RU2141195C1 - Method and apparatus for automatic in vitro plant multiplication - Google Patents

Abstract

FIELD: agriculture, in particular, vegetative plant multiplication. SUBSTANCE: method involves placing explants into cultural cells-vessels interconnected so as to define flexible ribbon; filling cells with liquid nutritive medium and covering them with film; moving ribbon to horizontal position; cutting explants lengthwise into parts by means of set of knives and transplanting members, when explant root and stalk top grow; transferring explant parts into similar cultural cells of other similar ribbon. Regulation of air humidity in upper cell chambers by changing number of vent openings in cells allows in vitro plant growing to be intensified. EFFECT: increased efficiency, reduced manual work, intensified plant growth and improved plant adaptability upon transplanting. 13 cl, 16 dwg

Description

 The invention relates to the field of agriculture and is intended for vegetative propagation of plants in vitro, for example grapes, wormwood, lemon, apple, cherry, potato, stevia, cloves, etc.

 A known method and device for aseptic propagation of cells or tissues (application EP N 0412621, A 01 H 4/00, 1991) (prototype method). A chain of interconnected individual sterile capsules is gradually unwound from a horizontal package in a container and moved vertically along a group of stations: at the first station, the capsules (bags) are opened using vacuum suction cups acting on the walls of the capsules and filled with nutrient medium; at the second station, explants are planted in capsules; at the third station, capsules containing nutrient medium and explants are brewed from above with a horizontal seam to create sterility in them; at the fourth station, the capsules are transferred from vertical to horizontal position and stacked in this position in a container in the form of packaging. The prototype method provides a series of sequential actions carried out in the device over the culture vessels intended for planting explants in them and subsequent cultivation of plants from these explants, namely, such as that in the device the culture vessels-capsules are interconnected in a chain from the package, fill them with a nutrient medium, then after the explants have landed, the capsules are closed and put into a container. In the prototype method, the device performs actions on the culture vessels-capsules into which explants are planted. However, the processes for obtaining explants and their landing in these capsules are carried out manually. The prototype method does not provide for any automatic actions on the culture capsule vessels in which the plants grew in vitro. Therefore, manually connected from each other the cultural vessels in which the plants grew in vitro are removed from the container, the capsules are opened, the plants are removed, these plants are cut into explants and the obtained explants are planted in the capsule culture vessels.

 The disadvantage of the prototype method is the lack of automated actions on the culture vessels-capsules in which plants grew in vitro. The processes of extracting capsules from a container, opening them, extracting a plant from a capsule, cutting a plant into explants, capturing and planting explants into capsules are not automated. All these actions in the prototype are done manually. The implementation of these actions, performed in the prototype method manually, are the main labor costs when propagating plants in vitro. The prototype method does not fully automate the reproduction of plants at all stages in a closed cycle. Also, when plants are propagated by the prototype method, culture vessels in which plants were grown in vitro are not reused, which does not lead to correspondingly material savings. The use of the prototype method does not allow to obtain full-fledged plants capable of adapting to in vivo conditions, due to the lack of exchange between the gaseous medium inside the capsule culture vessel and the external gaseous medium. Therefore, additional transplantation of plants into culture vessels of a different type is required to obtain plants that are able to adapt to in vivo conditions. The prototype method allows plants to be propagated only at an intermediate stage, while most plants need to be propagated at the last stage, before being transplanted into the soil, in vivo. The prototype method does not allow plants to propagate in a closed cycle. It takes a lot of time and manual labor to reproduce plants in vitro at the last stage, before planting them in soil, in vivo.

 A known method and device for aseptic propagation of cells or tissues (application EP N 0412621, A 01 H 4/00, 1991) (prototype device). In the prototype device for the propagation of plants used culture vessels - individual sterile capsules (bags), which are interconnected in a chain. Capsules are made in the form of bags of organic polymer with soft walls, with one open side. The chain of interconnected individual sterile capsules (bags) is in horizontal packaging, in a container, with the possibility of its (chain) gradual extraction from this container, transfer from horizontal to vertical position and transport in this position in the device. In the device at the first station (the place of action over the capsules) there is a device for the formation of culture vessels from upright and interconnected capsules (bags) with their upper side open, with the possibility of opening these capsules (bags) by expanding their soft walls to the sides using vacuum suction cups. At the first station, there is also a device for filling the capsules (pouches) with a nutrient medium through the holes in the upper part of the pouches (their walls are pushed apart by vacuum suction cups). At the second station, capsules filled with a nutrient medium are in the open state for explants to land on them. At the third station, a device is installed with the possibility of closing the capsules (bags) by brewing holes in their upper parts with a horizontal seam in order to create sterile conditions inside the capsules. At the fourth station there is a chain of interconnected capsules (pouches) filled with nutrient medium, with explants inside, with the possibility of moving the chain of capsules from vertical to horizontal and folding it into a container in the form of a horizontal package. In the prototype device there are devices for performing actions on the culture vessels-capsules into which explants are planted. However, there are no devices that allow automated planting of explants into these capsules (bags), and also devices that automatically receive these explants: extract capsules with plants grown in them from a horizontal package, open capsules, extract plants from them, cut plants into explants, capture and plant explants in other culture vessels filled with nutrient medium. In the prototype device there are no devices that allow you to re-grow plants in culture capsule vessels, and in the same sense, there are no actions provided by the device's devices on capsules with plants grown in them. The culture vessels-capsules in the prototype are sealed hermetically, and they do not have any openings for exchanging the gas medium inside the culture vessels with the external environment, which does not allow to obtain plants capable of effective adaptation. Thus, in the prototype device there are no devices with the ability to automatically perform the main actions when propagating plants in vitro, namely to extract capsules with plants grown in them from a container, open capsules, cut plants into explants, capture these explants and plant them in other culture capsule vessels. When plants are propagated by the prototype device, the main operations of this process are performed manually, and automated actions are performed only on the culture vessels-capsules into which explants are planted. In capsules hermetically sealed with a horizontal seam, there are no openings allowing exchange of the gaseous medium inside the culture vessel-capsule with the external medium without violating sterility. For this reason, when plants are propagated by the prototype device, plants grow that are not capable of effective adaptation after their transplantation into the soil, into the greenhouse. Therefore, before planting plants in vivo, it is required to obtain them in other culture vessels, the actions of which are not provided for in the prototype device. And since when propagating plants in vitro, the cultivation of plants already intended for transplantation in vivo requires the greatest amount of work (it is necessary to propagate more plants than in the previous stages) at the last in vitro propagation stage before adaptation in vivo, the lack of the above devices in the prototype device significantly reduces the efficiency of in vitro propagation.

