Device and method for determining distribution of plant carbon assimilate source library

Abstract

The invention relates to the technical field of isotope labeling, and provides a device and a method for measuring plant carbon assimilation substance source library distribution. The device for measuring the distribution of the plant carbon assimilation substance source library comprises a generator, a leaf chamber, a concentration monitor and a main controller, and the leaf chamber is connected with the generator; the concentration monitor is connected with the leaf chamber; the main controller is used for establishing a standard curve between the CO2 concentration and the 13CO2 mass fraction according to the CO2 concentration data detected by the concentration monitor and the mass fraction of the 13CO3-containing solution in the generator; according to the CO2 concentration detected in real time, the mass fraction of the current 13CO2 is obtained through the standard curve; and calculating the concentration value of 13CO2 based on the CO2 concentration and the mass fraction of 13CO2. According to the device for measuring the distribution of the plant carbon assimilate source library, high-precision and low-cost control on the concentration of the < 13 > CO2 labeled gas is realized under the condition of not depending on an expensive isotope analyzer.

Description

Technical Field

[0001] This invention relates to the field of isotope labeling technology, and in particular to an apparatus and method for determining the allocation of plant carbon assimilate source libraries. Background Technology

[0002] In plant physiology and metabolism research, stable isotope labeling techniques (such as...) 13 C-CO2 labeling is a key tool for elucidating the transport and distribution mechanisms of carbon assimilation products. For cash crops such as cotton, the opposite leaf of the boll is the core photosynthetic source organ, and its photosynthetic efficiency and the ability to transport assimilates to the boll directly determine the quality and yield of cotton fibers.

[0003] Currently, various leaf chamber systems have been used for plant photosynthesis measurement and isotope labeling. For example, Evans et al. designed a dynamic flow leaf chamber with temperature control, airflow homogenization, and rapid gas switching functions, mainly for Arabidopsis research; Kölling et al. developed a system for single leaves... 14 CO2 labeling device; Ćeranić et al. reported on achieving uniform labeling throughout the plant using a custom-designed growth chamber. 13 C and 15 Methods for marking N.

[0004] However, the aforementioned systems rely on external gas sources or simple reactions, lack effective control over gas concentration, flow rate, and distribution, have low labeling efficiency, and require expensive detection equipment, resulting in high costs for the leaf chamber system. There is a lack of existing technologies that can achieve both low cost and high accuracy. 13 A system that controls CO2 concentration and is applicable to a variety of plant organs, supporting high-throughput parallel experiments. Summary of the Invention

[0005] This invention provides an apparatus and method for determining the allocation of plant carbon assimilate source libraries, thereby overcoming the shortcomings of low labeling efficiency and expensive detection equipment in the prior art.

[0006] This invention provides an apparatus for determining the allocation of a plant carbon assimilate source pool, comprising: a generator for producing CO2, wherein the CO2 includes... 12 CO2 and 13 CO2; at least one assimilation unit, each assimilation unit forming a loop with the generator, each assimilation unit comprising: a leaf chamber having a cavity for containing plant tissue, the leaf chamber being connected to the generator; a concentration monitor connected to the leaf chamber for monitoring the CO2 concentration within the leaf chamber; and a main controller electrically connected to the concentration monitor for using the CO2 concentration data detected by the concentration monitor, combined with the CO2 concentration in the generator. 13The mass fraction of CO3 solution, establishing the relationship between CO2 concentration and... 13 A standard curve relating CO2 mass fractions; based on the real-time detected CO2 concentration, the current... 13 The mass fraction of CO2; based on the CO2 concentration and the 13 The mass fraction of CO2 was calculated. 13 CO2 concentration value.

[0007] According to the present invention, an apparatus for determining the allocation of a plant carbon assimilate source library is provided, wherein the leaf chamber comprises: a leaf chamber body having the cavity, the leaf chamber body having opposing first and second plates, the first plate having a first groove, the second plate having multiple pairs of air inlets and outlets, a generator connected to a pair of air inlets and outlets, and a concentration monitor connected to a pair of air inlets and outlets; and a cover covering the leaf chamber body and sealingly connected to it, the cover being a light-transmitting cover having a first and second through groove, the first through groove communicating with the first groove, the first groove for accommodating the plant petiole, the cavity for accommodating the plant leaf, and the first groove, the first through groove, and the second through groove being sealed with flexible sealant.

[0008] According to the present invention, an apparatus for determining the allocation of a plant carbon assimilate source pool is provided, wherein the leaf chamber further includes an elastic element disposed in the first groove, and the elastic element is provided with a second groove for accommodating the petiole of the plant.

[0009] According to the present invention, an apparatus for determining the allocation of plant carbon assimilate source pools is provided, wherein the second groove is provided with sealant.

[0010] According to the present invention, an apparatus for determining the allocation of plant carbon assimilate source pools is provided, wherein the leaf chamber further includes a flexible sealing membrane, and the bottom plate of the leaf chamber body is provided with a third through groove, and the flexible sealing membrane is disposed at the third through groove.

[0011] According to the present invention, an apparatus for determining the allocation of a plant carbon assimilate source library is provided, wherein the assimilation unit further includes a flow pump disposed in the circulation loop, the flow pump being used to regulate the flow rate of the circulating gas in the circulation loop, and the flow pump being electrically connected to the main controller.

