A device and method for collecting plant root exudates and delivering nutrient solution

Abstract

The application discloses a device and method with functions of plant root exudate collection and nutrient liquid delivery, and belongs to the technical field of plant root exudate collection. The device comprises a control module, a collection module and a collection module, and the modules are connected through pipelines. The control module comprises an adjustable pressure vacuum pump and a controller, and the adjustable pressure vacuum pump is electrically connected with the controller. The method and device can efficiently extract plant root exudates in different growth periods of plants without damaging the growth state of plants, and the results are stable and the data are reliable. The device mainly comprises a control module, a collection module and a collection module. The modules are connected through pipelines, and can be quickly replaced when a single module has a problem, facilitating maintenance and maintenance. The device is reliable in performance, can be buried in soil and other substrates for a long time, is not damaged, and does not affect the extraction efficiency; the device is intelligent, efficient and continuous, greatly improving the extraction efficiency and flexibility.

Description

Technical Field

[0001] This invention relates to the field of plant root exudate collection technology, and in particular to a device and method that combines plant root exudate collection and nutrient solution delivery. Background Technology

[0002] As a crucial link between the aboveground parts of plants and the soil, the root system drives the dynamic exchange of substances, energy, and information in the rhizosphere during plant growth by releasing root exudates. Root exudates are the sum of organic matter actively or passively released by plant roots into the surrounding medium, encompassing three main categories: low molecular weight substances (such as organic acids, amino acids, soluble sugars, and plant hormones), high molecular weight substances (such as mucilage and extracellular enzymes), and cell shed material (such as root hair fragments). These exudates play a central role in rhizosphere nutrient activation, pollutant degradation (such as the bioremediation of polycyclic aromatic hydrocarbons), and plant-microbe signal transduction by altering soil physicochemical properties and regulating the structure and function of the microbial community.

[0003] Furthermore, recent studies have shown that the rate of root exudate input is closely related to plant growth strategies. The magnitude of root exudate input flux can be used as a functional trait for classifying plant nutrient utilization strategies, and it is of great significance in plant growth and evolutionary ecology. Therefore, conducting qualitative and quantitative research on the input of chemical components of root exudates is of great ecological significance. It is the prerequisite and foundation for further enriching and elucidating research areas such as plant growth nutrient cycling mechanisms, allelopathic mechanisms, and the maintenance of rhizosphere microecological structure and function.

[0004] Correspondingly, numerous techniques and methods for collecting root exudate components have emerged. However, because plant roots grow in underground ecosystems composed of soil matrix, plant root exudates can only be distributed within a narrow area centered on the root system. In addition, root exudates are characterized by low content, complex composition, and easy assimilation and utilization by microorganisms, making the timely and effective collection of plant root exudates, especially in-situ collection in the field, a difficult and crucial aspect of rhizosphere ecology research.

[0005] Currently, root exudate extraction methods mainly include indoor collection methods and in-situ field collection methods. Indoor collection primarily targets the collection of plant root exudates under controlled indoor conditions. Based on whether the collection device (method) can collect root exudates continuously and cyclically, it is divided into non-continuous root exudate collection methods and continuous root exudate collection methods, as well as destructive water extraction methods and non-destructive continuous collection methods.

[0006] Discontinuous root exudate collection methods mainly consist of traditional methods, which can be categorized into solution culture collection, soil culture collection, and substrate culture collection based on the culture medium. These methods primarily involve removing plant roots after a certain growth cycle, washing the roots with deionized water, and extracting root exudates using deionized water or a trace inorganic salt solution in a light-protected environment. However, discontinuous root exudate collection methods typically face challenges such as low exudate content, cumbersome sample collection and processing with potential sample contamination, and difficulty in real-time, dynamic collection of root exudates. For example, Oburger et al. (Oburger E, Dell'mour M, Hann S, et al. Evaluation of a novel tool for sampling root exudates from soil-grown plants compared to conventional techniques. Environmental and Experimental Botany, 2013, 87: 235-247) invented a root box system for collecting maize root exudates. The most distinctive feature of this root box is the narrow, soilless vertical slit at the bottom for root growth. Once the corn roots have grown into the slit, nutrient solution can be injected at the higher part of the slit, and root exudates can be extracted through an opening at the lower part. This device does not disrupt the natural growth environment between the roots and the soil, causing no damage to the roots, making it a relatively good root exudate collection device. However, it also has certain drawbacks. It cannot realistically simulate the wild growth environment, and the isolated roots, once separated from the soil substrate, may experience disturbances in plant hormone signals, potentially leading to collected root exudates that are not from natural conditions, thus causing experimental bias.

