Plant callus culture device

Abstract

The invention relates to the technical field of callus culture, in particular to a plant callus culture device which comprises a culture unit and a detection unit, and the detection unit detects the culture temperature of a culture medium, the oxygen content of the culture medium and the temperature of the environment where the culture medium is located; the image unit is used for acquiring an image of the callus in the culture medium to determine the volume change of the callus; the analysis unit controls the cultivation temperature to change periodically to adjust the hormone concentration, determines the induction trend and growth stage of the calluses in a plurality of grids, correspondingly determines the light quality ratio and COpulse parameters, and determines whether light breathing needs to be inhibited or not and whether cooperative triggering conditions are met or not; and the adjusting unit starts and closes the external temperature control device, executes a first adjusting instruction to drive the burette to move, starts the spectrum light source and the injection valve according to the determined illumination ratio and COpulse parameters, and starts the piezoelectric ceramic array according to the cooperative triggering condition. The method is matched with the dynamic physiology of the plant callus in a dynamic culture mode.

Description

Technical Field

[0001] The invention relates to the technical field of callus culture, in particular to a plant callus culture device. Background Art

[0002] The plant callus culture apparatus is an experimental equipment system used to induce, cultivate, and maintain plant callus under sterile conditions. Plant tissue culture involves isolating ex vivo plant tissue (organs or cells) and culturing them in culture medium under sterile conditions. Using specific conditions, cells undergo dedifferentiation, causing previously differentiated and functional cells to deviate from their original developmental path, losing their original state and function and returning to an undifferentiated callus state. This process is achieved by adding plant growth regulators to the culture medium, which prompt differentiated cells to alter their metabolic pathways and prepare for the re-initiation of cell division. Once callus is formed from the explant, the ratio of plant growth regulators can be adjusted to promote the differentiation of shoots and roots.

[0003] Chinese Patent Publication No.: CN209201779U discloses a plant callus subculture and proliferation culture device, including a double-screw culture tube, the upper screw mouth is provided with an exhaust and pressure relief port and a culture medium replenishing port, and the lower screw mouth is provided with a culture medium discharge port; a multi-port stirring and mixing container, having a culture medium return port, an air input port and a culture medium output port, a return liquid pipe inserted into the culture medium return port, the return liquid pipe bypasses a return liquid peristaltic pump and is then connected to the culture medium discharge port; an air pipe is inserted into the air input port, the outer end of the air pipe is connected to positive pressure sterile air; an infusion pipe is inserted into the culture medium output port, the infusion pipe bypasses an infusion peristaltic pump and is then connected to the culture medium replenishing port; a stirring device is used to fully mix the culture medium liquid in the multi-port stirring and mixing container with sterile air. It can be seen that the plant callus subculture and proliferation culture device has the following problems:

[0004] The culture device adopts a static culture method, and the static environment generated is often inconsistent with the dynamic physiology of plant callus;

[0005] The culture process creates hormone redundancy in order to maintain a static environment for the callus tissue, while increasing the carbon dioxide concentration to inhibit photorespiration causes carbon loss. Summary of the Invention

[0006] To this end, the present invention provides a plant callus cultivation device to overcome the problems in the prior art of mismatch between the static environment of the cultivation device and the dynamic physiology of the plant callus, which results in hormone redundancy and carbon loss.

[0007] To achieve the above-mentioned object, the present invention provides a plant callus cultivation device, comprising:

[0008] A cultivation unit comprising a base, a cultivation cover, a cultivation wall and a culture dish;

[0009] a detection unit connected to the analysis unit and configured to detect in real time the culture temperature of the culture medium, the oxygen content of the culture medium, and the ambient temperature of the culture device;

[0010] an imaging unit connected to the analyzing unit and configured to acquire an image of the callus tissue in the culture medium and determine a volume change of the callus tissue;

[0011] The analysis unit is used to control the cultivation temperature to change periodically according to the ambient temperature, adjust the hormone concentration according to the periodic conditions, determine the induction trend and growth stage of the callus tissue in several grids according to the volume change, and determine the light quality ratio and Pulse parameters are combined with the oxygen content of the culture medium to determine whether photorespiration needs to be inhibited and whether there are synergistic triggering conditions;

[0012] The regulating unit is connected to the regulating unit and is used to open and close the external temperature control device according to the transmission signal of the analysis unit, execute the first regulating instruction to drive the burette to move and adjust the hormone concentration according to the determined light ratio and The pulse parameters turn on the spectrum light source and the injection valve, and turn on the piezoelectric ceramic array according to the coordinated triggering conditions.