 The basis of the invention is the task to create a method of automated propagation of plants in vitro, producing actions on culture vessels and on plants grown in them under sterile in vitro conditions that provide complete automation of all processes of plant propagation in vitro, in contrast to the actions in the prototype, automated cutting plants for explants and planting explants in culture vessels, obtaining complete plants capable of adaptation when they are planted in the soil, in vivo, as well as repeated growing plants from the remains of stems with roots in the culture vessels where the plants were cut into explants, which eliminates the cost of manual labor at all stages of the closed plant propagation cycle in vitro, increases the rate of propagation, saves materials (re-growing plants in culture vessels), plant growth in vitro is accelerated, the survival rate of plants when they are planted in the soil, in vivo conditions is increased, and therefore, in general, the economic costs necessary for plant propagation are reduced vitro.

 The problem is solved in that in a method of automated propagation of plants in vitro, containing essential features common with the prototype, including a series of sequential actions by the device on the culture vessels into which the explants are planted, such as that the cultural cells connected together in the form of a ribbon (chain) capsule vessels (pouches) or cells are unwound from the packaging, fill them with nutrient medium, explants are planted in them (in the prototype, manually), and these capsule culture vessels are closed dots) or cells to create sterility in them and put the tape (a chain of capsules or cells) in a package for growing plants in vitro from explants; according to the invention, it is introduced that the device with the plants grown in its cell culture vessels is unwound from a roll onto a tripod, due to the flexibility of moving this tape from vertical to horizontal, open the cell by removing the coating film from the base of the tape, then the stems of the grown plants with sets of knives and transfer legs are cut in the upper measures of cells on explants, which are then planted with the same sets in two or more cells of another, moving in the horizontal plane of a sterile tape, into which a liquid nutrient medium is preliminarily poured, after which these cells are closed by attaching a coating film to the base of the tape, transfer this tape from horizontal to vertical position and wrap it into a roll on a tripod, and into the lower chambers and cameras for planting explants of the tape cells, in the upper chambers of which the stems were cut into explants, to they add fresh liquid nutrient medium to the dispenser, after which these cells are closed by attaching a coating film to the base of this tape, due to its flexibility they are also transferred from horizontal to vertical position, wound onto a roll on a tripod and from the rest of the stems in the chamber for planting the explant with their roots plants are re-grown in the lower chamber. Depending on how the knives and interlocking legs are arranged in their set, they either first cut the stem into explants, and then grab them with interchangeable legs, or first grab the stem of the plant in places of future explants, and then cut this stem into explants with knives. The humidity in the upper chamber and the aeration of the liquid nutrient medium in the lower chamber of the tape cell is controlled by the number and diameter of grooves made on the basis of the tape, as well as by the size of the holes in the lower wall of the chamber for landing the explant. Common features of the prototype method with the proposed method are actions on cultural capsule vessels (bags) or cells into which explants are planted, interconnected in the form of a chain (tape), such as the fact that the capsules are unwound from the packaging and delivered to the device, where the culture vessels are filled with nutrient medium and explants are planted in them. Then, the combined culture vessels-capsules (bags) or cells are closed to create sterile conditions in them and wound in the form of a package.