[0007] A continuous root exudate collection and culture system involves transplanting plants into a culture medium (typically quartz sand) with a root exudate collector at the bottom. A suitable amount of culture solution is periodically added to the container. As the solution flows downwards, it washes down the exudates, continuously enriching the organic matter within the collector. Meanwhile, the culture solution, containing inorganic ions, is pumped back into the container by air bubbles, creating a continuous, undisturbed circulation of the liquid. A selective adsorption resin column is typically connected to the bottom of the culture solution to adsorb organic matter. When root exudates flow through the resin, they are adsorbed onto the surface, achieving both enrichment and separation of organic matter. After a certain period, the resin is collected, and the root exudates are eluted from it using a suitable eluent. For example, the national invention patent document (CN119073205A) describes a laboratory hydroponic plant device and method for collecting root exudate ions. This method primarily involves a laboratory hydroponic plant device for collecting root exudate ions. Part of the plant roots enter the root exudate ion collection device, and plant root exudates are collected from the device. Buffer solution and methanol solvent are added to the root exudate collection tank, and after elution, a solution of plant root exudate ions is obtained. This solution is then freeze-dried to obtain the root exudate ions. In summary, the solution culture collection method is simple to operate, causes minimal damage to the roots during the collection process, is unaffected by the elements in the soil particles themselves on the composition of the exudate, and allows for better control of aseptic conditions, thus avoiding the decomposition and utilization of the exudate by microorganisms.

[0008] However, the following problems are common in plant root exudates collected from solution culture: (1) During solution culture, the plant roots are immersed in the solution, resulting in poor root aeration, inhibited root respiration, weakened physiological functions, and the potential production of toxic gases under hypoxic conditions, which can affect the normal growth of the plant. Therefore, the collection time for root exudates collected by solution culture should be minimized. (2) Plants cultured in solution generally have less root hair development, and their rhizosphere microenvironment (such as aeration, nutrient distribution, root branching structure, microorganisms, etc.) differs significantly from the soil environment under natural conditions. They also lack the mechanical wear of fine roots by soil particles and the feedback effect of rhizosphere microorganisms on root growth. These factors directly affect the morphological characteristics of the root system and the chemical composition and content of root exudates. Therefore, this method can only represent the root exudation of plants under solution culture conditions and cannot accurately reflect the root exudation under natural conditions.

[0009] In-situ collection methods for root exudates can be broadly categorized into in-situ extraction and non-destructive in-situ monitoring methods, depending on whether the exudates are collected using specialized equipment under in-situ conditions. For example, Phillips et al. (Phillips RP, Erlitz Y, Bier R, et al. New approach for capturing soluble root exudates in forest soils. Functional Ecology, 2008, 22: 990-999) designed a static in-situ collection system specifically for collecting root exudates from woody plants in forest ecosystems. This method involves digging up well-developed, intact root systems in the field, washing away soil particles attached to the roots with nutrient solution, placing the roots on a tray of a specific size, covering them with a 1:1 sand-soil mixture, culturing for 2-3 days, washing them again, and then placing them in small glass tubes to begin collecting root exudates. This method overcomes the long-standing limitation of in-situ collection of forest root exudates and provides a better method for their collection. However, the device also has some drawbacks: (1) the glass syringe is relatively small and cannot keep the roots in their natural shape in the syringe, which may affect the secretion rate of the secretions; (2) because the syringe is filled with glass beads, the filter tube may be blocked when the root secretions are filtered; (3) the sealing film used by the device may create gaps between the root system and the root system, causing certain pollution to the secretions in the soil environment. Therefore, the in-situ collection of root secretions faces great challenges: on the one hand, its concentration is low, its composition is complex and it is easily metabolized by microorganisms; on the other hand, existing technologies mostly rely on indoor controlled conditions (such as hydroponics and sand culture) or destructive sampling (such as traditional water extraction), which makes it difficult to truly reflect the composition of secretions under the interaction of rhizosphere microorganisms and plants in the natural soil environment, and it is impossible to conduct long-term non-destructive dynamic monitoring of perennial woody plants (such as forest tree species). In addition, although the field in-situ collection technology partially retains the ecological authenticity, it is affected by environmental fluctuations, microbial degradation and interspecific competition, resulting in low recovery rate of target components and difficulty in quantitative analysis, which seriously restricts the in-depth analysis of rhizosphere carbon cycle, allelopathy and ecological feedback mechanism. Therefore, there is an urgent need to develop an efficient root exudate collection system that is compatible with the in-situ soil-microbial environment and supports continuous non-destructive collection in order to overcome technical bottlenecks and provide accurate data support for the study of underground ecological processes.

[0010] In summary, existing technologies have three major shortcomings: First, while indoor cultivation methods (such as hydroponics and sand culture) can control sterile conditions, they neglect the synergistic metabolic effects between rhizosphere microorganisms and plants, resulting in the absence of key signaling molecules (such as microbial metabolites) in secretions; second, while in-situ field collection technologies preserve natural ecological feedback, they are unable to eliminate the interference of environmental noise (such as light and interspecific competition) on the composition of secretions; and third, existing devices generally lack non-destructive continuous monitoring capabilities and cannot dynamically track the temporal changes in secretions during the plant's growth cycle. Summary of the Invention

[0011] The technical problem to be solved by this invention is how to solve the problems of cumbersome operation and poor data accuracy of existing plant root exudate collection devices.

