for labeling 13C isotope of pinus massoniana seedlings, and relates to the technical field of forest seedling culture. In order to solve the problem that an existing in-situ marking device cannot accurately control key environmental factors influencing seedling growth, the following technical scheme is provided: the in-situ marking device comprises a marking chamber, a gas storage chamber and a reaction chamber, and the marking chamber comprises a first chamber, a second chamber and a third chamber which are separated from one another; a sensor assembly, a cooler and a humidifier are arranged on the side wall of the second chamber, the interior of the first chamber is in a vacuumized state, and 13CO2 gas generated in the reaction chamber is sequentially introduced into the gas storage chamber and the marking chamber for gas storage and marking reaction; pinus massoniana seedlings to be marked are arranged on the porous plate of the second chamber, and an alkaline solution is arranged in the third chamber. The sensor assembly, the cooler and the humidifier are in communication connection with the automatic control system. According to the method, the marking environment can be controlled more easily, normal growth of pinus massoniana seedlings can be marked more easily, and the marking efficiency is improved.

Description

Technical Field

[0001] The present invention relates to the technical field of forest seedling cultivation, and in particular to a method for growing Masson pine seedlings. 13 C isotope labeling device and method. Background Art

[0002] Masson pine is the main pioneer tree species for afforestation in southern mountainous areas of my country, with strong adaptability, rapid growth and high productivity. Against the backdrop of global climate change and the need for afforestation in difficult sites, in-depth research on the distribution and regulation of photosynthetic products in Masson pine seedlings, and 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 equipment and methods, it is difficult to accurately analyze the distribution law of photosynthetic products in Masson pine seedlings, which has become one of the "bottlenecks" in basic research on Masson pine seedlings and seedling quality control. 13 C stability isotope labeling is widely used in the study of carbon pool cycle in the atmosphere-plant-soil. It can be used to study the carbon dynamics of plant photosynthesis such as carbon assimilation, transportation, accumulation and reuse and the formation of assimilation capacity. However, the existing plant 13 There are many disadvantages of C isotope labeling devices, for example, they cannot control or cannot accurately control key environmental factors such as temperature, humidity and CO2 concentration that affect seedling growth. 13 C isotope labeling cannot accurately calculate the cost of expensive chemicals (Na2 13 CO3、Ba2 13 CO3, etc.) is usually achieved by adding excessive amounts of chemicals, which results in unnecessary waste. 13 C isotope labeling needs to be carried out in a closed environment and usually in clear weather. Therefore, the ambient temperature will rise rapidly during the labeling process, greatly affecting plant photosynthesis, and even causing the death of seedlings in severe cases. Existing technologies usually reduce the ambient temperature through water cooling or air cooling, but the effect is extremely limited. Summary of the invention

[0004] The object of the present invention is to provide a method for preparing a seedling of Masson pine 13 The invention discloses a device and method for C isotope labeling 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 invention to solve the above technical problems is as follows:

[0006] A kind of seedling for Masson pine 13A device for C isotope labeling, comprising: a labeling chamber, a gas storage chamber and a reaction chamber which are connected to each other, the labeling chamber comprising a first cube and a second cube arranged 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 porous plate arranged inside, a sensor assembly, a cooler and a humidifier are arranged on the side wall of the second chamber, and the tops of the first cube and the second cube are sealed by a sealing cover;

[0007] The first chamber is in a vacuum state, and the reaction chamber is generated. 13 CO2 gas will 13 CO2 gas is stored in the gas storage room. 13 CO2 gas will 13 CO2 gas is released into the second chamber; Masson pine seedlings to be marked are set on the porous plate, an alkaline solution is set in the third chamber, and the sensor assembly, cooler and humidifier are respectively connected to the automatic control system for communication.

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

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

[0010] Preferably, a liquid inlet channel is provided at a side end of the second chamber, the liquid inlet channel is communicated with the third chamber, and a liquid discharge channel is provided at the bottom of the third chamber.