[0012] According to the present invention, an apparatus for determining the allocation of a plant carbon assimilate source pool is provided, wherein the assimilation unit further includes a barometer disposed in the circulation loop, the barometer being used to detect the pressure of the circulation loop.

[0013] According to the present invention, an apparatus for determining the allocation of plant carbon assimilate source pools is provided, wherein the assimilation unit further includes a buffer controller, the buffer controller being disposed in the circulation loop, the buffer controller being used to reduce the difference between the environmental parameters in the leaf chamber and the external environmental parameters, and the buffer controller being electrically connected to the main controller.

[0014] The present invention also provides a method for determining the allocation of plant carbon assimilate source pools based on the apparatus described above, comprising: establishing a relationship between CO2 concentration and... 13 Standard curve of CO2 mass fraction; obtaining the first mass fraction of carbon in source organ dry matter and 13 The first isotopic percentage of C, the second mass fraction of carbon in the dry matter of the organoids, and 13 The percentage of the second isotope of C; the source organ and the stem organ are placed in the leaf chamber for photosynthesis, and the CO2 concentration in the leaf chamber is obtained. Based on the CO2 concentration, the standard curve is used to control... 13 The CO2 concentration was within the set range; the third mass fraction of carbon in the labeled source organ dry matter was obtained at different time points. 13 The percentage of the third isotope of C, the fourth mass fraction of carbon in the dry matter of the organ, and 13 The fourth isotope percentage of C; the carbon assimilation efficiency of the source organ is calculated based on the first mass fraction, the first isotope percentage, the third mass fraction, and the third isotope percentage; the carbon assimilation efficiency of the library organ is calculated based on the second mass fraction, the second isotope percentage, the fourth mass fraction, and the fourth isotope percentage.

[0015] According to the method provided by the present invention, the establishment of CO2 concentration and 13 The steps for establishing a standard curve between CO2 mass fractions include: keeping the total mass of the carbon isotope salt solution in the generator constant, and changing the concentration of CO2 in the solution. 13 The mass fraction of the CO3 solution was obtained. 13 The concentration of CO2 produced by a CO3 solution at different mass fractions; based on the reaction... 13 CO2 and containing 13 Based on the principle that the mass fraction of CO3 in the solution is equal, we obtain... 13 The mass fraction of CO2; 13 Using the mass fraction of CO2 as the x-axis and the concentration of CO2 as the y-axis, a curve was fitted to obtain the relationship between the CO2 concentration and the... 13 Standard curve of CO2 mass fraction.

[0016] The device for determining the allocation of plant carbon assimilate source libraries provided by this invention, by setting up a concentration monitor and a main controller, and by establishing a CO2 concentration standard curve, can obtain the carbon assimilate source library allocation based on the real-time detected CO2 concentration. 13 The mass fraction of CO2 can then be calculated from the CO2 gas content. 13 The concentration of CO2 was measured, enabling the determination of CO2 concentration without relying on expensive isotope analyzers. 13 High-precision, low-cost quantitative control of CO2 labeled gas concentration. Attached Figure Description

[0017] To more clearly illustrate the technical solutions in this invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of this invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0018] Figure 1 This is one of the structural schematic diagrams of the device provided by the present invention for determining the allocation of plant carbon assimilate source libraries.

[0019] Figure 2 yes Figure 1 The diagram shows a schematic of the main body of the leaf chamber.

[0020] Figure 3 yes Figure 1 The diagram shows a schematic of the cover structure of the leaf chamber.

[0021] Figure 4 This is the second schematic diagram of the device for determining the allocation of plant carbon assimilate source pools provided by the present invention.

[0022] Figure 5 This is the third schematic diagram of the device for determining the allocation of plant carbon assimilate source pools provided by the present invention.

[0023] Figure 6 This is a curve showing the dynamic change of CO2 concentration when there are no leaves in the leaf chamber.

[0024] Figure 7 Different NaH 13 Dynamic curve of CO2 concentration change when CO3 mass fraction is used.

[0025] Figure 8 It is a standard curve of CO2 concentration.

[0026] Figure 9 This is a CO2 concentration control curve using a threshold-triggered replenishment method.

[0027] Figure 10This is a stable CO2 concentration curve using PID closed-loop control.

[0028] Figure label: 10. Generator; 11. Third pipeline; 12. Fourth pipeline; 20. Assimilation unit; 21. Leaf chamber; 22. First pipeline; 23. Second pipeline; 24. Concentration monitor; 25. Buffer controller; 26. Barometer; 27. Flow pump; 28. Buffer controller; 29. ​​Main controller; 30. Computer; 211. Leaf chamber body; 212. First protrusion; 213. Second protrusion; 214. Detection module; 215. Flexible sealing membrane; 212. Cover; 2111, cavity; 2112, first groove; 2113, first air inlet; 2114, first air outlet; 2115, second air inlet; 2116, second air outlet; 2121, first through groove; 2122, second through groove. Detailed Implementation

[0029] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of this invention. All other embodiments obtained by those skilled in the art based on the embodiments of this invention without creative effort are within the scope of protection of this invention.

[0030] The following is combined with Figures 1-10 The present invention describes the apparatus and method for determining the allocation of plant carbon assimilate source libraries.