[0012] The present invention solves the above-mentioned technical problems through the following technical means:

[0013] The first aspect of the present invention provides a device for collecting plant root exudates and delivering nutrient solution, comprising a control module, a collection module, and a collection module, wherein the modules are connected by pipelines; the control module includes an adjustable pressure vacuum pump and a controller, wherein the adjustable pressure vacuum pump is electrically connected to the controller; the collection module includes a container and a double-ended connector, wherein the double-ended connector is fixedly connected to the container; the collection module includes a hollow fiber tube and a connector, wherein the hollow fiber tube and the connector are fixedly connected.

[0014] Preferably, the adjustable pressure vacuum pump is provided with a positive pressure port, a negative pressure port and a pressure regulating valve; the positive pressure port or the negative pressure port is connected to the double-connector through a gas supply pipe.

[0015] Preferably, the double-ended connector and the connecting head are connected via an infusion tube.

[0016] Preferably, the infusion tube is also equipped with a flow meter.

[0017] Preferably, the controller is programmed to control the operating mode, pressure, and operating time of the adjustable pressure vacuum pump.

[0018] Preferably, the control module is used to control the pressure and time.

[0019] Preferably, the collection module is used to store nutrient solution and root exudates.

[0020] More preferably, the dual-connector is used to connect the adjustable pressure vacuum pump and the acquisition module, serving to seal and transfer liquid.

[0021] Preferably, the acquisition module is used to deliver nutrient solution to plant roots and extract plant root secretions.

[0022] Preferably, the connector consists of a rigid PE tube and a flexible silicone tube, wherein the rigid PE tube is connected to the hollow fiber tube via the flexible silicone tube.

[0023] Preferably, the connector is a quick-connect fitting with an integrated structure, which connects directly to the hollow fiber tube. (The quick-connect fitting mainly consists of a 2-point to 2-point male thread quick-connect fitting or a 2-point to 4-point male thread crossover fitting, the number of which depends on the number of hollow fiber tubes.) Preferably, the hollow fiber tube can be made into various shapes, such as circular (e.g., Figure 2 (A) Target-shaped (e.g.) Figure 2 (B) Multiple fiber bundles (such as...) Figure 2 (C, D)

[0024] A support frame can be used to shape the hollow fiber tube. The support frame is a product printed from acrylic sheet and mainly serves to fix the hollow fiber tube, thereby achieving the required spatial arrangement.

[0025] Preferably, the membrane material of the hollow fiber tube is polyvinylidene fluoride (PVDF), and the inner lining material is polyester (PET), which are products of existing processes.

[0026] When preparing the acquisition module, one end of the hollow fiber tube can be plugged with glue as needed and allowed to dry for later use. The glue used is CT-8121 type AB glue.

[0027] The preparation method of the acquisition module is as follows: insert the unblocked side of the hollow fiber tube into the external thread side of the quick connector, then seal it with glue and let it dry. The glue is a slow-drying silicone XZX-8231, which is an existing technology product.

[0028] The aforementioned data acquisition module has been iterated and upgraded four times to produce different products based on different needs. Figure 2 ).

[0029] A second aspect of the present invention provides a method for collecting and delivering nutrient solution from plant root exudates, implemented based on the above-mentioned device for collecting and delivering nutrient solution from plant root exudates, comprising the following steps: (1) Module assembly: Connect the control module, collection module, and acquisition module together and test the overall airtightness. If the airtightness is good, proceed to the next step. The acquisition module is buried in the planting substrate before operation, and then plant seeds are planted in the substrate. (If some modules have airtightness issues, the corresponding modules need to be replaced.) (2) Delivery of nutrient solution: Nutrient solution is added to the container, and the pumping pressure is set through the controller program. Nutrient solution is pumped in daily through an adjustable pressure vacuum pump. (The specific pumping volume needs to be determined based on the plant's growth environment. The purpose of this step is to utilize the principle that plant roots tend to grow in areas with sufficient nutrients and water, allowing the plant roots to accumulate around the collection module, thereby maximizing the efficiency of the subsequent negative pressure suction collection module.) (3) Collect plant root exudates: At the corresponding stage of plant growth, root exudates are extracted. First, phosphate buffer is added to the container, and positive pressure is set through the controller program to inject phosphate buffer into the plant substrate. After a period of time, negative pressure is set through the program to aspirate the root exudates.

[0030] (The purpose of the phosphate buffer is to further dissolve root exudates in the phosphate buffer without affecting the growth of plant roots.) Preferably, the extraction process is automated by the controller, which automatically controls the start and stop of the adjustable pressure vacuum pump based on the real-time feedback information from the flow meter, thereby quantitatively extracting root exudates.