[0011] Preferably, the sealing cover comprises hinges and fasteners respectively arranged at the side ends, and a plurality of silicone gaskets are arranged at the bottom of the sealing cover.

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

[0013] A method for improving the above-mentioned method for masson pine seedlings 13 The method for measuring the labeling efficiency of a C isotope labeled device comprises the following steps:

[0014] S1. Determine and optimize the volume of the labeling chamber: First, measure the space required for the Masson 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 without affecting the normal photosynthesis of the plants;

[0015] S2. Perform starvation treatment and prepare the labeling environment: Cultivate the Masson pine seedlings to be labeled in a low-concentration CO2 environment to improve labeling efficiency; fix the plants in the second chamber of the labeling room, add NaOH solution to an appropriate liquid level to completely absorb the original CO2 in the room, and use a vacuum pump to evacuate the first chamber to further ensure that there is no interfering CO2;

[0016] S3, accurately calculate and generate 13 CO2 gas: Based on the volume of the labeling chamber, the initial CO2 concentration requirement, the labeling time, and the CO2 consumption rate of the Masson pine seedlings, the required Na 13 CO3 mass; then, the calculated amount of Na 13 CO3 and excess sulfuric acid H2SO4 are added to the reaction chamber to generate 13 CO2 is guided to the gas storage room for temporary storage for subsequent use;

[0017] S4. Set up and start the automatic control system: configure the parameters of the automatic control system, including the upper and lower thresholds of CO2 concentration, temperature and humidity; after connecting the power supply, the system automatically adjusts according to the preset conditions, that is, when the CO2 concentration, temperature or humidity is detected to deviate from the preset range, the solenoid valve, cooler or humidifier is opened or closed respectively to maintain the stability of the marked indoor environment;

[0018] S5. Complete marking and detection 13 C abundance: After the labeling experiment, the labeling chamber was opened, the labeled Masson pine seedlings were transferred and the whole plant was sampled; after the samples were separated and processed, the isotope mass spectrometer was used to accurately determine the content and ratio of carbon isotopes in the plant components, and the labeling experiment was calculated based on the measurement results. 13 C labeling efficiency.

[0019] The Na added in step S3 13 The mass calculation formula of CO3 is:

[0020]

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

[0022]

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

[0024] 1. The marking chamber with an insulating chamber (first chamber) provided by the present invention is used to extract the air inside the first chamber through a vacuum pump connected to the first chamber, so that the pressure inside the first chamber becomes low, the number of molecules actively moving inside is reduced, and it is in a relatively vacuum state. Thereby reducing the thermal conductivity coefficient of the marking chamber as a whole, being able to isolate the internal and external environments through the first chamber in clear weather, reducing heat conduction, and slowing down the temperature rise inside the marking chamber. The ambient temperature is easier to control, which is more beneficial to the normal growth of the marked Masson pine seedlings. At the same time, the relative vacuum environment of the first chamber further improves the sealing of the marking chamber.

[0025] 2. The marking room with automatic control system provided by the present invention can realize automatic control of environmental values ​​(temperature, humidity and CO2 concentration) in the marking room by connecting components such as sensors, microcontrollers, solenoid valves, coolers, humidifiers and power supplies. For temperature control, although the heat insulation chamber effectively isolates the heat brought by heat conduction and convection, there is still a part of the heat through thermal radiation, and the heat insulation chamber cannot affect the heat brought by thermal radiation. Through the dual cooling mechanism of the heat insulation chamber and the cooler, the effect of temperature control can be better achieved.