[0013] Furthermore, the base is located at the bottom of the cultivation device, and an external temperature control device is provided inside the base. The cultivation wall is a transparent structure connected to the base. The cultivation cover is located on the top of the cultivation wall. The culture dish is located inside the cultivation wall and on the top of the base.

[0014] The inner wall and bottom of the culture dish are provided with grid scales and height scales, and the interior of the culture cover is provided with a moving device, a moving rack and a spectrum light source.

[0015] The moving device is connected to the micro camera and the burette, and can drive the micro camera and the burette to move on the moving frame;

[0016] The outer cover wall of the cultivation cover is provided with a display device, and the display device is used to display the cultivation temperature and light quality ratio;

[0017] Wherein, the spectral light source is a multi-spectral LED array.

[0018] Furthermore, the analysis unit calculates the difference between the ambient temperature and the standard temperature.

[0019] If the difference between the ambient temperature and the standard temperature is greater than the difference evaluation value, the analyzing unit sends a signal to the regulating unit to regulate the power of the external temperature control device so that the difference between the internal temperature and the culture temperature is within the standard deviation value;

[0020] If the difference between the ambient temperature and the standard temperature is less than or equal to the difference evaluation value, the analyzing unit sends a signal to the regulating unit to turn off the external temperature control device.

[0021] Furthermore, if the difference between the ambient temperature and the standard temperature is less than or equal to the difference evaluation value,

[0022] The analysis unit controls the incubation temperature to change periodically. If the incubation temperature is lower than the lower limit temperature of the cycle, the analysis unit determines that the incubation temperature does not meet the periodic condition;

[0023] If the cultivation temperature rises to the upper limit temperature of the cycle within the initial cycle, the analysis unit determines that the cultivation temperature meets the periodic conditions and is in the rising stage, and outputs the grid control adjustment unit where the callus tissue is in the first induction trend to execute the first adjustment instruction.

[0024] Furthermore, the first regulating instruction is to drive the burette to move to the grid where the callus tissue in the first induction trend is located, and titrate the corresponding growth regulator or cytokinin to regulate the hormone concentration in the culture dish.

[0025] Furthermore, the analysis unit calculates the average growth rate of all callus tissues and the volume growth rate of callus tissues in each grid according to the volume change;

[0026] If the volume growth rate is greater than 0.7 times the average growth rate, the analysis unit determines that the callus tissue within the grid is in the second induction trend;

[0027] If the volume growth rate is less than or equal to 0.7 times the average growth rate, the analysis unit determines that the callus tissue in the grid is in the first induction trend and outputs the grid to the adjustment unit.

[0028] Furthermore, the analysis unit determines the growth stage of the callus tissue according to the average growth rate.

[0029] If the average growth rate is less than the first standard growth rate, the analysis unit determines that the growth stage of the callus tissue is the induction stage;

[0030] If the average growth rate is greater than or equal to the first standard growth rate and less than the second standard growth rate, the analysis unit determines that the growth stage of the callus tissue is the proliferation stage;

[0031] If the average growth rate is greater than or equal to the second standard growth rate, the analysis unit determines that the callus tissue is in the differentiation stage.

[0032] Furthermore, the analysis unit determines the light quality ratio of red and blue light in the spectral light source and the injection valve according to different growth stages of rice callus. Pulse parameters;

[0033] If the oxygen content of the culture medium is less than the critical content, the analysis unit determines that the respiratory intensity of the callus tissue exceeds the range and needs to inhibit photorespiration, and the adjustment unit turns on the multi-spectrum LED array for blue light irradiation and turns on the injection valve to spray conduct pulse;

[0034] If the oxygen content of the culture medium is greater than or equal to the critical content, the regulating unit turns on the multi-spectrum LED array for red light irradiation to promote light system repair.