 The known and proposed methods differ in that 1) in the prototype method, a chain of interconnected capsules (pouches) is unwound from a horizontal package into containers (capsules are laid by a snake, for example, from left to right, from right to left, etc.), and in the proposed method they (cells) are unwound from a roll; 2) in the prototype method, actions are performed in the device over interconnected capsules (bags) in an upright position, and in the inventive method, over those in a horizontal position; 3) in the prototype method, culture vessels are formed from capsules (pouches) by pushing the walls of the capsules (pouches) to the sides using vacuum suction cups, and in the proposed method, the culture vessels-cells have a rather rigid structure, retain their shape and therefore there is no need for the formation of cultural vessels from them, since they are already ready for filling the nutrient medium in them and landing explants; 4) in the prototype method, the explants are planted in the culture vessels-capsules (bags) manually, in the proposed method - automatically using a set of knives and transfer legs; 5) in the prototype method, the culture vessels-capsules (bags) are closed to create sterility in them by brewing the upper open side of the upright bags with a horizontal seam, and in the proposed method, the culture vessels-cells are closed by attaching a tape to the base (located in a horizontal position) thermoactive coating film; 6) in the prototype method, culture vessels-capsules (bags) interconnected in a chain are folded in the form of horizontal packaging into a container (snake, from left to right, from right to left, etc.), and in the proposed method, the culture vessels of the tape wound onto a roll on a tripod (in a roll, the cells are in an upright position); 7) in the prototype method, explants are manually planted in capsules (bags) filled in the device with a nutrient medium. In the proposed method, the landing of explants is carried out in the device by a set of knives and transfer legs in automatic mode. From all of the above features of the method - the prototype and the proposed method, the common features for both methods are selected. The differences of the proposed method from the prototype are as follows. 1) In the proposed method, actions are performed on the culture vessels in which the plants grew in vitro, as well as on the plants grown in them, namely, such that the tape with the plants grown in its culture vessel cells is unwound from a roll on a tripod and due to flexibility, they are transferred from a vertical (in this position plants grew in cells in the light room) to a horizontal position, the cells are opened by removing the coating film from the base of the tape, and the stems of the grown plants are cut with knives of a set of knives and feathers accessory legs to the explants and capture the explants with the interlocking legs of these kits. In the prototype method, the device does not perform any action on the culture vessels-capsules (bags) in which the plants grew in vitro, as well as on the plants grown in them. Obviously, the actions, such as removing the culture vessels-capsules (pouches) from the package, opening the capsules (pouches), extracting plants from them, cutting plants into explants, capturing explants for planting, are carried out manually in the prototype method. 2) In the proposed method, the transfer legs that captured the explants into which the plant was cut with knives transfer these explants to the culture vessels of the cell of another tape. The transplant paws plant explants in these cells with the liquid nutrient medium pre-poured into them by the dispenser. In the prototype method, actions are performed on the culture vessels-capsules into which the explants are planted, but the operation of the explants landing is performed manually. 3) In the proposed method, cells with liquid medium dispenser poured into them and explants planted with transfer legs are closed by attaching a coating film to the base of the horizontal tape. In the prototype method, the culture vessels-capsules (pouches) located in an upright position are closed by brewing their upper, open part with a horizontal seam. 4) In the proposed method, the closed culture vessels-cells with explants planted in them, combined in the form of a flexible tape, are transferred from horizontal to vertical position, and in the prototype method, the chain of interconnected capsules (bags) is, on the contrary, transferred from vertical to horizontal. 5) In the proposed method, closed culture vessels-cells, interconnected in the form of a tape, filled with nutrient medium and planted explants, are wound upright on a tripod in a vertical position, and in the prototype method, culture vessels-capsules (bags), on the contrary, in a horizontal position (with a snake, for example, from left to right, from right to left, etc.), they are folded into a container in the form of a horizontal package. 6) In the proposed method, in the culture vessels, cells in the upper chambers of which the plants were cut into explants, with the remains of the stems in the chambers for planting explants and roots in the lower chambers, add fresh liquid nutrient medium. A certain amount of medium remains after growing in the plant cell. In the horizontal position of the cell, the medium spreads over its chambers (except for the upper one). Therefore, in which of the chambers the nutrient medium will be added: to the lower chamber, to the chamber for explant landing or in two at once, it does not matter, since the remains of stems in the chamber for explant landing have roots in the lower chamber, and after the cells are closed and When they are placed in a vertical position, the entire medium will flow into the lower chamber. Then these cells are closed by attaching the coating film to the base of the tape, transfer this tape from horizontal to vertical position and wind it in this position into a roll on a tripod. In the prototype method, all these actions are absent, since no operations are performed in the device on the culture vessels in which the plants grew in vitro, as well as on these plants. 7) In the proposed method, plants are grown in cell culture vessels from both explants and from parts of stems with roots remaining after cutting the explants. In the prototype method, plants are grown only from explants. 8) In the proposed method, you can either first cut the plant into explants, and then grab them with transplant paws, or first grab the plant with transplant paws in places of future explants, and then cut it with knives. In the prototype method there are no cutting operations in the device of the plant for explants and their capture. These actions are performed in the prototype manually. 9) In the proposed method, the air humidity in the upper chamber and the aeration of the liquid nutrient medium in the lower chamber of the tape cell is controlled by the number and diameter of grooves made on the basis of the tape, as well as the size of the holes in the lower wall of the chamber for landing the explant. In the prototype method, these operations are absent. Culture capsule vessels (pouches) are sealed with a horizontal seam. Thus, the proposed method for a number of essential features of a technical solution to the problem - automation of plant propagation in vitro differs from the prototype method. The proposed method allows you to fully automate the process of plant propagation in vitro, resulting in reduced labor and materials costs, and also provides re-growth of plants in cultured cell vessels in which the plants were cut into explants. Also, by regulating air humidity and aeration of the environment, the ability of plants to adapt in vivo is significantly improved. Obtained in vitro plants are suitable for planting in the soil. Plants propagated by the prototype method are not suitable for transplantation in vivo, where the air humidity is lower than in culture capsule vessels (bags) that are closed (sealed) hermetically.

 The basis of the invention is the task of creating a device for automated propagation of plants in vitro, where plants are propagated in culture vessels with openings made in them, which will ensure both the exchange of the gaseous medium inside the culture vessels with the external environment, and the sterility inside the culture vessels. Also in the culture vessels, supporting and structural elements are necessary to fix the planted explant and maintain the plant in a certain position. The device must contain a device with the possibility of cutting the plants grown in the culture vessels into explants, their capture and planting in other culture vessels with liquid nutrient medium poured into them in the device. The design of the proposed device should automate the cutting of plants into explants, their capture and planting. In the prototype device there is no such device, and these operations are performed manually. The design of the culture vessels should ensure the re-closure and opening of the culture vessels (re-cultivation of plants from parts of stems with roots remaining after cutting the plants into explants). The exchange of the gaseous medium inside the culture vessels with the external environment should contribute to the production of complete plants that take root well after they are planted in vivo. Structural elements made in the culture vessels make it possible to cut plants into explants and capture them with high reliability.