[0026] 3. The Na required for the calculation reaction provided by the present invention 13 The formula for the mass of CO3 is calculated by labeling the volume of the chamber, the rate at which the Masson pine seedlings consume carbon dioxide, and the Na 13 Calculate the required Na2CO3 molecular weight, gas molar volume and other data 13 The quality of CO3 is improved by eliminating the addition of excessive Na 13 CO3 (usually 2-5 times the required amount) is used to ensure the accuracy of the experiment while avoiding unnecessary waste. In addition, the formula for calculating the labeling efficiency provided by the present invention is calculated by calculating the consumption of Masson pine seedlings. 13 C and fixed as photosynthetic products 13 The ratio of C can accurately evaluate the labeling efficiency of the experiment and provide a basis for subsequent analysis. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 The present invention is used for Masson pine seedlings 13 A schematic diagram of the structure of a C isotope labeling device;

[0028] Figure 2 It is the composition and connection method of the automatic control system;

[0029] Figure 3 It is the logic decision flow chart of the automatic control system;

[0030] Figures 1 to 3 The reference numerals shown in the figure represent respectively: 1-marking chamber, 11-first chamber, 111-support column, 112-vacuum pump interface, 12-second chamber, 121-porous plate, 122-liquid inlet channel, 13-third chamber, 131-liquid discharge 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 DESCRIPTION

[0031] The technical solution of the present invention is described clearly and completely below in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative work are within the scope of protection of the present invention.

[0032] Please refer to Figure 1-3 This embodiment provides a method for growing Masson pine seedlings. 13 C isotope labeling device and a method for labeling Masson pine seedlings 13 In order to achieve the above object, according to a first aspect of the present invention, a method for labeling Masson pine seedlings is provided. 13 The device for C isotope labeling comprises: 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 this embodiment, the marking chamber 1 is made of high-density polyethylene (HDPE) material that is heat-insulating, corrosion-resistant, and has a light transmittance greater than 95%. It is in the shape of a cube or a cuboid, and is divided into a first cube in the outer layer and a second cube in the inner layer. The marking chamber consists of three chambers, wherein the first chamber 11 is composed of a cavity in the middle of the inner and outer layers of the cube, and the five center surfaces of the inner and outer layers of the cube except the top are provided with cylindrical support columns 111 to maintain the stability of the marking chamber. The first chamber is provided with a vacuum pump interface 112, which is used to connect a vacuum pump to extract the air in the first chamber 11, forming a vacuum barrier to isolate the heat exchange between the inner and outer environments. The second chamber 12 in the upper layer of the second cube and the third chamber 13 in the lower layer are cavities located inside, separated by a porous plate 121 in the middle, the second chamber 12 is used to place the Masson pine seedlings to be marked, and the third chamber 13 is used to place an alkaline solution that absorbs carbon dioxide. A liquid inlet 122 leading from the top to the bottom is provided at the corner of the second chamber 12, which is used to add alkaline solution to the third chamber 13. A drain port 131 is provided at the bottom of the third chamber 13, which is used to discharge the alkaline solution in the third chamber 13. The tops of the inner and outer layers of the marking chamber 1 are both designed with an openable and closable sealing cover 14, one side of the sealing cover 14 is connected to the marking chamber 1 through a hinge 141, and the other three sides are equipped with buckles. A card slot matching the buckle is provided at the top of the marking chamber 1, and the buckle and the card slot constitute a fastener 142. Silicone gaskets 143 are installed at the positions where the sealing cover 14 contacts the top edge of the marking chamber 1 to ensure air tightness. In addition, a connection port leading to the second chamber 12 is provided on the side of the marking chamber 1.

[0034] The gas storage chamber 2 is made of polyethylene material and is cylindrical in shape. It is used to store the gas generated by the reaction. 13 CO2 gas. Connection ports are arranged on both sides of the gas storage chamber 2, one of which is connected to the connection port of the marking chamber 1 through a rubber hose.

[0035] The reaction chamber 3 is made of glass material and is in the shape of a wide-mouth bottle, which is used for chemical reactions to generate the required 13 CO2 gas. A connection port is provided on the side of the reaction chamber 3, which is connected to another connection port of the gas storage chamber 2 through a rubber hose. A rubber plug is provided on the top of the reaction chamber 3, which is plugged with the rubber plug after adding chemicals to ensure air tightness.