[0035] Furthermore, the analysis unit sends a signal to the adjustment unit according to different growth stages of the callus tissue, and turns on the spectrum light source according to the first light quality ratio and the second light quality ratio for irradiation, and turns on the injection valve according to the pulse parameters to spray. ;

[0036] During the induction period, the regulating unit turns on the multi-spectrum LED array to perform main blue light irradiation according to the first light quality ratio;

[0037] When red light irradiation is performed during the differentiation period, the regulating unit turns on the multi-spectrum LED array to perform main red light irradiation according to the second light quality ratio.

[0038] Furthermore, when the cooperative triggering condition is met, the analysis unit controls the adjustment unit to activate the piezoelectric ceramic array to apply mechanical stress vibration to the callus tissue in the culture medium;

[0039] The synergistic triggering condition is that the oxygen content of the culture medium is less than the critical content, the callus tissue is in the proliferation stage, and the culture temperature is in the rising stage.

[0040] Compared with the existing technology, the beneficial effect of the present invention is that during the natural growth process of rice, the synthesis and degradation of hormones (such as IAA, ZR, and ABA) fluctuate in a circadian rhythm. The present device uses an external temperature control device to achieve periodic temperature fluctuations in rice callus tissue, simulating the impact of the diurnal temperature difference (such as warming during the day and cooling at night) on the endogenous hormone rhythm in the natural environment of rice. The temperature cycle is synchronized with the glycolysis oscillation cycle of rice cells (about 5-7 hours), which can enhance energy utilization efficiency. At the same time, it avoids the cultivation defect of constant hormone concentration in traditional culture that leads to cell "desensitization" (receptor downregulation). Pulse release maintains cell responsiveness and breaks the steady-state culture mode. Compared with the constant temperature cultivation in the existing technology, the proportion of S-phase cells in callus tissue is greatly increased and the browning rate is greatly reduced. The core purpose is to precisely regulate the biological response (hormone release) through non-natural but biomimetic physical stimulation (temperature), thereby increasing the induction success rate of rice callus tissue cultivation.

[0041] Furthermore, the device integrates a millimeter-level grid scale at the bottom of the culture bottle and combines it with image recognition technology to achieve non-destructive measurement of callus volume. The grid in which the callus is located is determined by the grid scale and the image of the callus, and the volume growth rate of the callus in the grid is calculated based on the volume change of the callus. The induction trend of the callus in the grid is determined based on the average growth rate, providing a target grid to be moved to for the subsequent adjustment unit to execute the first adjustment instruction; at the same time, the analysis unit determines the growth stage of the callus based on the average growth rate of the callus, providing a basis for the subsequent adjustment of the light quality ratio.

[0042] Furthermore, rice callus activates the photorespiration pathway under light, leading to carbon flow to glycine / serine rather than biomass accumulation, requiring high concentrations of Inhibit photorespiration, but also inhibit cell proliferation; in traditional methods, the light intensity is fixed or a simple day-night cycle The concentration is constant. This device promotes proliferation through blue light and differentiation through red light. Pulse optimization of carbon metabolism improves the carbon fixation efficiency of callus culture, and coupled with the above-mentioned hormone concentration regulation of phosgene to promote cell cycle synchronization, so that more cells are at the peak of energy demand, and the metabolic activity signal is captured by the oxygen content of the culture medium to trigger Compared with static culture, pulse precise carbon supplementation not only has higher induction efficiency but also increases callus dry weight faster.

[0043] Furthermore, traditional cognition holds that mechanical stress mainly promotes cell differentiation, but in rice callus tissue it exhibits the counterintuitive effect of inhibiting differentiation and enhancing pluripotency. This device uses a piezoelectric ceramic array to generate micro-vibrations to simulate the periodic wind or raindrop impacts on plants in nature, which can stimulate and activate rice callus tissue to achieve more efficient dedifferentiation and regeneration. The core lies in breaking the "static culture" paradigm and using dynamic physical signals to optimize cell behavior.