 The problem is solved in that in a device for automated propagation of plants in vitro, containing essential features common with the prototype, such as culture vessels interconnected in the form of a tape (chain), separated from each other by jumpers and moved in the device from one of their packaging in another, with the possibility of filling these culture vessels with a nutrient medium, planting explants in them and closing them for further growing from plant explants in vitro, according to the invention, that flax is introduced This consists of a base in the form of a leaf with cultured cell vessels stamped in it, in the cell there is an upper chamber for the growth of the stele of the plant, a lower chamber for the development of its roots and a chamber for planting the explant located between the upper and lower chambers, which is separated from the lower chamber of the lower a wall with a groove for accommodating the lower part of the explant, bounded on both sides by protrusions with the possibility of attaching a coating film to them, with holes in the lower wall for overflowing of the nutrient medium into the lower After transferring the tape from horizontal to vertical, the lower chamber and the chamber for landing the explant are wider than the upper chamber, grooves are stamped on the base of the tape connecting the upper chamber and the camera for landing the explant with a hole in the ribbon bridge, and perforations are made with the possibility of engagement them with a toothed drum, the device includes a set of knives with interchangeable paws, and grooves are stamped in the cells of the tape with the possibility of movement of knives in them while cutting plants into explants and capture of these explants with transfer legs on both sides of each groove, the zigzag grooves stamped on the base of the tape connecting the chamber for planting the explant in the cell and each section of the upper chamber bounded by the walls of the grooves for cutting plants with a hole in the tape made between its cells, the film the coating is made of thermoactive material with the possibility of heating it joining the base (except for the sides of the tape, where the perforation holes are made) of the tape, into the cells of which are poured a rich nutrient medium and explants were planted, as well as ribbons, in the cells of which there are parts of stems with roots remaining after cutting the stems into explants in the upper cells of the cells for subsequent cultivation in vitro of plants in the light room, both from explants and from parts of the stems with roots, sets of knives and interchangeable legs can be installed in the device, in which the knives and interchangeable legs are rigidly connected to each other with the possibility of movement in the grooves of the cells towards each other, while the knives are first cut into the stalk l plants for explants, which are then captured by transplant legs, or the device contains sets of knives and transplant legs, in which the knives and transplant legs are not rigidly connected to each other, and one of the walls of each groove of the upper chamber of the cell the ability to first capture the stem of the plant in places of future explants, and then cut it into explants by sliding each knife from top to bottom along the inclined wall of the groove of the cell. The proposed device differs from the prototype device by the presence of a set of knives and transfer legs with the possibility of automated cutting of plants into explants, their capture and planting of these explants in the culture cell vessels filled with liquid nutrient medium. The prototype device does not have any device for cutting plants into explants, their capture and planting, which requires labor to perform these operations manually. In more detail, the differences between the proposed device and the prototype are as follows. 1) In the prototype device, the culture vessels-capsules are made of organic polymer with soft walls in the form of bags, interconnected by sides and having open upper sides. In the proposed device, the culture vessels-cells that retain their shape are stamped on a flat sheet of the base of the tape, the flexibility of which is ensured by the flexibility of the jumpers between the cells and the presence of holes between the cells in the jumpers. The cells are open on the side (capsules (bags) in the prototype are open on the upper side). 2) In the prototype device, the culture vessels-capsules (bags) with soft side walls with the possibility, after filling them with a nutrient medium and landing in them explants, to close the culture vessels-capsules (bags) by hermetically sealing their open upper side with a horizontal seam. In the proposed device, the culture vessels of the cells are made with the possibility of their closure by attaching a thermoactive coating film to the base of the tape (excluding the sides of the tape, where perforation holes are made, which serve to engage gear drums for them). Of all these characteristics of the prototype device and the claimed device, the common features for both of these solutions are selected. The design of the claimed device differs from the design of the prototype in the following ways. 1) In the proposed device in the culture vessels there are structural elements with the ability to maintain in a predetermined position a planted explant and a growing plant (lower and upper chambers, a chamber for planting an explant, with walls delimiting them, and the width of the upper chamber is less than the width of the lower chamber and chamber for planting explant), as well as ensuring reliable cutting of the plant with knives into explants and capture of these explants by the transfer legs of the set of knives and transfer legs installed in the device (pa s in the upper chamber of the cell). The culture vessels-capsules (bags) of the prototype device are made in the form of bags with soft walls and there are no structural elements inside them. 2) In the proposed device, on the basis of the tape, zigzag grooves are made with the possibility of exchanging the gas medium inside the culture vessels with the external medium after attaching the coating film to the base of the tape, which improves the quality of the grown plants and accelerates their growth. In the prototype device, the culture vessels-capsules (bags) after filling them with nutrient medium and landing explants in them are closed hermetically, which does not allow to obtain full-fledged plants capable of adapting to in vivo conditions. 3) In the proposed device, sets of knives and transfer legs are installed with the ability to automatically cut the explants grown in the culture vessel cells to capture these explants (moreover, the explants are cut and captured directly in the upper chamber of the cell, simultaneously along the entire length of the plant stem), transfer explants to the cells, with the nutrient medium previously poured into them in the device and disembarking them in these cells, which eliminates the cost of manual labor for performing these operations. In the design of the prototype there is no device for cutting plants into explants and planting explants, and these operations are done manually. Thus, the proposed device in comparison with the prototype device will reduce labor costs, increase the efficiency of in vitro propagation and improve the quality of plants grown in vitro.

 The invention is illustrated by drawings.

 In FIG. 1 schematically shows a ribbon with cells for automated propagation of plants in vitro and planting them for adaptation to in vivo conditions; in FIG. 2 - a tape cell ("a" is a top view, "b" is a section along A-A to "a", "c" is a section along BB in "a", "d" is a section along BB in “a”, “d” - section along the G-D to “a”, “e” - the section along the D-D to “a”, “g” - the section along the E-E to “a”). In FIG. 3 schematically shows an automatic machine for propagating plants in vitro; in FIG. 4 - a tape cell, knives with transfer legs and transfer legs without knives in the upper position before cutting the plant into explants and capturing the explants; in FIG. 5 - a tape cell, knives with interchangeable legs and interchangeable legs without knives in the lower position before cutting the plant into explants and capturing the explants; in FIG. 6 - transfer legs that captured the explant, two frames are in the upper position before the explant is displaced; in FIG. 7 - interchangeable legs in the open position, two frames are located on the base in the lower position after the landing of the explant; in FIG. 8 - drive movement of a set of knives with interchangeable legs; in FIG. 9 - the movement device in the machine sets of knives with interchangeable legs; in FIG. 10 - design of the knife in the form of a plate with interchangeable legs, not rigidly connected to the knife.

 For propagation of plants by the proposed method and device for automated propagation of plants in vitro using culture vessels-cells of the tape (Fig. 1, 2). The tape consists of a base 1 in the form of a sheet with cells 2 stamped in it for placement of a nutrient medium and an explant, separated from each other by jumpers 3. A coating 4 of one sheet of a transparent thermoactive film is attached to the tape. In the cell there is a lower chamber 5 for root growth, placement of a liquid medium and an upper chamber 6 for the growth of the plant stem. At the base of the tape are perforation holes 7 for transporting the tape.