[0031] Preferably, the collected root exudates are concentrated using a vacuum freeze dryer to a volume of 3-5 ml, then stored as plant root exudates, which can be sent to the company for testing of plant non-targeted metabolomics according to experimental requirements.

[0032] Preferably, in step (3), the time period is specifically 5 to 10 minutes.

[0033] Preferably, in step (3), the root exudates are extracted repeatedly after an interval of 0.5-1h, for a total of 3-6 times, in order to fully extract the exudates.

[0034] The beneficial effects of this invention are as follows: 1. This invention provides an apparatus and method for collecting plant root exudates. The method and apparatus can efficiently extract plant root exudates at different growth stages of plants without damaging the plant's growth state, and the results are stable and the data is reliable.

[0035] 2. The device of this invention mainly comprises three parts: a control module, a collection module, and a sampling module. These modules are connected by pipelines, allowing for quick replacement in case of a problem with a single module, facilitating maintenance and upkeep. The method of this invention can effectively extract root exudates, with extraction efficiency and accuracy similar to solution culture collection methods, but it can also supplement key metabolite components that cannot be extracted by solution culture collection methods.

[0036] 3. The device of this invention is reliable and can be buried in substrates such as soil for extended periods without damage or impact on extraction efficiency. Furthermore, the extraction method does not affect the plant itself, enabling in-situ extraction of root exudates throughout the plant's entire life cycle. This device and method are intelligent, efficient, and continuous, greatly improving extraction efficiency and flexibility.

[0037] 4. This invention develops a root exudate collection system that combines in-situ environmental simulation, microbial coexistence compatibility, and high-precision dynamic monitoring. It basically meets the following requirements: (1) minimizing root disturbance and supporting long-term continuous sampling; (2) being compatible with natural soil-microbe-plant interaction networks; and (3) integrating in-situ enrichment and rapid stable preservation technologies, which basically overcomes the limitations of existing methods in terms of ecological authenticity, operational efficiency, and data reliability, and provides technical support for the study of plant-soil interaction mechanisms.

[0038] 5. This device and method influence root development and extract root exudates by supplying nutrients to the root system. The efficiency and accuracy of extraction are similar to solution culture collection methods, but it can also supplement key metabolite components that cannot be extracted by solution culture collection methods. The device is reliable and can be buried in substrates such as soil for extended periods without damage or impact on extraction efficiency. This device and method are intelligent, efficient, and continuous, greatly improving extraction efficiency and flexibility. Attached Figure Description

[0039] Figure 1 This is a schematic diagram of the overall acquisition device in Embodiment 1 of the present invention; Figure 2 These are actual structural diagrams of different products using the data acquisition module in Embodiment 1 of the present invention; Figure 3 This is a schematic diagram of the data acquisition module support structure in Embodiment 1 of the present invention; Figure 4 This is a diagram showing the experimental setup of the data acquisition module's effect on root growth in Embodiment 1 of the present invention. Figure 5 This is an experiment in Embodiment 1 of the present invention to show the root distribution of wheat after 30 days of growth, which was conducted by the data acquisition module. Figure 6 The percentage of fresh weight of roots affected by the data acquisition module in the root growth experiment of Embodiment 1 of the present invention; Figure 7 The experiment in Example 1 of this invention showed the growth of potted plants and the status of the data collection module in the experiment on the effect of root exudate extraction on plant growth. Figure 8 This is a diagram showing the species composition of root exudates extracted from three different treatment groups in Example 1 of the present invention; Figure 9 This is a graph showing the differences between the root exudates extracted from three different treatment groups in Example 1 of the present invention. Figure 10 This is a Venn diagram of key metabolic components extracted from root exudates in three different treatment groups in Example 1 of the present invention.

[0040] In the diagram: 1. Controller; 2. Adjustable pressure vacuum pump; 21. Positive pressure port; 22. Negative pressure port; 3. Double-ended connector; 4. Container; 5. Flow meter; 6. Acquisition module; 61. PE rigid pipe; 62. Silicone flexible hose; 63. Hollow fiber tube; 64. Support; 65. Quick connector. Detailed Implementation

[0041] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Unless otherwise defined, the technical terms used below have the same meaning as understood by those skilled in the art.

[0042] Unless otherwise specified, the test materials and reagents used in the following examples are commercially available or prepared by known methods.

[0043] Unless otherwise specified, all techniques or conditions described in the embodiments can be performed in accordance with the techniques or conditions described in the literature in this field or in the product manual. Unless otherwise specified, the quantitative experiments in the following embodiments are all repeated three times or more, and the results are averaged.

[0044] Example 1: 1.1 Specific preparation of the acquisition module: like Figure 1 As shown, a collection module is needed to extract root exudates, and a collection module 6 needs to be prepared.