[0044] Furthermore, the timing of starting the external temperature control device is determined by comparing the difference between the ambient temperature and the standard temperature with the difference evaluation value, thereby eliminating the adverse effects of the external ambient temperature on the subsequent temperature regulation to achieve periodic changes in the cultivation temperature. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] Figure 1 This is a unit connection diagram of a plant callus cultivation device according to an embodiment of the present invention;

[0046] Figure 2 This is a schematic diagram of the main structure of the cultivation unit in an embodiment of the present invention;

[0047] Figure 3 This is a bottom-up structural diagram of a culture cover according to an embodiment of the present invention;

[0048] Figure 4 Schematic diagram of the front structure of the culture cover in an embodiment of the present invention;

[0049] In the figure: 1-base, 2-culture dish, 3-culture cover, 4-culture wall, 21-grid scale, 22-height scale, 31-burette, 32-display device, 33-movable stand, 34-spectral light source, 35-moving device. DETAILED DESCRIPTION

[0050] In order to make the objects and advantages of the present invention more clearly understood, the present invention is further described below in conjunction with embodiments; it should be understood that the specific embodiments described herein are merely used to explain the present invention and are not intended to limit the present invention.

[0051] The preferred embodiments of the present invention are described below with reference to the accompanying drawings. It should be understood by those skilled in the art that these embodiments are only used to explain the technical principles of the present invention and are not intended to limit the scope of protection of the present invention.

[0052] It should be noted that, in the description of the present invention, terms such as "up", "down", "left", "right", "inside", and "outside" indicating directions or positional relationships are based on the directions or positional relationships shown in the accompanying drawings. This is only for the convenience of description and does not indicate or imply that the device or element must have a specific orientation, be constructed and operated in a specific orientation. Therefore, it cannot be understood as a limitation on the present invention.

[0053] Furthermore, it should be noted that, in the description of the present invention, unless otherwise expressly specified or limited, the terms "mounted," "connected," and "connected" should be understood in a broad sense. For example, they may refer to fixed connections, detachable connections, or integral connections; mechanical connections or electrical connections; direct connections or indirect connections through an intermediate medium; and internal communication between two components. Those skilled in the art will understand the specific meanings of the above terms in the present invention based on specific circumstances.

[0054] See also Figure 1-Figure 4 As shown, Figure 1 This is a unit connection diagram of a plant callus cultivation device according to an embodiment of the present invention; Figure 2 This is a schematic diagram of the main structure of the cultivation unit in an embodiment of the present invention; Figure 3 This is a bottom-up structural diagram of a culture cover according to an embodiment of the present invention; Figure 4 Schematic diagram of the front structure of the culture cover in an embodiment of the present invention.

[0055] The present invention provides a plant callus cultivation device, comprising:

[0056] The cultivation unit comprises a base 1, a cultivation cover 3, a cultivation wall 4 and a culture dish 2;

[0057] a detection unit connected to the analysis unit and configured to detect in real time the culture temperature of the culture medium, the oxygen content of the culture medium, and the ambient temperature of the culture device;

[0058] an imaging unit connected to the analyzing unit and configured to acquire an image of the callus tissue in the culture medium and determine a volume change of the callus tissue;

[0059] The analysis unit is used to control the cultivation temperature to change periodically according to the ambient temperature, adjust the hormone concentration according to the periodic conditions, determine the induction trend and growth stage of the callus tissue in several grids according to the volume change, and determine the light quality ratio and Pulse parameters are combined with the oxygen content of the culture medium to determine whether photorespiration needs to be inhibited and whether there are synergistic triggering conditions;

[0060] The regulating unit is connected to the regulating unit and is used to open and close the external temperature control device according to the transmission signal of the analysis unit, execute the first regulating instruction to drive the burette to move and adjust the hormone concentration according to the determined light ratio and The pulse parameters turn on the spectrum light source and the injection valve, and turn on the piezoelectric ceramic array according to the coordinated triggering conditions.

[0061] The base 1 is located at the bottom of the cultivation device and is provided with an external temperature control device (not shown) inside. The cultivation wall 4 is a transparent structure connected to the base 1. The cultivation cover 3 is located on the top of the cultivation wall 4. The culture dish 2 is located inside the cultivation wall 4 and on the top of the base 1.

[0062] The inner wall and bottom of the culture dish 2 are provided with a grid scale 21 and a height scale 22. The interior of the culture cover 3 is provided with a moving device 35, a moving frame 33 and a spectrum light source 34.