[0031] like Figure 1 , Figure 2 and Figure 3 As shown, in an embodiment of the present invention, the apparatus for determining the allocation of plant carbon assimilate source pools includes: a generator 10 and at least one assimilation unit 20. The generator 10 is a sealed container containing a carbon isotope salt solution; optionally, the carbon isotope salt solution may be NaH+. 13 CO3 + NaH 12 CO3 solution, KaH 13 CO3+KaH 12 CO3 solution, carbon isotope salt solution, and citric acid can react to produce CO2, which includes... 12 CO2 and 13CO2. The generator 10 is connected to the assimilation unit 20 to form a circulation loop. The CO2 generated in the generator 10 enters the assimilation unit 20 for photosynthesis, and the oxygen produced by photosynthesis then returns to the generator 10. In this embodiment, the assimilation unit 20 can be one or more. When the number of assimilation units 20 is multiple, such as... Figure 4 As shown, the CO2 produced by the generator 10 can enter each assimilation unit 20 to participate in photosynthesis, and the oxygen produced by each assimilation unit 20 enters the generator 10 to simultaneously label multiple plant tissues.

[0032] Each assimilation unit 20 includes a leaf chamber 21, a concentration monitor 24, and a main controller 29. The leaf chamber 21 has a cavity 2111 for containing plant tissue. The leaf chamber 21 is connected to the generator 10. The concentration monitor 24 is connected to the leaf chamber 21 and is used to monitor the CO2 concentration within the leaf chamber 21. The main controller 29 is electrically connected to the concentration monitor 24 and is used to combine the concentration data detected by the concentration monitor 24 with the concentration of CO2 contained within the generator 10. 13 The mass fraction of CO3 solution, establishing the relationship between CO2 concentration and... 13 A standard curve showing the relationship between CO2 mass fractions was obtained, and the CO2 concentration in the gas was determined based on the real-time detected CO2 concentration. 13 The mass fraction of CO2, based on CO2 concentration and 13 The mass fraction of CO2 in the CO2 gas was calculated. 13 CO2 concentration.

[0033] Specifically, the concentration monitor has 24 pairs. 12 CO2 is relatively sensitive and can be accurately read under normal, unlabeled natural conditions. 12 The concentration value of CO2, but when the gas contains isotopes 13 When CO2 is present, the concentration monitor 24 will not be able to accurately reflect the concentration of CO2 gas. 12 The actual concentration of CO2. In this embodiment, NaHCO3 was selected for the experiment, and the total mass of NaHCO3 participating in the reaction was kept constant (e.g., 0.02 g), while the concentration of NaHCO3 was systematically changed. 13 The mass ratio of CO3 in total NaHCO3 (e.g., m(NaHCO3)) 13 CO3) / m(NaH 13 CO3+ NaH 12 (CO3) = 0, 20%, 40%, 60%, 100%. Different mass ratios of NaH2O. 13The concentration of CO2 produced by CO3 varies. Concentration monitor 24 detects the CO2 concentration at each mass fraction and sends the data to main controller 29. Main controller 29 uses the measured CO2 concentration as the ordinate (Y). During labeling, considering that plant photosynthesis has a minimal impact on isotope ratios, it is assumed that photosynthesis does not change the isotope ratio within the volume, and that CO2 contains... 13 The mass fraction of CO2 and NaH 13 The mass fraction of CO3 in the total NaHCO3 is equal, with 13 Using the mass fraction of CO2 as the x-axis, a curve was fitted to obtain the relationship between CO2 concentration and... 13 The standard curve equation for the mass fraction of CO2 is Y = f(X). In the subsequent actual labeling process, generator 10 produces CO2 gas with a known total molar amount. The main controller 29 receives the reading Y from the concentration monitor 24 in real time and calls the pre-stored standard curve equation Y = f(X). Through calculation or table lookup, the mass fraction of CO2 in the current gas mixture can be uniquely determined. 13 The actual mass fraction X of CO2 in the system can be accurately calculated using the known total molar amount of CO2. 13 Real-time CO2 concentration.

[0034] For example, using 0.02 g of pure NaHCO3 as the reactant, the reaction was run continuously for 3600 seconds. Concentration monitor 24 acquired the CO2 concentration within leaf chamber 21 every second. The monitoring results are as follows: Figure 6 As shown, after the injection of NaHCO3 solution (430 s), the CO2 concentration in chamber 21 rapidly increased from the background level (463.5 ppm). The concentration peaked at approximately 1000 seconds, then entered a relatively stable plateau period, finally reaching approximately 8196.8 ppm at 3600 seconds. Throughout the process, the concentration monitor 24 demonstrated good continuous monitoring capability and dynamic response characteristics; the data curve was smooth, with no abnormal jumps, indicating that the gas was uniformly mixed in the reflux path and the device operated stably.

[0035] In this embodiment, the concentration monitor 24 is an NDIR-based sensor, which is inexpensive and greatly reduces the cost of devices for determining the allocation of plant carbon assimilate source libraries.

[0036] Optionally, in this embodiment, NaH 13 CO3 in total NaHCO3 (including NaH) 12 CO3 and NaH 13 The mass fraction of CO3 can be determined according to the experimental requirements. 13The CO2 abundance can be flexibly adjusted between 5% and 99%. To balance labeling effectiveness and cost, a range of 20% to 50% is preferred. The total concentration of the NaHCO3 solution can be configured between 0.1 mol / L and 1.0 mol / L. By controlling the solution volume (typically 1-10 mL) and concentration injected into the generator 10, the total amount of CO2 gas generated can be precisely controlled, achieving milligram-level accurate labeling.