[0045] like Figure 2 As shown, the first generation (such as Figure 2 (A) and second generation (such as) Figure 2 The equipment method for the B) acquisition module is as follows: First, cut the silicone tubing 62 into 3-5cm sections for later use. Pass both ends of the hollow fiber tube 63 through the cut silicone tubing 62 and insert it vertically into the PE rigid tube 61. Then, put the silicone tubing 62 onto the outside of the PE rigid tube 61 and seal it with slow-drying silicone XZX-8231. Let it air dry indoors.

[0046] The second-generation acquisition module requires the hollow fiber tube 63 of the first-generation acquisition module to be inserted into the bracket 64, which is made by cutting an acrylic plate.

[0047] The third-generation acquisition module (as shown in Figure C) requires the hollow fiber tube 63 to be cut to the required length. One side of the hollow fiber tube 63 is sealed with glue and allowed to dry. The glue used is CT-8121 type AB glue. The specific sealing method involves mixing the AB glue according to the configuration requirements and then using a vacuum pump to defoam it. This step is to prevent excessive air bubbles in the glue from affecting the sealing effect. Align one side of the prepared hollow fiber tube 63, immerse it in the glue for 5-10 seconds, and then hang it vertically for one day until dry. As needed, insert the un-glue side of the dried hollow fiber tube 63 into the quick connector 65, seal it with slow-drying silicone XZX-8231, and allow it to air dry indoors.

[0048] The initial steps for the fourth-generation acquisition module (as shown in Figure D) are the same as those for the third generation. After preparing the hollow fiber tubes 63, insert the glue-free side into the bracket 64, invert it vertically, and gather all the glue-free sides of the hollow fiber tubes 63 together. Secure them with cable ties, then use scissors or a guillotine to cut the glue-free side flat and insert it into the quick connector 65. Seal it with slow-drying silicone XZX-8231 and allow it to air dry indoors. The specific travel details and related brackets are as follows... Figure 2 and Figure 3 As shown.

[0049] 1.2 A device that combines the functions of collecting plant root exudates and transporting nutrient solution (structural schematic diagram shown in figure) Figure 1 As shown, the system includes a control module, a collection module, and a data acquisition module, all connected by pipelines. The control module includes an adjustable pressure vacuum pump 2 and a controller 1, with the adjustable pressure vacuum pump 2 electrically connected to the controller 1. The adjustable pressure vacuum pump 2 is equipped with a positive pressure port 21, a negative pressure port 22, and a pressure regulating valve. The positive pressure port 21 or the negative pressure port 22 is connected to a double-connector 3 via a gas supply pipe. The double-connector 3 is connected to a connector via a liquid supply pipe. A flow meter 5 is also installed on the liquid supply pipe. The controller 1, through a programmed control, controls the operating mode, pressure, and operating time of the adjustable pressure vacuum pump 2.

[0050] The collection module includes a container 4 and a double connector 3, which are fixedly connected to the container 4; the acquisition module 6 includes a hollow fiber tube 63 and a connector, which are fixedly connected to each other.

[0051] The connector consists of a rigid PE tube 61 and a flexible silicone tube 62. The rigid PE tube 61 is connected to the hollow fiber tube 63 through the flexible silicone tube 63; or the connector is a quick-connect fitting 65 with an integral structure, which is directly connected to the hollow fiber tube 63.

[0052] Hollow fiber tube 63 can be made into various shapes, such as circular shapes (e.g.) Figure 2 (A) Target-shaped (e.g.) Figure 2 (B) Multiple fiber bundles (such as...) Figure 2 (C, D) 1.3 A method for collecting plant root exudates and delivering nutrient solution, implemented based on the above-mentioned device that combines plant root exudate collection and nutrient solution delivery, includes the following steps: (1) Module assembly: Connect the control module, collection module, and acquisition module together and test the overall airtightness. If the airtightness is good, proceed to the next step. The acquisition module is buried in the planting substrate before operation, and then plant seeds are planted in the substrate. (2) Delivery of nutrient solution: Nutrient solution is added to the container, and the pumping pressure is set through the controller program. Nutrient solution is pumped in daily through an adjustable pressure vacuum pump. (3) Collect plant root exudates: At the corresponding stage of plant growth, root exudates are extracted. First, phosphate buffer is added to the container, and positive pressure is set using the controller program to inject the phosphate buffer into the plant substrate. After an interval of 5-10 minutes, negative pressure is set using the program to aspirate the root exudates. This extraction is repeated every 0.5-1 hour, for a total of 3-6 times to ensure thorough extraction. The collected root exudates are then concentrated using a vacuum freeze dryer to a volume of 3-5 ml, and stored. This is the plant root exudate, which can be sent to the company for testing of the plant's non-targeted metabolomics according to experimental requirements.