 In the cell of the tape 2 there is a chamber 8 for displacing the explant, which is separated from the lower chamber 5 by the lower wall 9. In the lower wall, holes 10 are stamped for the medium to flow from the chamber 8 to place the explant in the lower chamber 5. Aeration of the liquid medium is also carried out through the holes 10 the lower chamber 5. On the lower wall 9 are the protrusions 11, which limit on both sides the lower groove 12 to accommodate the lower part of the explant. A cover 4 is attached to the protrusions 11 and the protrusions 13 located at the edges of the lower wall 13, which are at the same level with the base 1, which prevents the explant from falling into the lower chamber and prevents the coating from sagging under the weight of the medium located in the lower chamber 5, after transferring the tape from horizontal in vertical position. The upper chamber 6 is separated from the chamber 8 for placement of the explant by the upper wall 14, which prevents the medium from entering the upper chamber 6, but does not inhibit the growth of the stem at the explant. On the upper wall 14 there are upper protrusions 15 defining the upper groove 16 on both sides to accommodate the upper part of the explant. To the protrusions 15, located at the same level with the base 1, a cover 4 is attached, which prevents the explant from falling out of the groove 16 after the tape is put into vertical position. If the explant slides down and its upper part falls out of the groove 16, it is held by the leaves in the lower groove 12 from falling into the lower chamber 5. In the upper chamber 6 of the ribbon cell 2, grooves 17 are stamped in which the knives move during cutting the plant into explants. Each groove 17 is bounded below by a wall 18 and from above by a wall 19. The lower chamber 5 and chamber 8 for placing the explant are wider than the upper chamber 6, as a result of which the width of the explant is less than the width of the chamber 8 for placing the explant, which excludes its parts (leaves) from reaching the base tapes while attaching the coating. Grooves 20 connecting the upper chamber 6 and grooves 21 connecting the chamber 8 to accommodate the explant with the external environment are stamped on the basis of the tape. Through the grooves 20 and 21, the gas is exchanged between the cell and the external environment, which ensures the physiological processes of photosynthesis and respiration in plants. The grooves 20 and 21 go from the chambers 6 and 8 to the hole 22 between the cells of the tape. Hole 22 promotes the flexibility of the tape when moving the tape from vertical to horizontal, and vice versa, provides air exchange of cell 2 with the external environment, and also reduces the size of the roll of tape in the case of the distance between the cells of the tape corresponding to the distance between the plants in a row when they are planted to adapt to in vivo conditions - the cells of each subsequent turn of the tape in the roll go into the holes between the cells. In addition, material is saved - the plastic of which the tape is made. To eliminate clogging of the plantation with tape residues, the tape and the tape coating are made of a material that decomposes over time under the action of microorganisms or under the influence of ultraviolet radiation during pretreatment before planting for adaptation, or under the influence of moisture on the outside of the tape and coating, or by combining various materials that can decompose under the action of various factors in the layered production of tape.

 For plant propagation by the proposed method, the device (Fig. 3) is placed in two articulated laminar boxes 23 (dustproof chambers of the UO-BG type or in similar laminar boxes) with an adjustable air flow rate, which ensures sterility of the process. The device contains a roll of sterile tape 24, roll 25 of a sterile thermoactive film of coating 4, which is attached to the base 1 of the tape, a roll of tape 26 with grown plants intended for cuttings, from which the thermoactive film coating is removed and twisted into a roll 27. The coating 4 of the thermally active film is disconnected from the base 1 of the tape 26 by heating with a figured roller 28. The sterile nutrient medium is poured into the cells of the sterile tape 24 by the dispenser 29. The nutrient medium is in the tank 30. Cutting of plants to explants and planting of explants in the cells 2 of the sterile tape 24 carry out sets of 31 knives with interchangeable legs. Knives and transfer legs are disinfected in microwave ovens 32. The movement of the tape 26 with grown plants intended for cuttings, and the tape with planted explants is carried out by engaging the perforation holes 7 with a toothed drum 33. To the base 1 of the tape, the medium poured into the cells 2 and the planted explants they attach a coating 4 of a thermoactive film by heating it with a figured roller 34. The tape is transferred from horizontal to vertical position and wound into a roll 35.

 Further propagation of plants by the claimed method is as follows (Fig. 3): the medium is dispensed with dispenser 29 simultaneously in three cells, the plants are simultaneously cut into explants in three cells 2 with three sets of 31 knives with transfer legs, the explants are simultaneously planted in three cells with three sets of 31 knives with transfer legs and at the same time sterilize three sets of knives with transfer legs in three microwave ovens 32. The tape, in which the stems are cut into explants in the upper chamber 6, is reused for cutting pressing in its cells 2 plants from the remains of the stems in chamber 8 with roots in the lower chamber 5. For this, fresh medium is dispensed into the cells 2 of this tape with the dispenser 36 into the lower chamber 5 and into the chamber 8 for planting the explant, a coating 4 is placed on the roll 37, by heating it with a figure roller 38 and after transferring the tape from horizontal to vertical, it is wound into a roll 39. Re-growing plants from the remains of stems with roots after cutting the plant into explants in the tape cells not only saves material and costs, widely regarded as one for the manufacture of tape, but also accelerates plant growth, thanks to the roots of the stem residues.

 In the proposed device for propagation of plants in vitro, the propagation of cuttings of plants on explants is carried out in the cells of the tape using a set of knives and transfer legs. Knives and transfer legs are rigidly connected in pairs. The knives protrude forward and to the side of the transfer legs by a few millimeters. Knives with transfer legs and transfer legs without knives are arranged in pairs, alternating from the bottom of the stem to the top of the stem as follows: knives with transfer legs, transfer legs with knives, transfer legs without knives, etc. depending on the number of explants into which the plant is cut.

 In the proposed device, in a set of knives with interchangeable legs (Fig. 4), knives 40 with interchangeable legs 41 and interchangeable legs 42 without knives are closed when cutting plants into explants and capturing explants, namely they move towards each other in a plane perpendicular to the base 1 tape. The explants are sliced and captured in the upper chamber 6. The knives move during the cutting of the plant into explants in special grooves 17. On both sides of the walls 18, 19 of the grooves 17, the transfer legs 41, 42 capture the explants. Due to the presence of the grooves 17, the cell 2 can be of any shape. From the remaining part of the stem in chamber 8 for planting an explant with roots in the lower chamber 5, plants are re-grown in the same tape, rewound from roll 26 to roll 39. To cut the plant into explants and capture the explants, knives 40 with transfer legs 41 and transfer legs 42 without knives translate from the upper position (Fig. 4) to the lower position (Fig. 5). In this case, the knives go into the grooves 17, and the interchange legs 41, 42 are located on both sides of the walls 18, 19 of the grooves 17.