[0063] The moving device 35 is connected to the micro camera 36 and the burette 31, and can drive the micro camera 36 and the burette 31 to move on the moving frame 33;

[0064] The outer cover wall of the cultivation cover 3 is provided with a display device 32, and the display device 32 is used to display the cultivation temperature and light quality ratio;

[0065] Wherein, the spectral light source is a multi-spectral LED array.

[0066] A piezoelectric ceramic array is also provided inside the base, which can generate a small deformation under the action of an electric field to cause vibration impact. The culture medium is located in the culture dish;

[0067] The cultivation device can also be connected to the injection valve through a through hole provided on the top of the cultivation cover 3, and the required hormone solution can also be added through the through hole;

[0068] In this embodiment, the plant callus culture device is a rice callus culture device, which performs callus culture on rice.

[0069] The detection unit detects the culture temperature of the culture medium and the ambient temperature of the culture device in real time through a temperature sensor, and transmits the culture temperature and the ambient temperature to the analysis unit;

[0070] The analysis unit analyzes the culture temperature and the ambient temperature. If the difference between the ambient temperature and the standard temperature is greater than the difference evaluation value,

[0071] The analysis unit then adjusts the power of the external temperature control device according to the culture temperature and the ambient temperature, and sends a signal to the adjustment unit to control the difference between the internal temperature and the culture temperature to be within the standard deviation value;

[0072] If the difference between the ambient temperature and the standard temperature is less than or equal to the difference evaluation value, the analysis unit sends a signal to the adjustment unit to turn off the external temperature control device;

[0073] Wherein, the standard temperature is 25°C, and the difference evaluation value is 3.

[0074] In practice, the external temperature control device is a thermoelectric cooler located inside the base.

[0075] Specifically, the timing of starting the external temperature control device is determined by comparing the difference between the ambient temperature and the standard temperature with the difference evaluation value, eliminating the adverse effects of the external ambient temperature on the subsequent temperature control to achieve periodic changes in the cultivation temperature.

[0076] When the difference between the ambient temperature and the standard temperature is less than or equal to the difference evaluation value, the analyzing unit controls the culture temperature to change periodically based on the detected culture temperature of the culture medium in the culture device, and controls the regulating unit to periodically regulate the hormone concentration in the culture dish;

[0077] The analysis unit controls the regulation unit to adjust the hormone concentration in the culture medium according to the periodicity of the temperature fluctuation in the device.

[0078] When the incubation temperature is lower than the lower limit temperature of the cycle, the analysis unit determines that the incubation temperature does not meet the periodic condition;

[0079] When the cultivation temperature rises to the upper limit temperature of the cycle in the initial cycle, the analysis unit determines that the cultivation temperature meets the periodic condition and is in the rising stage, and outputs the grid where the callus tissue in the first induction trend is located to the control adjustment unit to execute the first adjustment instruction;

[0080] The initial period is 3-6 hours, the upper limit temperature of the period is 30°C, and the lower limit temperature of the period is 28°C.

[0081] The first adjustment instruction is to drive the burette to move to the grid where the callus tissue in the first induction trend is located, and titrate the corresponding growth regulator or cytokinin to adjust the hormone concentration in the culture dish;

[0082] The titration amount of the growth regulator or cytokinin is equal to the product of the difference between the average growth rate and the volume growth rate and the correction coefficient, and the correction coefficient can be adjusted within the range of 0.5-1.5 according to the specific difference between the average growth rate and the volume growth rate;

[0083] Specifically, the analysis unit controls the periodic change of the incubation temperature by actively adjusting the temperature through an external temperature control device.

[0084] Specifically, during the natural growth of rice, the synthesis and degradation of hormones (such as IAA, ZR, and ABA) fluctuate in a circadian rhythm. This device uses an external temperature control device to achieve periodic temperature fluctuations in rice callus tissue, simulating the impact of the day and night temperature difference (such as warming during the day and cooling at night) in the natural environment of rice on the rhythm of endogenous hormones. This synchronizes the temperature cycle with the glycolysis oscillation cycle of rice cells (about 5-7 hours), which can enhance energy utilization efficiency. At the same time, it avoids the cultivation defect of constant hormone concentration in traditional culture that leads to cell "desensitization" (receptor downregulation). Pulse release maintains cell responsiveness and breaks the steady-state culture mode. Compared with the constant temperature cultivation in the existing technology, the proportion of S-phase cells in the callus tissue is greatly increased, and the browning rate is greatly reduced. The core purpose is to accurately regulate the biological response (hormone release) through non-natural but bionic physical stimulation (temperature), thereby increasing the induction success rate of rice callus cultivation.