[0036] The sensor assembly 4 includes a temperature sensor, a humidity sensor and a CO2 concentration sensor, which are installed inside the second chamber 12 of the marking room 1 and are used to detect the temperature, humidity and CO2 concentration in the marking room.

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

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

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

[0040] The automatic control system uses a microcontroller to design the control circuit, including hardware connection and software programming. The hardware connection is: connecting the temperature, humidity and CO2 concentration sensors to the analog input port of the microcontroller to read the temperature, humidity and CO2 concentration; connecting the solenoid valve 5, humidifier 7 and cooler 6 to the digital output port of the microcontroller to control them to open or close. The software programming is: writing logic to read the sensor data and control the opening or closing of the solenoid valve 5, humidifier 7 and cooler 6 according to the preset value. Integrate the microcontroller into the entire automatic control system to ensure that it can work with the solenoid valve 5, humidifier 7, sensor assembly 4, cooler 6 and other components.

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

[0042] S1. Determine the size of the labeling chamber: Under the same concentration requirement, the smaller the labeling chamber volume, the more 13 Therefore, under the premise of meeting the experimental requirements and not affecting the normal photosynthesis of the Masson pine seedlings to be marked, the volume of the marking chamber was reduced as much as possible.

[0043] S2. Starve the marked Masson pine seedlings: Place the marked plants in a low-CO2 concentration environment for a certain period of time to help improve13 C marking efficiency. Place the marking chamber on a flat surface, open the inner and outer sealing covers, place the plant to be marked in the second chamber and fix it; add NaOH solution to the liquid inlet to absorb CO2, and the added NaOH solution should not exceed the porous plate; close the inner and outer sealing covers, and use a vacuum pump to evacuate the first chamber to a vacuum state. After 30 minutes, open the drain port to drain the NaOH solution in the marking chamber, and close the drain port after all the NaOH solution has been drained.

[0044] S3. Calculate the Na required for labeling 13 The mass of CO3 and the generation 13 CO2 gas: Na 13 CO3 or Ba 13 CO3 is expensive, so its usage must be accurately calculated to avoid waste. 3 ), the initial carbon dioxide concentration required for the labeling test is a (ppm), the labeling time is t (h), and the carbon dioxide consumption rate of the Masson pine seedlings to be labeled is b (ppm / h), and the added Na 13 The mass of CO3 is:

[0045]

[0046] Add the corresponding mass of Na according to the calculation 13 CO3 and excess 1 mol.L -1 The H2SO4 solution is added to the reaction chamber to generate 13 The CO2 gas is guided by a connected rubber hose into the gas storage chamber for temporary storage.

[0047] S4. Set the parameters of the automatic control system and start marking: 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 solenoid valve. When the CO2 concentration is higher than the preset range, the control system automatically closes the solenoid valve. When the temperature is higher than the preset range, the control system automatically turns on the cooler. When the temperature is lower than the preset range, the control system automatically turns off the cooler. When the humidity is lower than the preset range, the control system automatically turns on the humidifier. When the humidity is higher than the preset range, the control system automatically turns on the humidifier.

[0048] S5. Sampling and testing of Masson pine seedlings to be marked 13 C abundance: After the labeling experiment is completed, open the labeling box and move the labeled seedlings to a place far away from the labeling box for sampling; sample the whole labeled seedlings, separate the samples according to the test requirements, weigh them separately, dry and grind them, and use an isotope mass spectrometer to determine the content and ratio of carbon isotopes in plant components. 13 C and 13 The actual loss of C, the labeling efficiency of the labeling experiment is calculated

[0049]