 The process of disembarking the explant into the chamber 8 for disembarking the explant and the design of the set 31 (Fig. 3) of knives with interchangeable legs are presented in FIG. 6, 7. After cutting the plant into explants with knives 40 and capturing the explants with transfer legs 41, 42, the set of knives with transfer legs is shifted to a sterile tape unwound from roll 24 with the medium poured into its cells. The transfer legs 42 (Fig. 6) carrying the explant are located above the chamber 8 for disembarking the explant. The ends 43 of the transfer legs 42 are movably connected to the frame 44 and can move along the guide rails 45, 46, 47, 48. Two frames on which the transfer legs 42 carrying the explant are located (one foot captures the lower part of the explant, the other upper) are connected between themselves rails 49, 50, 51, 52. Two pairs of ends 43 of the transfer legs 42 are interconnected by ribs 53, 54, 55, 56. A tightening spring 57 is attached to the ribs 55, 56, which closes the transfer legs 42 to capture the explant after cutting the plant to explants. Under the action of the tightening spring 57, the interlocking legs 42 hold the explant until it disembarks into the chamber 8 for disembarking the explant. In the middle of the ribs 55, 56, at the attachment points of the tightening spring 57, two rails 58, 59 are movably attached, which are movably connected by their upper ends to the platform 60. The platform 60 can move down the rod 61, fixedly connected to the jumper 62, which is fixedly connected to the guides strips 46, 48 of two frames. Two interconnected frames are movably connected to stiffeners 63, 64, 65, 66 of the base 67 of the set of knives with interchangeable legs and can move up and down. To the ribs 68, 69 of the base 67 of the set of knives with interchangeable legs and to the midpoints of the rails 50, 52 connecting the two frames, there are fastening springs 70, 71, which stretch when two frames move down the stiffening ribs 63, 64, 65, 66 of the base 67 sets of knives with interchangeable legs. Depending on the depth of the cell, the movement of the two frames 44 down is stopped by the stops 72, 73, 74, 75. The movement to the sides of the ends 43 of the transfer legs 42 along the guide rails 45, 46, 47, 48 during the opening of the transfer legs 42 when stopping the explants, the stops 76 The reverse movement of the ends 43 of the transfer legs 42 is stopped by the ratchets 77. A spring 57 pulling together two pairs of ends 43 of the transfer legs 42 is more rigid in tension than two pulling springs 70, 71 supporting two frames in the upper position. During the cutting of the plant into explants (Fig. 6), the clamping bar 78, fixedly connected to the guide bar 45 of the frame 44, presses the plant stem against the walls 18, 19 of the grooves 17, which ensures the capture of the explants in a certain place of the transfer legs and the exact landing of the explants in the grooves 12, 16 cameras 8 for the landing of explants.

 Reproduction of plants in the proposed device using a set of knives with interchangeable legs is as follows. Two frames (Fig. 6) 44, interchange legs 42 which captured the explant, are in the set of knives with interchange legs. If there is an explant in the transfer legs 42, the photocell gives a signal for the movement of the tape 24 with the medium poured into the cells 2. After moving the tape, a signal is sent to move down the core of the solenoid. The pipe, which is a continuation of the core of the solenoid, has a cone-shaped hole at the lower end. The upper end of the rod 61 has a pointed shape. When the solenoid core moves downward, the pointed end of the rod 61 enters the conical hole of the pipe. Two interconnected frames 44 together with the base 67 of the set of knives with interchangeable legs are displaced, which ensures the exact location of the interchangeable legs over the chamber 8 for landing the explant. With further movement of the solenoid core downward, the pipe presses on the platform 60. Since the springs 70, 71 are easier to stretch than the spring 57 (Fig. 7), two frames move down the stiffening ribs 63, 64, 65, 66 of the base 67 to the stop of the rails 50 52 into the stops 72, 73, 74, 75, after which, under the pressure of the solenoid core, the platform 60 moves down the shaft 61. The rails 58, 59, movably connected to the platform 60, push the transfer legs 42 apart to the end of the ends 43 of the transfer legs 42 into the stops 76, while stretching the spring 57. The lower part of the stem of the explant tries ditsya in the groove 12, the top - in the groove 16. The explants are planted into the chamber for landing explant. The shaft of the solenoid moves up. The reverse movement of the ends of the 43 transfer legs along the guide bars 45, 46, 47, 48 under the action of the tightening spring 57 is restrained by the ratchets 77. Under the action of the tightening springs 70, 71, the two frames 44 are raised to the upper position. The rod 61 is released from the pipe, which is a continuation of the core of the solenoid. The set of knives with interchangeable legs moves, the next pair of interchangeable legs carrying the explant is located above the chamber 8 for landing the explant. In the absence of an explant in the transfer legs, the photocell does not send a signal to shift the tape 24, as a result of which no signal is generated for the movement of the solenoid shaft. The next set of knives with interchangeable legs is shifted to the chamber 8 for the landing of the explant.

 After planting the explants, the set of knives with transfer legs is shifted to the microwave oven 32, where the knives 40 with transfer legs 41 and the transfer legs 42 without knives are sterilized. All frames 44 of the set of knives with interchangeable feet are moved to the lower position, as a result of which the knives and interchangeable legs enter the slots in the lid of the microwave oven 32 by pressing on all platforms 60 of the set. If the explant was not displaced from the transfer paws 41, 42 (there was no explant in the transfer paws and they remained in the closed position, since the photocell did not send a signal to shift the tape and, therefore, a signal was not generated to move the core of the solenoid), the solenoid located above the microwave oven 32, not only puts all the frames 44 of the kit in the lower position, but also opens the transfer legs 41, 42 by pressing on the platform 60.

 After sterilization, a set of knives with interchangeable legs is shifted to tape 26 with grown plants to cut the plant into explants and capture the explants. The accuracy of the location of the knives 40 with interchange legs 41 and interchange legs 42 without knives above the grooves 17 of the upper chamber 6 is ensured in the same way as when landing the explant. Knives 40 with transfer legs 41 and transfer legs 42 located on the frames 44 of the set are moved from the upper position (Fig. 4) to the lower position (Fig. 5) by pressing on the spring plates of the ratchets 77 under the influence of the solenoid located above the tape 26 sec grown plants. Since it takes more force to bend the spring plates of ratchets 77 than to stretch the springs 70, 71, first the set frames 44 are shifted to the lower position, and then the spring plates of the ratchets 77 are bent, the obstacle to the movement of the end 43 of the transfer legs 42 towards each other is removed under the action of a tightening spring 57. Knives 40 cut the plant into explants, transfer legs 41, 42 capture the explants. The core of the solenoid cut the plant on the explant moves up. All frames 44 of the set are raised to the upper position under the influence of tightening springs 70, 71. The set of knives with interchangeable legs 41, 42, which captured the explants, is shifted to the tape 24 with the medium for planting explants poured into the cells. The whole process of reproduction begins again in a closed cycle.