[0037] The apparatus for determining the allocation of plant carbon assimilate source libraries provided in this invention, by setting up a concentration monitor and a main controller, establishes a correlation between CO2 concentration and... 13 A standard curve relating CO2 mass fractions can be obtained based on real-time CO2 concentration measurements. 13 The mass fraction of CO2 can then be calculated from the CO2 gas content. 13 The concentration of CO2 was measured, enabling the determination of CO2 concentration without relying on expensive isotope analyzers. 13 High-precision, low-cost quantitative control of CO2 labeled gas concentration.

[0038] like Figure 2 As shown, the leaf chamber 21 includes a leaf chamber body 211 and a cover 212. The leaf chamber body 211 includes a base plate and multiple plates, which surround the base plate to form a cavity 2111. A pair of plates facing each other are a first plate and a second plate. The first plate has a first groove 2112 for accommodating the petiole of the plant. The second plate has multiple pairs of air inlets and outlets, one pair of which is connected to the generator 10, and the other pair is connected to the concentration monitor 24. The cover 212 covers the leaf chamber body 211 and is sealed to it. In this embodiment, the cover 212 is a light-transmitting cover, and it has a first through groove 2121 and a second through groove 2122, with the first through groove communicating with the first groove. In the plant tissue placement chamber, the plant petiole is placed into the first groove 2112 through the first through groove 2121, and the plant leaf is placed into the cavity 2111 through the second through groove 2122. Then, the first groove 2112, the first through groove 2121, and the second through groove 2122 are sealed with flexible sealant. CO2 generated in the generator 10 can enter the leaf chamber 21 through the air inlet for photosynthesis, and the generated oxygen enters the generator 10 through the air outlet.

[0039] In this embodiment, the leaf chamber body 211 is made of non-toxic, chemically stable, and easily processed plastic, such as polymethyl methacrylate (acrylic), polycarbonate (PC), or ABS engineering plastic, to ensure the device's portability and durability. The cover 212 has a light transmittance of greater than or equal to 90%, and its material can be acrylic sheet or tempered glass, to ensure that the light required for photosynthesis during the marking process can fully enter the leaf chamber 21.

[0040] The leaf chamber body 211 and the cover 212 are sealed with a sealing ring. The first groove 2112 can be sealed with flexible sealant after the petiole is placed in it. The first through groove 2121 and the second through groove 2122 are also sealed with flexible sealant, which can be removed after use. In this embodiment, by providing a cavity within the leaf chamber body 211, plant tissues with large leaf areas, such as cotton boll-aligned leaves, can be marked. Furthermore, based on the morphology of the plant tissue to be marked, a support identical to the plant tissue morphology can be formed on the base plate of the leaf chamber body 211 using 3D printing to better support the plant tissue.

[0041] The device for determining the allocation of plant carbon assimilate source library provided in this embodiment of the invention has a leaf chamber that can accommodate plant tissues of different sizes, making it highly applicable. The leaf chamber body and the cover are sealed together, and the first groove, the first through groove and the second through groove are all sealed with flexible sealant, which has a good sealing effect and minimal interference with plant tissues, thus avoiding damage to plant tissues and improving the accuracy of detection.

[0042] Furthermore, in an embodiment of the present invention, the leaf chamber 21 further includes an elastic member disposed within the first groove 2112, and the elastic member has a second groove for accommodating the petiole of the plant. In this embodiment, the elastic deformation of the elastic member allows the second groove to accommodate petioles of different diameters, improving the versatility of the leaf chamber 21.

[0043] Furthermore, a sealant, such as sealing clay or silicone gel, can be filled into the second groove. This type of sealant has excellent plasticity, inertness, and sealing properties, and can perfectly conform to the petiole surface. While achieving ultra-high airtightness, it causes almost no mechanical damage to plant tissue, thus improving the airtightness of the leaf chamber.

[0044] like Figure 2 As shown, in an embodiment of the present invention, the blade chamber 21 further includes a flexible sealing membrane 215. The bottom plate of the blade chamber body 211 is provided with a third through groove. The flexible sealing membrane 215 is disposed at the third through groove. The flexible sealing membrane 215 can undergo elastic deformation. The flexible sealing membrane 215 is used to adjust the air pressure in the blade chamber 21 so that the pressure difference between the air pressure in the blade chamber 21 and the ambient air pressure is less than 0.1 kPa.

[0045] like Figure 2 As shown, in an embodiment of the present invention, a pair of first protrusions 212 and a pair of second protrusions 213 are provided in the cavity 2111 of the leaf chamber body 211, with the pair of second protrusions 213 located between the pair of first protrusions 212. A first air inlet 2113, a first air outlet 2114, a second air inlet 2115, and a second air outlet 2116 are provided on the second plate of the leaf chamber body 211. The first air inlet 2113 and the first air outlet 2114 are connected to the generator 10, and the second air inlet 2115 and the second air outlet 2116 are connected to the concentration monitor 24. Each first protrusion 212 is provided with a first channel, so that the first air inlet 2113 communicates with the cavity 2111 through the first channel, and the first air outlet 2114 communicates with the cavity 2111 through the first channel. Each second protrusion 213 is provided with a second channel, so that the second air inlet 2115 communicates with the cavity 2111 through the second channel, and the second air outlet 2116 communicates with the cavity 2111 through the second channel. The length of the first protrusion 212 is greater than the length of the second protrusion 213, so as to deliver CO2 to the plant tissue.