[0050] 1. Marking room size determination

[0051] The present invention is now described in detail using the carbon isotope labeling test applied to Masson pine seedlings. In this case, the test materials are derived from the seed garden of the National Masson pine Seed Base of Ma'anshan Forest Farm, Duyun City, Guizhou Province. Container seedlings were raised at the end of March 2021, and seedling raising and management were carried out in accordance with conventional methods. In September 2021, Masson pine seedlings with consistent growth were selected for testing. A total of 3 groups of seedlings with different numbers of plants were set up for the test, of which Group 1 had 9 plants, Group 2 had 18 plants, and Group 3 had 27 seedlings. The test cultivation pots have a diameter of 15 cm and a height of 18 cm. According to the diameter of the cultivation pots, it can be calculated that each Masson pine seedling occupies an area of ​​0.0225 m 2 For the 1st, 2nd and 3rd test groups, the marking chambers were designed with inner dimensions of 0.5x0.5x0.5m, 0.7x0.7x0.5m and 0.8x0.8x0.5m, and outer dimensions of 0.6x0.6x0.6m, 0.8x0.8x0.6m and 0.9x0.9x0.6m. The inner bottom areas were 0.25, 0.49 and 0.64m, respectively. 2 , slightly larger than the total area occupied by the seedlings to be marked (see Table 1), so that the seedlings to be marked do not block each other and the volume of the marking room is reduced as much as possible. In addition, the height of the test seedlings is between 18.73-26.71cm, so the height of the three sets of marking rooms is 0.5m, of which the lower layer is 0.1m high and the upper layer is 0.4m high, and the upper and lower layers are separated by a porous plate.

[0052] Table 1 Marking box scheme selection

[0053]

[0054] 2. Carbon isotope labeling

[0055] The weather on the day of marking was sunny, with a maximum temperature of 27 degrees and a minimum temperature of 16 degrees. The marking period was 10:00-14:00. Place the marking chamber on a stable surface, open the inner and outer sealing covers of the marking chamber, and evenly place the Masson pine seedlings to be marked on the porous plate of the second chamber. Add 1 mol·L -1 Add NaOH solution to the lower layer of the marking chamber, with the solution level below the porous plate. Close the inner and outer sealing covers, use a vacuum pump to remove the air in the first chamber to make it a relative vacuum state. Start timing, and allow the Masson pine seedlings to be marked to "starve" for 30 minutes. After the starvation reaction is over, open the drain port to drain all the NaOH solution, and close the drain port.

[0056] generate13 The chemical for CO2 is Na2 13 CO3( 13 C abundance>98%) and 1 mol·L -1 H2SO4 solution, the chemical reaction equation is Na2 13 CO3+H2SO4→Na2SO4+H2O+ 13 CO2. The Na2 required for the three labeling chambers is calculated according to the following formula 13 CO3:

[0057]

[0058] Where m is the required Na2 13 CO3 mass; a (ppm) is the starting point of the marker test 13 CO2 concentration; x(m 3 ) is the volume of the marking box; b (ppm / h) is the rate at which the plant consumes carbon dioxide; t (h) is the marking time; the coefficient 22.4 is the molar volume of the gas under standard conditions (24°C and 1atm) (22.4L / mol); the coefficient 106.99 is the Na2 13 The molecular weight of CO3 is (106.99 g / mol). 13 The initial concentration of CO2 was set to 1000 ppm; the volumes of the inner marking chambers of groups 1, 2, and 3 were 0.125 m 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 marking time of the three groups of experiments was 4h. As shown in Table 2, the Na2 13 CO3 were 0.755, 1.599 and 2.073 g respectively (see Table 2). Therefore, 0.755, 1.599 and 2.073 g of Na2 13 CO3 and excess 1 mol.L -1 of H2SO4.