 Sets of knives with interchangeable legs are shifted in the device after performing one operation to perform another as follows (Fig. 8). Axes 81, 82 are attached to the sides 79, 80 of the base 67 of the set of knives with interchangeable legs, which move along the perimeter of the groove 83 formed by the stop strips 84, 85. On the axis 82, which protrudes above the stop strips 84, 85, there is a sleeve 86, which is rigidly connected to the sleeve 87. In turn, the sleeve 86 rotates on the axis 82. The axis 88 is movably connected to the sleeve 87 and fixedly connected to the chain link 89. The chain is on the gear 90. The gear 90 is rigidly connected to the gear 91, on which the chain 92. Thus, chain 92 leads in motion ue gear 91 which in turn drives the gear 90 and located thereon circuit 89. After rigidly connected to a chain link 89 axle 88, bushing 87, 86, the axis 82 is driven by a base 67 with a set of knives interchange paws.

 In FIG. 9 shows the process of performing the same operation with three sets of knives with transfer legs. To each of the chains 93, 94, 95 are attached 6 sets of knives with interchangeable legs. A set of 96 knives with interchangeable legs cuts the plant into explants in the upper chamber 6 in cell 2 of the tape 26 with the grown plants. Tape 26 is shifted immediately to 3 cells. A set of 96 knives with interchangeable legs cuts the plant in every third cell 2, leaving the plants in two cells for cutting with sets 97, 98. A set of 99 knives with interchangeable legs transplant explants into every third cell 2 of tape 24, leaving two cells with medium for the explant landings with sets 100, 101. For the knives 93, 94, 95, sets 102, 103, 104 connected to the chains, the knives with the transfer legs of the frame 44 are in the upper position on the base 67 and those transfer legs 41, 42 that carried out the explants are unclenched. Sets of knives 102, 103, 104 with transfer legs connected to chains 93, 94, 95 are sterilized in three microwave ovens 105, 106, 107. Above each of sets 96, 97, 98, cutting plants into explants, there is a solenoid, which moves all the frames of each set on the base to the lower position, and then pressing all the ratchets 77 causes the knives 40 to interlock with the transfer legs 41 and the interlock legs 42 without knives to close, as a result of which the plants are cut into explants and the explants are captured by the transfer legs 41, 42 in cells 2 tapes 26. Above each One of the sets of explants 99, 100, 101, which makes the landing of explants, is the explant landing solenoid, which puts the two interconnected frames 44 into the lower position and then opens the transfer legs 41, 42 by pressing on the platform 60. Above each of the sets 102, 103 104, in which the knives 40 with transfer legs 41, 42 are sterilized in the microwave ovens 105, 106, 107, there is a solenoid that, by pressing all the platforms 60 of the sets 102, 103, 104, puts all the frames 44 on the base 67 to the lower position, and then spreads the knives 40 with interchangeable legs 41 and transplant legs 42 without knives, since with a small size of the plant, not all transplant legs 41, 42 capture and plant the explant, and therefore some of them are in closed position. Kits 108, 109, 110 with sterile knives and interchangeable legs connected to chain 93 are ready for cutting plants for explants and for capturing explants. Each of the chains 93, 94, 95 with attached sets of knives with transfer legs has an independent drive from electric motors with an adjustable number of revolutions through gearboxes that rotate gears 111, 112, 113, connected by a chain gear to gears 91, which in turn, they are rigidly connected to the gears 90. The need for independent movement of each of the chains 93, 94, 95 is dictated by the fact that plants grow in the cells of the tape different in length and, for example, while one set 99 connected to a chain 93, there will be explants in all transplant legs 41, 42 (the plant has grown over the entire length of the cell), in another set 100 connected to another chain 94, only half of the transplant legs 41, 42 will carry explants, since the plant grew only half the length cell top chamber. Therefore, in this case, the chain 94 is shifted by a larger number of links compared to the chain 93, which provides a high speed of landing of explants and eliminates the absence of explants in the cells of the tape.

 For cutting plants into explants and capturing explants, another set of knives and transfer legs can be used (Fig. 10): knife 114 in the form of a plate, not rigidly connected to the transfer legs 115, 116, 117, 118. The knife 114 is located on a movable rail 119, which can move up and down in the knife holder 120. The knife 114 draws the spring 121 down. The movable rail 119 is mounted on the holder 120 with a platform 122 and a crescent-shaped washer 123. The knife holder 120 is mounted on the movable rail 124. On the movable rail 124, limiters 125, 126 for horizontal movement of the holder 120 and compression spring 127 are installed. The interlocking tabs 115, 116, capturing the explant from above and below on one side, and 117, 118, capturing the explant on the other hand, are interconnected below the fastening on the sleeve 128. The sleeve 128 is mounted on a movable rail 124.

 The knife 114 cuts the plant into explants by moving the movable rail 124, on which it is mounted, from top to bottom and sliding the knife blade 114 along the wall 129 of the groove 130 of the upper chamber 6 of the cell 2. The stalk falls between the sharp edge of the wall 129 of the groove 130 of the upper chamber 6 of the cell 2 and the lower edge of the blade of the knife 114. The angle α of the sharpening of the knife 114 is equal to or may be slightly larger than the angle β of the inclination of the wall 129 of the groove 130, which allows the blade of the knife 114 to slide on the wall 129 of the groove 130 of the cell 2 with the smallest gap. The knife 114 can abut against the bottom 130 of the groove of the cell without damaging it due to the spring 121. The spring 127 presses the knife against the wall 129 of the groove 130 of the cell 2 while cutting the plant stem into explants. When the knife 114 moves downward along the wall 129 of the groove 130 of the cell 2, the knife 114 moves to the right, compressing the spring 127. By fixing the stops 125, 126 on the movable rail 124, the location of the knife 114 is adjusted relative to the grooves 130 of the cell 2 of the tape. The transfer legs 115, 116 and 117, 118 are closed when the knife 114 presses the plant against the wall 129 of the groove 130 of the cell 2 before cutting the stem into the explants, as a result of which the explant is captured in a strictly defined location of the transfer legs, which allows it to be precisely planted in the grooves 12, 16 cameras 8 for the landing of the explant. The locking mechanism of the interlocking legs 115, 116 and 117, 118 can be made, for example, in the form of hinge levers that closes the interlocking legs during the engagement of the explant under the action of a tightening spring and opens under the action of the solenoid with their locking in the open position by the ratchet mechanism.