[0046] like Figure 2 As shown, in an embodiment of the present invention, the leaf chamber body 211 is further provided with a detection module 214, which is used to detect the temperature, humidity and light intensity inside the leaf chamber 21.

[0047] like Figure 1 As shown, in an embodiment of the present invention, the assimilation unit 20 further includes a flow pump 27, a barometer 26, and a buffer controller 25. The generator 10 is connected to the first air inlet 2113 of the blade chamber 21 via a first pipe 22 and to the first air outlet 2114 of the blade chamber 21 via a second pipe 23. The buffer controller 25 is disposed on the first pipe 22 and is used to reduce the difference between the environmental parameters inside the blade chamber 21 and the external environmental parameters. In this embodiment, the environmental parameters include: air pressure, oxygen concentration, and humidity, etc. The barometer 26 and the flow pump 27 are both disposed on the second pipe 23. The barometer 26 is used to detect the air pressure in the circulation loop. The flow pump 27 is used to adjust the flow rate of the circulating gas in the circulation loop to regulate the air pressure in the circulation loop.

[0048] Furthermore, in an embodiment of the present invention, both the flow pump 27 and the buffer controller 25 are electrically connected to the main controller 29. The calculation is based on the standard curve equation Y = f(X) and the total concentration of CO2. 13 After determining the CO2 concentration, the main controller 29 can use this calculated value as feedback. By adjusting the flow pump 27 and the buffer controller 25, it can regulate the air pressure, oxygen concentration, and other parameters within the blade chamber 21, thereby achieving the desired control within the blade chamber 21. 13 Closed-loop precise control of CO2 concentration ensures the quantification and accuracy of the labeling process.

[0049] Specifically, the main controller 29 controls the operation of the flow pump 27 to extract gas from the blade chamber 21. The buffer controller 25 can effectively buffer the pressure fluctuations caused by the addition of carbon dioxide or temperature changes, maintain the internal pressure stability of the assimilation unit 20, and keep the pressure fluctuation of the blade chamber less than 0.1 kPa. This protects the structural safety of the closed loop, reduces the "dead volume" of the gas circulation, and promotes the rapid and uniform distribution of the labeled gas in the assimilation unit 20. It also reduces the interference of pressure fluctuations on the accuracy of carbon dioxide concentration detection, and further improves the uniformity and efficiency of labeling.

[0050] Furthermore, the main controller 29 is electrically connected to the detection module 214 within the leaf chamber 21. The main controller 29 can precisely regulate key environmental factors within the leaf chamber 21, with its parameters set to reference the target plant's ambient growth temperature: a temperature range of 20℃ to 45℃, preferably 25℃ to 35℃, to adapt to the photosynthetic optimum temperature for most crops. The difference between the leaf chamber temperature and the ambient temperature is less than 0.5℃. An integrated or external LED light source provides an intensity range of 100 μmol·m⁻². -2 ·s -1 Up to 2000 μmol·m -2 ·s -1 Photosynthetically active radiation (PAR). For C3 crops such as cotton and rice, a light intensity of 800-1500 μmol·m⁻² is preferred. -2 ·s -1 .

[0051] like Figure 1 As shown, in an embodiment of the present invention, the assimilation unit 20 further includes a pair of regulators 28 connected to the generator 10. One of the regulators 28 is used to adjust the volume of the carbon isotope salt solution in the generator 10, and the other is used to adjust the volume of citric acid. The main controller 29 is also used to control the operation of the regulators 28 to change the volume of the carbon isotope salt solution and the volume of citric acid in the generator 10.

[0052] like Figure 4 As shown, in an embodiment of the invention, each generator 10 is connected to an assimilation unit 20 to calibrate a plant tissue. Multiple devices for determining the allocation of plant carbon assimilate source libraries are connected to a computer 30 to send the calculated carbon assimilate allocation efficiency to the computer 30 for storage.

[0053] like Figure 5As shown, in an embodiment of the present invention, the generator 10 is connected to the first pipeline 22 of each assimilation unit 20 via the third pipeline 11, and the generator 10 is connected to the second pipeline 23 of each assimilation unit 20 via the fourth pipeline 12 to form a high-throughput parallel system, thereby enabling the labeling of multiple plants or multiple source organs and sink organs.

[0054] The device for determining the allocation of plant carbon assimilate source libraries provided in this invention is applicable not only to major field crops such as cotton, soybean, rice, wheat, and corn, but also to horticultural crops such as tomatoes and cucumbers, as well as branches or single leaves of fruit trees (such as apples and citrus). Through customized micro-chambers, this device is also suitable for whole-plant or partial organ studies of model plants such as Arabidopsis and tobacco, achieving full-scale coverage from model plants to economic crops. The gas mixing and concentration stabilization rate of the assimilation unit is closely related to the total volume of the device. Under standard operating conditions, the time required for the CO2 concentration in the chamber to reach a stable state within ±5% of the target value (i.e., the system response time) is less than (3 minutes × system volume correction factor K), where the volume correction factor K = total device volume (L) / 10 (L). For example, for a device with a total volume of 1.5L, the concentration stabilization time is expected to be less than 1.5 minutes. This device has a rapid response capability and can meet the needs of dynamic experiments.