[0059] Make sure all devices (sensors, microcontrollers, solenoid valves, coolers, humidifiers) are connected to power, start the microcontroller and load the pre-written control program. Set the preset range of CO2 concentration to 900-1000ppm; set the preset range of temperature to 22-27 degrees; set the preset range of humidity to 45%-55%. When the CO2 concentration is lower than 900ppm, the control system automatically opens the solenoid valve to add CO2 to the marking room. When the CO2 concentration is higher than 1000ppm, the control system automatically closes the solenoid valve and stops adding CO2; when the temperature is higher than 27 degrees, the control system automatically opens the cooler to reduce the temperature. When the temperature is lower than 22 degrees, the control system automatically turns off the cooler and stops cooling; when the humidity is lower than 45%, the control system automatically turns on the humidifier to increase the humidity. When the humidity is higher than 55%, the control system automatically turns off the humidifier and stops humidification. The control system continuously monitors the CO2 concentration, temperature and humidity, and automatically adjusts according to the actual situation. Record the changes in CO2 concentration, temperature and humidity for subsequent analysis. After the marking is completed, stop the control system manually or through program instructions.

[0060] Table 2 Na2 required for carbon isotope labeling experiment 13 CO3 Quality

[0061]

[0062] 3. Organs of Masson pine seedlings 13 The proportion of C and labeling efficiency

[0063] The marked Masson pine seedlings were taken out and continued to grow under the original light conditions. On the fourth day after the marking, the whole seedlings were dug up and sampled. The seedlings were fully collected according to the root, stem, leaf and other tissues. The samples were washed with clean water → deionized water → clean water → 1% hydrochloric acid → deionized water 3 times, then fixed at 105°C for 15 minutes, and then dried at 80°C to constant weight and weighed. After crushing, they were sieved with 100 mesh and mixed. The samples were measured by isotope mass spectrometer. 13 The abundance of C (δ 13 C) and 13 C content. The calculation formula is as follows:

[0064]

[0065] C i =T i ×W i

[0066]

[0067] Where: R S for 13 C / 12The absolute value ratio of C; R C (Carbon isotope standard ratio) = 0.0111802; F i (%)for 13 C atomic percentage; C i (g) is the total carbon content of each organ; W i (g) is the biomass of each organ; T i is the carbon content percentage of each organ; i is different organs; F n For unlabeled samples 13 C atomic percentage; P i (%) is the percentage of each organ 13 C distribution ratio; 13 C F Accumulation for all parts 13 As shown in Table 3, the root, stem and leaf 13 The concentrations of C were between 268.39–282.31‰, 192.78–212.68‰, and 305.33–350.10‰, respectively, indicating that the three size labeling schemes all had high labeling efficiencies. 13 The distribution of C in roots, stems and leaves ranged from 14.70% to 15.38%, 16.93% to 17.45% and 66.53% to 67.71%, respectively. These results indicate that the implementation of different sized labeling chambers will not affect the labeling effect of Masson pine seedlings.

[0068] In addition, the absorption of 13 C and 13 The actual loss ratio of C is used to evaluate 13 The labeling efficiency of C is calculated as follows:

[0069]

[0070] In the formula, P represents the labeling efficiency; 13 C F (g) is the absorbed 13 C total; y (h) is the duration of the labeling test; 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 of all schemes is 4 h. Substituting the above data into the formula, it can be calculated that the labeling efficiency of groups 1, 2 and 3 of the labeling experiment is 35.09%, 32.04% and 31.17%, respectively, which are improved compared with the 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 a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for masson pine seedlings 13 The device is labeled with C isotope, characterized in that include: A marking chamber (1), a gas storage chamber (2) and a reaction chamber (3) which are interconnected, the marking chamber (1) comprising a first cube and a second cube arranged inside the first cube, a first chamber (11) being formed between the first cube and the second cube, the second cube being divided into an upper second chamber (12) and a lower third chamber (13) by a porous plate (121) arranged inside, the side wall of the second chamber (12) being provided with a sensor assembly (4), a cooler (6) and a humidifier (7), and the tops of the first cube and the second cube are both sealed by a sealing cover (14); The interior of the first chamber (11) is in a vacuum state, and the reaction chamber (3) generates 13 CO2 gas will 13 CO2 gas is stored in the gas storage chamber (2), and the gas storage chamber (2) stores 13 CO2 gas will 13 CO2 gas is released into the second chamber (12); the Masson pine seedlings to be marked are placed on the porous plate (121); an alkaline solution is placed in the third chamber (12); and the sensor assembly (4), the cooler (6) and the humidifier (7) are respectively connected to an automatic control system for communication.