[0085] The inner wall and bottom of the culture dish are provided with grid scales and height scales. The imaging unit calculates the volume change of the callus tissue according to the grid scales and height scales. The analyzing unit distinguishes the callus tissue with a slower callus healing trend according to the volume changes of the callus tissues and outputs the grid in which the callus tissue is located to the regulating unit.

[0086] The imaging unit acquires images of the callus tissue in each grid using a micro camera according to an initial detection period, determines a volume change of the callus tissue based on the images, the grid scale, and the height scale, and the analyzing unit calculates an average growth rate of all callus tissues and a volume growth rate of the callus tissue in each grid based on the volume change;

[0087] If the volume growth rate is greater than 0.7 times the average growth rate, the analysis unit determines that the callus tissue within the grid is in the second induction trend;

[0088] If the volume growth rate is less than or equal to 0.7 times the average growth rate, the analysis unit determines that the callus tissue in the grid is in the first induction trend and outputs the grid to the adjustment unit.

[0089] The analyzing unit determines the growth stage of the callus according to the average growth rate,

[0090] If the average growth rate is less than the first standard growth rate, the analysis unit determines that the growth stage of the callus tissue is the induction stage;

[0091] If the average growth rate is greater than or equal to the first standard growth rate and less than the second standard growth rate, the analysis unit determines that the growth stage of the callus tissue is the proliferation stage;

[0092] If the average growth rate is greater than or equal to the second standard growth rate, the analysis unit determines that the growth stage of the callus tissue is the differentiation stage;

[0093] The first standard growth rate is 0.2 mm³ / day, and the second standard growth rate is 0.5 mm³ / day.

[0094] Specifically, this device integrates a millimeter-level grid scale at the bottom of the culture bottle and combines it with image recognition technology to achieve non-destructive measurement of callus volume. The grid in which the callus is located is determined by the grid scale and the image of the callus, and the volume growth rate of the callus in the grid is calculated based on the volume change of the callus. The induction trend of the callus in the grid is determined based on the average growth rate, providing a target grid to move to for the subsequent adjustment unit to execute the first adjustment instruction; at the same time, the analysis unit determines the growth stage of the callus based on the average growth rate of the callus, providing a basis for the subsequent adjustment of the light quality ratio.

[0095] The analysis unit determines the light quality ratio of red and blue light in the spectrum light source and the injection valve according to different growth stages of rice callus. Pulse parameters;

[0096] The detection unit uses a dissolved oxygen sensor to monitor the respiration intensity of the callus tissue in real time, and the analysis unit monitors the oxygen content of the culture medium in real time based on the dissolved oxygen sensor data.

[0097] If the oxygen content of the culture medium is less than the critical content, the analysis unit determines that the respiratory intensity of the callus tissue exceeds the range and needs to inhibit photorespiration, and the adjustment unit turns on the multi-spectrum LED array for blue light irradiation and turns on the injection valve to spray conduct pulse;

[0098] If the oxygen content of the culture medium is greater than or equal to the critical content, the regulating unit turns on the multi-spectrum LED array for red light irradiation to promote light system repair;

[0099] During implementation, the analysis unit sends a signal to the adjustment unit according to the different growth stages of the callus tissue, turns on the multi-spectrum LED array according to the first light quality ratio and the second light quality ratio, and turns on the injection valve to spray according to the pulse parameters. ;

[0100] During the induction period, the regulating unit turns on the multi-spectrum LED array to perform main blue light irradiation according to the first light quality ratio;

[0101] During the differentiation period, the regulating unit turns on the multi-spectrum LED array to perform red light main irradiation according to the second light quality ratio;

[0102] The first light quality ratio is blue light: red light equal to 7:3, and the second light quality ratio is blue light: red light equal to 3:7;

[0103] The critical content is 4 mg / L, the blue and red light of the LED lights of the spectrum light source are controllable, The pulse parameters of the pulse include the initial pulse duration and the initial concentration, the initial pulse duration is 5min, the initial The concentration is 1000ppm.