[0055] The apparatus for determining the allocation of plant carbon assimilate source libraries provided in this invention employs a custom-designed, fitted leaf chamber based on the shape of the plant's marker sites (e.g., achieved through 3D printing and sealing with a flexible sealing material gel). This allows for partial or complete sealing of leaves or other organs with minimal intervention, making it suitable for plants of different sizes and supporting high-throughput studies. It also allows for precise control of the precursor liquid (e.g., a specific concentration of NaH+). 12 CO3 / NaH 13 The reaction volume and concentration of the CO3 mixed solution are used to quantitatively generate and supply CO2 (including CO3 mixed solution) at the target concentration in a closed-loop gas circuit. 12 CO2 and 13 This method not only achieves precise labeling at the milligram level with extremely low single-use consumption (20-100mg), but more importantly, it allows for direct and convenient determination and control of the initial concentration and ratio of isotopes in the circulation system through precise solution preparation, overcoming the traditional reliance on expensive isotope spectrometers for real-time detection. Furthermore, this method can control and maintain key environmental factors such as total CO2 concentration, humidity, light, and temperature within the leaf chamber in accordance with the surrounding environment, reducing labeling interference with the plant. Under special circumstances, the temperature, humidity, and light conditions within the leaf chamber can also be adjusted to meet experimental requirements.

[0056] This invention also provides a method for determining the allocation of plant carbon assimilate source libraries, specifically including the following steps: Step 01: Establish CO2 concentration and 13 Standard curve of CO2 mass fraction.

[0057] Keeping the total mass of NaHCO3 participating in the reaction constant (e.g., 0.02 g), systematically change the NaH... 13 The mass ratio of CO3 in total NaHCO3 (e.g., m(NaHCO3)) 13 CO3) / m(NaH 13 CO3+ NaH 12 CO3) = 0, 20%, 40%, 60%, 100%. Different mass ratios of NaH2O. 13 The concentration of CO2 produced from CO3 varies, and the CO2 concentration value at each mass fraction is obtained. The measured CO2 concentration value is used as the ordinate (Y). During the labeling process, considering that plant photosynthesis has a small effect on the isotope ratio, it is assumed that photosynthesis does not change the isotope ratio within the volume, and the CO2 concentration is... 13 The mass fraction of CO2 and NaH 13 The mass fraction of CO3 in the total NaHCO3 is equal, with 13 Using the mass fraction of CO2 as the x-axis, a curve was fitted to obtain the relationship between CO2 concentration and... 13 The standard curve equation for the mass fraction of CO2 is Y = f(X). In the subsequent actual labeling process, generator 10 produces CO2 gas with a known total molar amount. The main controller 29 receives the reading Y from the concentration monitor 24 in real time and calls the pre-stored standard curve equation Y = f(X). Through calculation or table lookup, the mass fraction of CO2 in the current gas mixture can be uniquely determined. 13 The actual mass fraction X of CO2 in the system can be accurately calculated using the known total molar amount of CO2. 13 Real-time CO2 concentration.

[0058] For example, keeping the total mass of NaHCO3 constant at 0.02 g, by changing the NaH... 13 CO3 percentage (m(NaH) 13 CO 3 ) / m(NaH 13 CO3+ NaH 12 CO3) = 0, 0.2, 0.4, 0.6, 1) produces five kinds of... 13 Standard gases with CO2 concentrations (0%, 20%, 40%, 60%, 100%) were continuously monitored and recorded within leaf chamber 21. For example... Figure 7As shown, by analyzing the stable CO2 monitoring value at the experimental endpoint (t=1800s), linear regression analysis was performed to obtain the standard curve equation: Y = -1961.05X +2315.47 (R²). 2 = 0.9998). For example Figure 8 As shown, the X coordinate is 13 The theoretical mass fraction of CO2 (0-1), with the Y-axis representing the CO2 concentration (ppm). By monitoring the CO2 concentration values ​​in real time, the concentration of CO2 in the standard curve can be obtained. 13 The mass fraction of CO2 can be accurately calculated based on the known CO2 concentration produced by the reaction within leaf chamber 21. 13 The actual concentration of CO2.

[0059] Step 02: Obtain the first mass fraction of carbon in the dry matter of the source organ and 13 The first isotopic percentage of C, the second mass fraction of carbon in the dry matter of the organoids, and 13 The percentage of second isotopes of C.

[0060] Specifically, in order to better detect CO2 concentration and achieve better results 13 C-labeling, with a specific proportion selected in the experiment. 13 C. Before the labeling experiment, source organs (leaves) and sink organs (fruits, spikes, etc.) were collected from plants of the same type and with the same growth stage as the test plants. After blanching (105℃, 30 min), drying (80℃ to constant weight), and grinding into powder, the first mass fraction of carbon (fCs) in the dry matter of the source organs was determined using an elemental analyzer, and the second mass fraction of carbon (fCk) in the dry matter of the sink organs was determined using a stable isotope ratio mass spectrometer. 13 The percentage of the first isotope of carbon (A%13s-pre), in the dry matter of the organ deposit. 13 The second isotope percentage of C, A%13k-pre.