Bibliographic Information
1. USSR author's certificate N 1341114, B 65 D 85/52, 09.30.87.

 2. Application WO 88/04520, A 01 G 7/00, 06/30/88.

 3. Application WO 91/15110, A 01 H 4/00, 10.17.91.

 4. Application WO 91/03929, A 01 H 4/00, 04/04/91.

 5. Application EP 0412621, A 01 H 4/00, 02/13/91.

Claims (13)

1. A method of automated propagation of plants in vitro, including unwinding from a package of culture vessels-cells joined together in the form of a tape, filling them with a nutrient medium, planting explants in them, closing these culture vessels-cells to create sterility in them and laying the tape (chains culture vessels-cells) in a package for growing plants from explants, characterized in that the tape with the plants grown in its cells is unwound from a roll on a tripod, then due to its flexibility they are transferred from a vertical horizontal, open the cell by removing the coating film from the base of the tape, cut the stems of the grown plants in the upper chambers of the cells into explants, which are then planted in the cells of another sterile tape moving in the horizontal plane, where liquid nutrient medium is preliminarily poured, after which these cells close by attaching the coating film to the base of the tape, transfer the tape from horizontal to vertical and wind it onto a roll on a tripod, and into the cells of the tape in the upper chambers Then the stems were cut into explants, they were added with fresh liquid nutrient medium, then they were closed by attaching a coating film to the base of this tape, then, due to their flexibility, they were transferred from horizontal to vertical position, wound onto a roll on a tripod, where from the rest of the stems in the planting chamber explant with their roots in
lower chamber re-grown plants.  2. The method according to claim 1, characterized in that first, in the cell of the tape, the plant is cut with knives into explants, which are then captured by interchangeable paws.  3. The method according to claim 1, characterized in that first, in the cell of the tape, the grafting paws capture the plant along the entire length of its stem in places of future explants, and then cut the stem with knives into explants.  4. The method according to claim 1, characterized in that the humidity in the upper chamber and the aeration of the liquid nutrient medium in the lower chamber of the tape cell is controlled by the number and diameter of grooves made on the basis of the tape, as well as by the size of the holes in the bottom wall of the chamber for landing the explant.  5. A device for automated propagation of plants in vitro, containing culture vessels-cells, interconnected in the form of a tape, separated from each other by jumpers and moved in the device from one package to another with the possibility of filling these culture vessels-cells with a nutrient medium, planting in them explants and their closure for further cultivation from plant explants in vitro, characterized in that the tape consists of a base in the form of a sheet with stamped cell culture vessels in it, in a cell There is an upper chamber for the growth of the plant stem, a lower chamber for the development of its roots, and an explant landing chamber located between the upper and lower chambers, which is separated from the lower chamber by a lower wall with a groove for accommodating the lower part of the explant, which is bounded on both sides by projections to them the coating film, and from the upper chamber - the upper wall with a groove for placing the upper part of the explant, limited on both sides by protrusions with the possibility of attaching to them the coating film, openings are made in the bottom wall for the nutrient medium to flow into the lower chamber after transferring the tape from horizontal to vertical, the lower chamber and the explant landing chamber are wider than the upper chamber, grooves are stamped on the tape base connecting the upper chamber and the explant landing chamber with a hole in the jumper tapes, and perforation holes are made with the possibility of engaging them with a gear drum, in the closed state of the cells, the coating film is attached to the base of the tape, the device including set of knives with interchange legs, and in the cells of the tape stamped grooves for movement in their knives during cutting plants, the explants and capture these explants interchange tabs on both sides of each groove.  6. The device according to claim 5, characterized in that zigzag grooves are stamped on the base of the tape connecting the chamber for planting the explant in the cell and each section of the upper chamber bounded by the walls of the grooves for cutting plants with an opening in the tape made between its cells.  7. The device according to claim 5, characterized in that the coating film is made of thermoactive material with the possibility of attaching it to the base of the tape by heating.  8. The device according to claim 5, characterized in that the tape with cells and the coating film are made of materials that decompose under the influence of external factors to ensure the further development of plants in vivo.  9. The device according to claim 5, characterized in that the set of knives with interchangeable legs consists of knives rigidly connected to interchangeable legs, and of interchangeable legs without knives, with each pair of interchangeable legs with knives or interchangeable legs without knives placed on the frame with the possibility of movement on the sides of the frame one towards the other, and the frame is movably fixed on the basis of a set of knives with interchangeable legs with the ability to move on the sides of the base: down - to carry out the processes of cutting the plant into explants and their grip, then landing explants and upwards - plant after cutting and gripping explants and explants after disembarkation.  10. The device according to claim 9, characterized in that the two frames, the interlocking legs of which capture one explant, are interconnected to move down the stiffening ribs of the base under the action of the core of the solenoid and upward under the action of tightening springs, and two pairs of knives with interchange legs or interchangeable legs without knives mounted on two frames connected to each other, interconnected with the possibility of movement along the guide rails of the frames one towards the other under the action of a tightening spring and in the opposite direction it under the action of the core of the solenoid, ratchets are installed on the frames with the possibility of restraining the reverse movement of knives with transfer legs and transfer legs without knives after they are apart from each other under the action of the core of the solenoid.  11. The device according to PP.9 and 10, characterized in that the frames are rigidly attached to the clamping strips with the possibility of pushing the stem of the plant to the walls of the grooves in the upper chamber of the ribbon cell in which the plant is cut.  12. The device according to claim 5, characterized in that in the set of knives and interchangeable legs, the interchangeable legs do not have knives, and the knives are not rigidly connected to the interchangeable legs, while the knives are made in the form of plates and installed with the possibility of cutting the plant stem into explants by movement knives along the walls of the grooves of the upper chamber of the tape cell, and the interlocking legs grasping the explant on one side in its lower and upper parts are connected together below their fastening on a sleeve placed on a movable rail, with the possibility of contact I have transplant paws that do not go one after the other if there is no explant in the places intended to capture the explant.  13. The device according to claim 5 or 12, characterized in that the knife is sharpened at an angle equal to or slightly greater than the angle of inclination of the wall of the groove of the tape cell, with the possibility of sliding the lower edge of the knife with the smallest clearance on the wall of this groove during cutting of the plant stem into explants .

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