2. The method for preparing Masson pine seedlings according to claim 1. 13 The device is labeled with C isotope, characterized in that A plurality of support columns (111) are provided between the first cube and the second cube.

3. The method for preparing Masson pine seedlings according to claim 1 13 The device is labeled with C isotope, characterized in that A vacuum pump interface (112) is provided at the side end of the first cube.

4. The method for preparing Masson pine seedlings according to claim 1 13 The device is labeled with C isotope, characterized in that A liquid inlet channel (122) is provided at a side end of the second chamber (12), the liquid inlet channel (122) is communicated with the third chamber (13), and a liquid discharge channel (131) is provided at the bottom of the third chamber (13).

5. The method for preparing Masson pine seedlings according to claim 1 13 The device is labeled with C isotope, characterized in that The sealing cover (14) comprises hinges (141) and fasteners (142) respectively arranged at side ends, and a plurality of silicone gaskets (143) are arranged at the bottom of the sealing cover (14).

6. The method for preparing Masson pine seedlings according to claim 1 13 The device is labeled with C isotope, characterized in that A solenoid valve (5) is provided on the pipeline connecting the marking chamber (1) and the gas storage chamber (2).

7. A method for improving the use of a method for growing Masson pine seedlings as claimed in any one of claims 1 to 6 13 The method for determining the labeling efficiency of a C isotope-labeled device is characterized in that, The following steps are involved: S1. Determine and optimize the volume of the labeling chamber (1): first measure the space required for the Masson pine seedlings to be labeled, then design and adjust the size of the labeling chamber (1) to ensure that the volume of the labeling chamber (1) is minimized under a given concentration requirement while not affecting the normal photosynthesis of the plants; S2. Perform starvation treatment and prepare a labeling environment: place the Masson pine seedlings to be labeled in a low-concentration CO2 environment to improve labeling efficiency; fix the plants in the second chamber (12) of the labeling chamber (1), add NaOH solution to an appropriate liquid level to completely absorb the original CO2 in the chamber, and use a vacuum pump to evacuate the first chamber (11) to further ensure that there is no interfering CO2; S3, accurately calculate and generate 13 CO2 gas: Based on the volume of the labeling chamber (1), the initial CO2 concentration requirement, the labeling time, and the CO2 consumption rate of the Masson pine seedlings, the required Na 13 CO3 mass; then, the calculated amount of Na 13 CO3 and excess sulfuric acid H2SO4 are added to the reaction chamber (3) to generate 13 CO2 is directed to the gas storage chamber (2) for temporary storage for subsequent use; S4, setting and starting the automatic control system: configuring the parameters of the automatic control system, including the upper and lower thresholds of CO2 concentration, temperature and humidity; after connecting the power supply, the system automatically controls according to the preset conditions, that is, when it is detected that the CO2 concentration, temperature or humidity deviates from the preset range, the solenoid valve (5), the cooler (6) or the humidifier (7) are opened or closed respectively to maintain the stability of the environment in the marking room (1); S5. Complete marking and detection 13 C abundance: After the labeling experiment, the labeling chamber (1) was opened, the labeled Masson pine seedlings were transferred and the whole plant was sampled; after the samples were separated and processed, an isotope mass spectrometer was used to accurately determine the content and ratio of carbon isotopes in the plant components, and the labeling experiment was calculated based on the measurement results. 13 C labeling efficiency.

8. The method for improving the seedlings of Masson pine according to claim 7. 13 The method for C isotope labeling efficiency is characterized in that The Na added in step S3 13 The mass calculation formula of CO3 is: 。 9. The method for improving the seedlings of Masson pine according to claim 7 13 The method for C isotope labeling efficiency is characterized in that The formula for calculating the labeling efficiency of the labeling test in step S5 is: 。

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