[0061] Step 03: Place the source organ and stem organ separately into leaf chamber 21 for photosynthesis, obtain the CO2 concentration inside leaf chamber 21, and based on the CO2 concentration, use the CO2 concentration and... 13 Standard curve control of CO2 mass fraction 13 The CO2 concentration is within the set range.

[0062] Specifically, the source organ (leaf) of the plant to be tested is sealed and placed in leaf chamber 21, and flow pump 27 is started and adjusted to a predetermined flow rate. NaH+ with a predetermined ratio and amount is then introduced via regulator 28. 13 CO3 + NaH 12 The CO3 mixed solution is injected into generator 10, producing 12 CO2 and13 CO2 gas is continuously introduced into the blade chamber 21. The concentration of CO2 is monitored in real time by a concentration monitor 24, and the concentration within the blade chamber 21 is calculated using a standard curve. 13 The concentration of CO2 is measured, and the flow rate of the flow pump 27 and the action of the buffer controller 25 are adjusted to... 13 The CO2 concentration value remained stable at the set value.

[0063] Accordingly, the bank organs of the plant to be tested are sealed in leaf chamber 21 and marked according to the above method.

[0064] Step 04: Obtain the third mass fraction of carbon in the dry matter of the labeled source organs at different time points. 13 The percentage of the third isotope of C, the fourth mass fraction of carbon in the dry matter of the organ, and 13 The fourth isotope percentage of C; the carbon assimilation efficiency of the source organ is calculated based on the first mass fraction, the first isotope percentage, the third mass fraction, and the third isotope percentage; the carbon assimilation efficiency of the deposit organ is calculated based on the second mass fraction, the second isotope percentage, the fourth mass fraction, and the fourth isotope percentage.

[0065] Specifically, after labeling, at different predetermined time points (e.g., 1 h, 24 h, 72 h, 144 h after labeling), the third mass fraction of carbon in the dry matter of the labeled source organs was collected. 13 The percentage of the third isotope of C, the fourth mass fraction of carbon in the dry matter of the organ, and 13 The fourth isotope percentage of C. The methods for obtaining the third mass fraction, third isotope percentage, fourth mass fraction, and fourth isotope percentage are the same as those for obtaining the first mass fraction, first isotope percentage, second mass fraction, and second isotope percentage, and will not be elaborated further here.

[0066] The carbon assimilation partition efficiency is calculated based on the following formula: Efficiency (s→k) = [Δ 13 Ck / (Δ 13 Cs+Δ 13 Ck) ]× 100%, where, Δ 13 Cs = D(Ws) × f(Cs) × [(A% 13 s-post - A% 13 s-pre) / 100], Δ 13 Ck = D(Wk) × f(Ck) × [(A% 13 k-post - A% 13 [k-pre) / 100], Where D(Ws) is the dry weight of the source organ, and D(Wk) is the dry weight of the sink organ; f(Cs) is the first mass fraction of carbon in the dry matter of the source organ, and f(Ck) is the second mass fraction of carbon in the dry matter of the sink organ; A%13s-pre is the dry weight of the source organ before labeling. 13 The first isotope percentage of C, and A%13s-post is the source organ labeled. 13 The third isotope percentage of C; A%13k-pre is the amount before organ labeling in the library. 13 The second isotope percentage of C, and A%13k-post is the labeling of the library organ. 13 Percentage of the fourth isotope of C. Δ 13 Cs represents the excess of source organ fixation during labeling. 13 Total C (g), Δ 13 Cs is calculated based on the third mass fraction and the first mass fraction. Δ 13 Ck represents the total amount (g) of 13C transported and accumulated in the reservoir organ at a specific point in time, i.e., Δ 13 Ck is calculated based on the fourth and second mass fractions. By calculating the distribution efficiency at different time points, a time-dynamic curve of carbon assimilate transport can be plotted, revealing the patterns of material flow between source and sink. Combined with Δ13Cs, the absolute transport volume can also be further calculated.

[0067] Figure 9 This is a CO2 concentration control curve using a threshold-triggered replenishment method. The curves in the figure show the control method during the labeling process, where when the CO2 concentration drops to a preset threshold (e.g., 1000 ppm), the concentration is gradually increased by adding a quantitative amount of reaction solution to maintain the labeling process. The figure also illustrates the concentration change curves of the labeled gas maintained over a wide range by monitoring the CO2 concentration and discretely adding reactants.

[0068] Figure 10 The figure shows the CO2 concentration stability curve using PID closed-loop control. The curve illustrates the control method and effect during the labeling process, where the PID controller dynamically adjusts the reaction solution injection rate based on concentration deviation to stabilize the CO2 concentration near the set target value (e.g., 1800 ppm). The figure also demonstrates the concentration change curve achieved by continuously fine-tuning the reactant addition rate through the PID feedback system to obtain a highly accurate and stable labeled gas concentration.

[0069] like Figure 10 As shown, it can be observed that in cotton, the transport efficiency of carbon assimilates to the boll typically peaks approximately x hours after labeling. Combined with Δ... 13Cs can calculate absolute transport volume. This method can accurately quantify the carbon contribution of a specific source leaf to its corresponding cotton boll, and is suitable for comparing source-sink relationships under different varieties and cultivation practices.