[0104] Specifically, rice callus activates the photorespiration pathway under light, causing carbon to flow to glycine / serine rather than biomass accumulation, requiring high concentrations of Inhibit photorespiration, but also inhibit cell proliferation; in traditional methods, the light intensity is fixed or a simple day-night cycle The concentration is constant. This device promotes proliferation through blue light and differentiation through red light. Pulse optimization of carbon metabolism improves the carbon fixation efficiency of callus culture, and coupled with the above-mentioned hormone concentration regulation of phosgene to promote cell cycle synchronization, so that more cells are at the peak of energy demand, and the metabolic activity signal is captured by the oxygen content of the culture medium to trigger Compared with static culture, pulse precise carbon supplementation not only has higher induction efficiency but also increases callus dry weight faster.

[0105] When the analysis unit meets the cooperative triggering conditions, the control adjustment unit turns on the piezoelectric ceramic array to apply mechanical stress vibration to the callus tissue in the culture medium to inhibit the edge differentiation of the callus tissue, and adjusts the blue light intensity and Concentration and vibration work synergistically;

[0106] In this embodiment, the piezoelectric vibration parameters of the piezoelectric ceramic array are a frequency of 50 Hz and an amplitude of 10 μm;

[0107] Specifically, the synergistic triggering condition is that the oxygen content of the culture medium is less than the critical content, the callus tissue is in the proliferation stage, and the culture temperature is in the rising stage.

[0108] Specifically, traditional cognition holds that mechanical stress mainly promotes cell differentiation, but in rice callus it exhibits the counterintuitive effect of inhibiting differentiation and enhancing pluripotency. This device uses a piezoelectric ceramic array to generate micro-vibrations to simulate the periodic wind or raindrop impacts on plants in nature, which can stimulate and activate rice callus to achieve more efficient dedifferentiation and regeneration. The core lies in breaking the "static culture" paradigm and using dynamic physical signals to optimize cell behavior.

[0109] Thus far, the technical solutions of the present invention have been described in conjunction with the preferred embodiments shown in the accompanying drawings. However, it will be readily understood by those skilled in the art that the scope of protection of the present invention is obviously not limited to these specific embodiments. Without departing from the principles of the present invention, those skilled in the art may make equivalent changes or substitutions to the relevant technical features, and the technical solutions after such changes or substitutions will fall within the scope of protection of the present invention.

[0110] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Those skilled in the art will readily appreciate that the present invention is susceptible to various modifications and variations. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention are intended to be within the scope of protection of the present invention.

Claims

1. A plant callus cultivation device, characterized in that: include: A cultivation unit comprising a base, a cultivation cover, a cultivation wall and a culture dish; a detection unit connected to the analysis unit and configured to detect in real time the culture temperature of the culture medium, the oxygen content of the culture medium, and the ambient temperature of the culture device; an imaging unit connected to the analyzing unit and configured to acquire an image of the callus tissue in the culture medium and determine a volume change of the callus tissue; The analysis unit is used to control the cultivation temperature to change periodically according to the ambient temperature, adjust the hormone concentration according to the periodic conditions, determine the induction trend and growth stage of the callus tissue in several grids according to the volume change, and determine the light quality ratio and Pulse parameters are combined with the oxygen content of the culture medium to determine whether photorespiration needs to be inhibited and whether synergistic triggering conditions are met; The regulating unit is connected to the regulating unit and is used to open and close the external temperature control device according to the transmission signal of the analysis unit, execute the first regulating instruction to drive the burette to move and adjust the hormone concentration according to the determined light ratio and The pulse parameters turn on the spectrum light source and the injection valve, and turn on the piezoelectric ceramic array according to the coordinated triggering conditions.

2. The plant callus cultivation device according to claim 1, characterized in that: The base is located at the bottom of the cultivation device, and an external temperature control device is provided inside the base. The cultivation wall is a transparent structure connected to the base. The cultivation cover is located on the top of the cultivation wall. The culture dish is located inside the cultivation wall and on the top of the base. The inner wall and bottom of the culture dish are provided with grid scales and height scales, and the interior of the culture cover is provided with a moving device, a moving rack and a spectrum light source. The moving device is connected to the micro camera and the burette, and can drive the micro camera and the burette to move on the moving frame; The outer cover wall of the cultivation cover is provided with a display device, and the display device is used to display the cultivation temperature and light quality ratio; Wherein, the spectral light source is a multi-spectral LED array.