[0053] 1.4 The impact of the data acquisition module on root growth experiment This experiment used a third-generation collection module, the preparation method of which is as described in the previous experiment. Three different treatment groups were set up in this experiment, and three different water supply methods were set up using collection module 6: bilateral water supply (nutrients + water), bilateral water supply (even water supply on both sides), and control (top water supply), to investigate whether these methods would affect the root distribution during wheat growth.

[0054] This experiment uses vermiculite as a matrix, and the hollow fiber tube 63 of the acquisition module 6 is arranged according to... Figure 4 The pipes were laid out as shown, and the substrate was filled in. Four wheat plants were evenly planted on top of the substrate. The wheat variety was Zhenmai 16.

[0055] Before the experiment begins, the control module, collection module, and acquisition module are set up according to... Figure 1 Connect as shown. Set the daily water supply and operating time for controller 1. Through the data acquisition module, supply 15ml of water to each pot daily at a set time. After 30 days of growth, cut open the pots to observe the direction of wheat root secretions and weigh the fresh weight of the wheat roots.

[0056] Experimental results are as follows Figure 5As shown in the figure. The results show that in the control treatment, the wheat root system was more evenly distributed. However, in the bilateral water supply treatment group, wheat roots were found to be concentrated on the nutrient-supply side and relatively less concentrated on the water-supply side. In the left and right water supply treatment group, wheat roots were concentrated around the hollow fiber tube 63 of the collection module 6. The experiment shows that the root exudate collection module 6 can affect the distribution of wheat roots and cause them to concentrate around the hollow fiber tube 63 of the collection module 6.

[0057] Based on the placement of the hollow fiber tubes (63), each pot was divided into three equal-area sections (left, center, and right). The fresh weight of the roots from different parts of the plant under different treatments was then measured. The results are as follows: Figure 6 As shown, through significant difference analysis, it was found that the proportion of fresh weight of roots on the left and right sides of different treatment groups was significantly higher than that in the middle. The experimental results show that the collection module 6 can affect the orientation of the root system and make the roots accumulate around the collection module.

[0058] 1.5 Experiment on the effects of root exudate extraction on plant growth The main purpose of this experiment is to verify whether the device and method can effectively extract root exudates and whether the extraction will affect plant growth.

[0059] A collection module 6, which is a fourth-generation collection module, is placed inside a flowerpot. Wheat that has grown to the jointing stage is then transplanted into the flowerpot. The wheat roots are then manually wrapped evenly around the hollow fiber tube 63 of the collection module 6 to maximize the extraction of root secretions.

[0060] This experiment used field soil as the growth substrate. Before the experiment began, the control module, collection module, and sampling module were set up according to... Figure 1 Connect as shown.

[0061] The controller 1 is set to determine the daily water supply and operating time. Through the collection module, 15ml of water is supplied to each pot daily at a set time and in a set quantity. Samples are taken 7 days and 30 days after transplanting.

[0062] The experimental treatment groups are shown in Table 1. On day 7, root exudates were extracted using deionized water. The specific steps are as follows: First, 30 ml of deionized water was added to reagent bottle 4. The deionized water was then injected into the plant substrate via controller 1. After 10 minutes, the adjustable pressure vacuum pump 2 was automatically activated under the control of controller 1, drawing in root exudates at a negative pressure of -0.2 MPa. When flow meter 5 detected the aspiration of 15 ml of the mixed liquid containing root exudates, controller 1 automatically stopped the adjustable pressure vacuum pump 2. This operation was repeated every 0.5 hours for a total of 3 times to ensure thorough extraction of root exudates.

[0063] In addition, a blank control and a hydroponic extraction control were added to investigate the extraction efficiency of this device and method. The hydroponic extraction method used in this study was as follows: the transplanted wheat roots were cleaned with deionized water, placed in a reagent bottle wrapped with tin foil, and 30 ml of deionized water was added quantitatively. The root exudates were collected after 6 hours.

[0064] To test whether phosphate-buffered saline (PBS) extraction could effectively improve extraction efficiency, this experiment, based on previous research, added a treatment group using 1xPBS extraction on day 30. The extraction procedure was the same as that for deionized water extraction, only the deionized water was replaced with an equal volume of 1xPBS. Furthermore, the root exudates extracted on day 30 were concentrated using a freeze dryer, with each treatment frozen to approximately 5 ml.

[0065] Since root exudates are mostly composed of acids, ketones, and other organic substances, in order to detect whether this equipment and method can effectively extract root exudates, the content of soluble organic carbon (SOC) in the test liquid of all treated and extracted root exudates was quantitatively detected. The experimental results are shown in Table 1.

[0066] The results show that, equivalent to water extraction, this device and method can effectively extract root exudates. Furthermore, through a control analysis comparing extraction with and without plants, the device and method extracted higher levels of SOCs under plant treatment, indicating effective extraction of root exudates.