[0070] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. An apparatus for determining the allocation of a plant carbon assimilate source pool, characterized in that, include: A generator for producing CO2, the CO2 comprising 12 CO2 and 13 CO2; At least one assimilation unit, each assimilation unit forming a loop with the generator, each assimilation unit comprising: Leaf chamber, the leaf chamber having a cavity for accommodating plant tissue, the leaf chamber being connected to the generator; A concentration monitor, connected to the leaf chamber, is used to monitor the CO2 concentration inside the leaf chamber; The main controller, electrically connected to the concentration monitor, is used to analyze the CO2 concentration data detected by the concentration monitor in conjunction with the concentration data of the generator. 13 The mass fraction of CO3 solution, establishing the relationship between CO2 concentration and... 13 A standard curve relating CO2 mass fractions; based on the real-time detected CO2 concentration, the current... 13 The mass fraction of CO2; based on the CO2 concentration and the 13 The mass fraction of CO2 was calculated. 13 CO2 concentration value.

2. The apparatus for determining the allocation of plant carbon assimilate source pools according to claim 1, characterized in that, The leaf chamber includes: The leaf chamber body has the cavity and has a first plate and a second plate with opposite sides. The first plate has a first groove and the second plate has multiple pairs of air inlets and outlets. The generator is connected to a pair of air inlets and outlets and the concentration monitor is connected to a pair of air inlets and outlets. A cover is placed on the leaf chamber body and sealed to the leaf chamber body. The cover is a light-transmitting cover. The cover has a first through groove and a second through groove. The first through groove is connected to the first groove. The first groove is used to accommodate the petiole of the plant. The cavity is used to accommodate the leaf of the plant. The first groove, the first through groove and the second through groove are sealed with flexible sealant.

3. The apparatus for determining the allocation of plant carbon assimilate source pools according to claim 2, characterized in that, The leaf chamber also includes an elastic element disposed in the first groove, and the elastic element has a second groove for accommodating the petiole of the plant.

4. The apparatus for determining the allocation of plant carbon assimilate source pools according to claim 3, characterized in that, The second groove is filled with sealant.

5. The apparatus for determining the allocation of plant carbon assimilate source pools according to claim 2, characterized in that, The leaf chamber also includes a flexible sealing membrane, and the bottom plate of the leaf chamber body is provided with a third through groove, and the flexible sealing membrane is disposed in the third through groove.

6. The apparatus for determining the allocation of plant carbon assimilate source pools according to claim 1, characterized in that, The assimilation unit also includes a flow pump, which is disposed in the circulation loop and is used to regulate the flow rate of the circulating gas in the circulation loop. The flow pump is electrically connected to the main controller.

7. The apparatus for determining the allocation of plant carbon assimilate source pools according to claim 1, characterized in that, The assimilation unit also includes a barometer, which is disposed in the circulation loop and is used to detect the pressure of the circulation loop.

8. The apparatus for determining the allocation of plant carbon assimilate source pools according to claim 1, characterized in that, The assimilation unit further includes a buffer controller, which is disposed in the circulation loop. The buffer controller is used to reduce the difference between the environmental parameters inside the leaf chamber and the external environmental parameters. The buffer controller is electrically connected to the main controller.

9. A method for determining the allocation of plant carbon assimilate source pools based on any one of claims 1-8, characterized in that, include: Establish CO2 concentration and 13 Standard curve of CO2 mass fraction; Obtain the first mass fraction of carbon in the dry matter of the source organ and 13 The first isotopic percentage of C, the second mass fraction of carbon in the dry matter of the organoids, and 13 The percentage of second isotopes of C; The source organ and stem organ were placed in the leaf chamber for photosynthesis, and the CO2 concentration in the leaf chamber was obtained. Based on the CO2 concentration, the standard curve was used to control the process. 13 The CO2 concentration is within the set range; Obtain the third mass fraction of carbon in the dry matter of the labeled source organs at different time points. 13 The percentage of the third isotope of C, the fourth mass fraction of carbon in the dry matter of the organ, and 13 Percentage of the fourth isotope of C; The carbon assimilation efficiency of the source organ is calculated based on the first mass fraction, the first isotope percentage, the third mass fraction, and the third isotope percentage. The carbon assimilation efficiency of the library organ is calculated based on the second mass fraction, the second isotope percentage, the fourth mass fraction, and the fourth isotope percentage.

10. The method according to claim 9, characterized in that, The establishment of CO2 concentration and 13 The steps involved in creating a standard curve for CO2 mass fractions include: To maintain a constant total mass of the carbon isotope salt solution within the generator, the content of the solution is changed. 13 The mass fraction of the CO3 solution was obtained. 13 The concentration of CO2 produced by a CO3 solution at different mass fractions; Based on the reaction 13 CO2 and containing 13 Based on the principle that the mass fraction of CO3 in the solution is equal, we obtain... 13 Mass fraction of CO2; by 13 Using the mass fraction of CO2 as the x-axis and the concentration of CO2 as the y-axis, a curve was fitted to obtain the relationship between the CO2 concentration and the... 13 Standard curve of CO2 mass fraction.

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