3. The plant callus cultivation device according to claim 1, characterized in that: The analysis unit calculates the difference between the ambient temperature and the standard temperature. If the difference between the ambient temperature and the standard temperature is greater than the difference evaluation value, the analyzing unit sends a signal to the regulating unit to regulate the power of the external temperature control device so that the difference between the internal temperature and the culture temperature is within the standard deviation value; If the difference between the ambient temperature and the standard temperature is less than or equal to the difference evaluation value, the analyzing unit sends a signal to the regulating unit to turn off the external temperature control device.

4. The plant callus cultivation device according to claim 3, characterized in that: If the difference between the ambient temperature and the standard temperature is less than or equal to the difference evaluation value, The analysis unit controls the incubation temperature to change periodically. If the incubation temperature is lower than the lower limit temperature of the cycle, the analysis unit determines that the incubation temperature does not meet the periodic condition; If the cultivation temperature rises to the upper limit temperature of the cycle within the initial cycle, the analysis unit determines that the cultivation temperature meets the periodic conditions and is in the rising stage, and outputs the grid control adjustment unit where the callus tissue is in the first induction trend to execute the first adjustment instruction.

5. The plant callus cultivation device according to claim 4, characterized in that: The first regulating instruction is to drive the burette to move to the grid where the callus tissue in the first induction trend is located, and titrate the corresponding growth regulator or cytokinin to regulate the hormone concentration in the culture dish.

6. The plant callus cultivation device according to claim 5, characterized in that: The analysis unit calculates the average growth rate of all callus tissues and the volume growth rate of callus tissues in each grid according to the volume change; If the volume growth rate is greater than 0.7 times the average growth rate, the analysis unit determines that the callus tissue within the grid is in the second induction trend; If the volume growth rate is less than or equal to 0.7 times the average growth rate, the analysis unit determines that the callus tissue in the grid is in the first induction trend and outputs the grid to the adjustment unit.

7. The plant callus cultivation device according to claim 6, characterized in that: The analyzing unit determines the growth stage of the callus according to the average growth rate, If the average growth rate is less than the first standard growth rate, the analysis unit determines that the growth stage of the callus tissue is the induction stage; If the average growth rate is greater than or equal to the first standard growth rate and less than the second standard growth rate, the analysis unit determines that the growth stage of the callus tissue is the proliferation stage; If the average growth rate is greater than or equal to the second standard growth rate, the analysis unit determines that the growth stage of the callus tissue is the differentiation stage.

8. The plant callus cultivation device according to claim 1, characterized in that: The analysis unit determines the light quality ratio of red and blue light in the spectrum light source and the injection valve according to different growth stages of rice callus. Pulse parameters; If the oxygen content of the culture medium is less than the critical content, the analysis unit determines that the respiratory intensity of the callus tissue exceeds the range and needs to inhibit photorespiration, and the adjustment unit turns on the multi-spectrum LED array for blue light irradiation and turns on the injection valve to spray conduct pulse; If the oxygen content of the culture medium is greater than or equal to the critical content, the regulating unit turns on the multi-spectrum LED array for red light irradiation to promote light system repair.

9. The plant callus cultivation device according to claim 8, characterized in that: The analysis unit sends a signal to the adjustment unit according to different growth stages of the callus tissue, and turns on the spectrum light source according to the first light quality ratio and the second light quality ratio for irradiation, and turns on the injection valve according to the pulse parameters to spray. ; During the induction period, the regulating unit turns on the multi-spectrum LED array to perform main blue light irradiation according to the first light quality ratio; When red light irradiation is performed during the differentiation period, the regulating unit turns on the multi-spectrum LED array to perform main red light irradiation according to the second light quality ratio.

10. The plant callus cultivation device according to claim 1, characterized in that: When the cooperative triggering condition is met, the analysis unit controls the adjustment unit to activate the piezoelectric ceramic array to apply mechanical stress vibration to the callus tissue in the culture medium; The synergistic triggering condition is that the oxygen content of the culture medium is less than the critical content, the callus tissue is in the proliferation stage, and the culture temperature is in the rising stage.

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