[0067] Results of root exudate extraction after 30 days showed that phosphate buffer extraction was more effective in extracting root exudates, and freeze-drying concentration increased the concentration of root exudates by 3 times.

[0068] To investigate whether phosphate buffer affects plant growth, this experiment continuously observed the growth status of wheat 14 days after extraction with phosphate buffer. Figure 7 The experimental results showed that extraction with phosphate buffer had no effect on plant growth. This experiment also investigated whether the soil environment would affect the collection module 6 itself over a long period. After the wheat matured, we extracted the deployed collection module from the soil. Air tightness testing showed that the soil environment had no effect on the performance of the collection module over a long period, further verifying the reliability of the device and method.

[0069] Table 1. Soluble organic carbon content in root exudates from different treatment groups

[0070] 1.6 Comparative Experiment of Root Exudates Extracted by Different Methods The preceding experiments have all demonstrated that this device and method can effectively extract root exudates. To further examine its extraction efficiency and the differences from existing mainstream extraction methods, further research is needed.

[0071] This experiment included three treatments: solution culture collection, soil culture collection, and extraction using a collection module. Each treatment had four parallel samples.

[0072] The data acquisition module extraction method processing group was pre-deployed with a fourth-generation data acquisition module 6. The selected wheat variety was Zhenmai 16, and the culture medium was field wheat soil.

[0073] Wheat was planted in small white buckets in each parallel sample, with 1 kg of soil in each bucket. The fertilizer ratio was 150 mg / kg nitrogen fertilizer (urea), 57.4 mg / kg phosphorus fertilizer (superphosphate), and 40.1 mg / kg potassium fertilizer (potassium sulfate).

[0074] Before the experiment, the seeds were disinfected with a 5% sodium hypochlorite solution for 5 minutes, rinsed with deionized water 3 times, and germinated at room temperature. The germinated seeds were then planted in small white buckets, with 4 seeds planted in each bucket.

[0075] Each parallel sample was replenished with water to 60% of its maximum field capacity, and water was replenished every two days. The treatment group using the collection module extraction method was irrigated through the collection module 6 to ensure that the wheat roots grew evenly around the hollow fiber tube 63 of the collection module 6, thereby improving the extraction efficiency of root exudates.

[0076] During the wheat flowering stage, root exudates are collected. The solution culture collection method mainly involves carefully separating the whole wheat plant from the soil for each parallel sample, while carefully washing the wheat roots with tap water until the soil adhering to the roots is clean. The roots are then rinsed with deionized water and placed in a 250ml reagent bottle protected from light by aluminum foil. 30ml of 1xPBS is added to the bottle, and the liquid is collected after 6 hours. This liquid is the root exudate extracted by the solution culture collection method.

[0077] The method for extracting root exudates by soil culture collection is as follows: the whole wheat plant under each parallel sample is carefully separated from the soil, and then the rhizosphere soil tightly attached to the wheat roots is collected, a total of 2g is collected, placed in a centrifuge tube, and then frozen and stored in a freezer at -80 degrees Celsius.

[0078] The extraction steps of the acquisition module extraction method are the same as those of the above experiments.

[0079] The root exudates extracted by the solution culture collection method and the collection module extraction method were placed in a freeze dryer for concentration and drying until about 5 ml of liquid remained. The liquid was then passed through a 0.22 μm water filter and placed in a brand new sterilized centrifuge tube for freezing, ready for sample delivery.

[0080] Root exudates extracted using three methods were sent to the company for non-targeted metabolomics analysis. The main instrument platform used was ultra-high performance liquid chromatography (UPLC): Waters UPLC Acquity I-Class PLUS; high-resolution mass spectrometry (HDMS): Waters UPLCXevo G2-XS QTof; chromatographic column: Acquity UPLC HSS T3 1.8um 2.1. 100mm, Waters. Specific analytical testing methods can be found in the relevant reference [Wang J, Zhang T, Shen X, et al. Serum metabolomics for early diagnosis of esophageal squamous cell carcinoma by UHPLC-QTOF / MS[J]. Metabolomics, 2016, 12(7):116.] William O. Slade,Emily G. Werth,Evan W. McConnell,Sophie Alvarez,Leslie M. Hicks. Quantifying Reversible Oxidation of Protein Thiols inPhotosynthetic Organisms[J]. Journal of The American Society for MassSpectrometry,2015,26(4). Tetrahydrobiopterin and alkylglycerol monooxygenase substantially alter the murine macrophage lipidome[J]. Proceedings of the National Academy of Sciences of the United States of America,2015,112(8). Kuhl C, Tautenhahn R, Böttcher C, et al. CAMERA: An IntegratedStrategy for Compound Spectra Extraction and Annotation of LiquidChromatography / Mass Spectrometry Data Sets[J]. Analytical Chemistry, 2012, 84(1):283-9.] Analysis of metabolomics data revealed differences in metabolite composition among the three treatment groups: solution culture (WW), soil culture (SS), and sampling module extraction (NN). Metabolite composition was matched and classified using the HMDB database, showing differences in species composition across the three treatment groups. However, the WW and NN treatment groups exhibited relatively similar species compositions. Figure 8 ).

[0081] PCoA analysis was performed on different treatment groups ( Figure 9 The analysis results showed that there were significant differences between the different treatment groups (Adonis analysis R). 2 =0.54 p<0.001), where the WW and NN treatment groups were similar in species composition.

[0082] Further analysis was conducted on the secretions of key components, and the differences in key components among different treatment groups were analyzed. Figure 10 The Venn diagram results show that the key components extracted by the three extraction methods have overlapping parts, but at the same time, the different extraction methods have extracted different key components to a certain extent.

[0083] The above experiments all demonstrate that this device and method can effectively extract root exudates, with extraction efficiency and accuracy similar to solution culture collection methods. However, it can also supplement key metabolite components that cannot be extracted by solution culture collection methods. This device is reliable and can be buried in substrates such as soil for extended periods without damage or impact on extraction efficiency. Furthermore, the extraction method does not affect the plant itself, enabling in-situ extraction of root exudates throughout the plant's life cycle. This device and method are intelligent, efficient, and continuous, greatly improving extraction efficiency and flexibility.

[0084] Compared with existing technologies, the advantages of this collection device and method are that it can efficiently extract plant root exudates without affecting the plant's growth status, and it can extract root exudates throughout the entire plant growth cycle.

[0085] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. 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. Such 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. A device that combines the functions of collecting plant root exudates and transporting nutrient solution, characterized in that, It includes a control module, a collection module and a collection module, and the modules are connected by pipelines; the control module includes an adjustable pressure vacuum pump (2) and a controller (1), and the adjustable pressure vacuum pump (2) is electrically connected to the controller (1); the collection module includes a container (4) and a double-ended connector (3), and the double-ended connector (3) is fixedly connected to the container (4); the collection module (6) includes a hollow fiber tube (63) and a connector, and the hollow fiber tube (63) and the connector are fixedly connected.

2. The apparatus according to claim 1, characterized in that, The adjustable pressure vacuum pump (2) is provided with a positive pressure port (21), a negative pressure port (22) and a pressure regulating valve; the positive pressure port (21) or the negative pressure port (22) is connected to the double-connector (3) through a gas supply pipe; the double-connector (3) is connected to the connector through a liquid supply pipe; the double-connector (3) is used to connect the adjustable pressure vacuum pump (2) and the acquisition module (6), and plays the role of sealing and transmitting liquid.

3. The apparatus according to claim 1, characterized in that, The controller (1) is responsible for controlling the working mode, pressure, and working time of the adjustable vacuum pump (2) by writing a program.

4. The apparatus according to claim 1, characterized in that, The collection module is used to store nutrient solution and root exudates; the control module is used to control the pressure and time; the collection module (6) is used to deliver nutrient solution to plant roots and extract plant root exudates.

5. The apparatus according to claim 1, characterized in that, The connector consists of a PE rigid tube (61) and a silicone flexible tube (62), and the PE rigid tube (61) is connected to the hollow fiber tube (63) through the silicone flexible tube (62).

6. The apparatus according to claim 1, characterized in that, The connector is a quick-connect fitting (65) with an integral structure, which is directly connected to the hollow fiber tube (63).

7. The apparatus according to claim 1, characterized in that, The membrane material of the hollow fiber tube (63) is polyvinylidene fluoride, and the inner lining material is polyester; the shape of the hollow fiber tube is circular, target-shaped, or multiple fiber bundles.

8. A method for collecting plant root exudates and transporting nutrient solution, characterized in that, The implementation of the device for collecting and delivering nutrient solution from plant root exudates as described in any one of claims 1-7 includes the following steps: (1) Module assembly: Connect the control module, collection module, and acquisition module together and test the overall airtightness. If the airtightness is good, proceed to the next step. The acquisition module is buried in the planting substrate before operation, and then plant seeds are planted in the substrate. (2) Delivery of nutrient solution: Nutrient solution is added to the container, and the pumping pressure is set through the controller program. Nutrient solution is pumped in daily through an adjustable pressure vacuum pump. (3) Collect plant root exudates: At the corresponding stage of plant growth, root exudates are extracted. First, phosphate buffer is added to the container, and positive pressure is set through the controller program to inject phosphate buffer into the plant substrate. After a period of time, negative pressure is set through the program to aspirate the root exudates.

9. The method according to claim 8, characterized in that, In step (3), the time period is specifically 5 to 10 minutes; the root exudate is extracted again after an interval of 0.5 to 1 hour, and this is done 3 to 6 times in total.

10. The method according to claim 8, characterized in that, The collected root exudates were concentrated and preserved using a vacuum freeze dryer.

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