Device and method for rapid separation of single spore strains of rice blast fungus

Abstract

The invention provides a device for rapidly separating monospore strains of rice blast fungus, comprising an operating table, a driving box, a picking needle and a strain carrying table; a light hole is arranged on the operating table, a driving system is arranged in the driving box, the strain carrying table is arranged along the circumference of the light hole and is used for carrying rice stalks to be operated, and the picking needle moves in horizontal and vertical directions under the drive of the driving system and the needle tip contacts the rice stalks.

Description

Technical Field

[0001] The invention relates to a microbiology technology and a plant protection technology, in particular to a device and a method for rapidly separating a monospore strain of rice blast fungus. Background Art

[0002] Rice blast is a worldwide disease caused by ascomycete fungi, which can cause a significant reduction in rice yields, with a 40% to 50% reduction in severe cases, or even a total loss of grains. Rice blast can occur throughout the entire growth period of rice, with the characteristics of long-term damage, diverse symptoms, and multiple infection sites. Due to the genetic complexity of rice blast fungus races and the rapid variation of avirulent genes, the resistance of resistant varieties will weaken or be lost after 3 to 5 years of promotion and planting. Real-time and accurate understanding and mastering of the composition of rice blast fungus populations and the dynamic trends of changes in avirulent genes of physiological races in each rice-growing area can provide a basis for the rational layout of disease-resistant rice varieties, and also help to screen and cultivate disease-resistant germplasm resources and explore local rice blast resistance resources. Clarifying the distribution of avirulent genes of local rice blast fungi provides a reliable reference for rice breeding.

[0003] Therefore, researchers need to isolate a large number of single-spore strains of rice blast fungus from the field in order to detect the pathogenicity, pathotype and fungicide resistance of different rice blast fungus strains. At present, the existing methods for isolating single-spore strains of rice blast fungus have cumbersome separation processes, high separation costs, long separation and purification cycles, easy contamination during the separation process, and the isolated strains are unlikely to be single-spore strains. These factors have greatly restricted the progress and level of researchers' research on the spatiotemporal dynamics of avirulent genotypes in rice blast fungus populations. Therefore, there is an urgent need for a method that can quickly isolate single-spore strains of rice blast fungus from rice blast-infected straw in the field. Summary of the invention

[0004] The purpose of the present invention is to provide a device and method for rapidly separating monospore strains of rice blast fungus.

[0005] A device for quickly separating single spore strains of rice blast fungus to achieve the purpose of the present invention comprises an operating table, a drive box, a picking needle, and a strain carrying table; a light hole is arranged on the operating table, a drive system is arranged in the drive box, the strain carrying table is arranged along the circumference of the light hole and is used to carry rice stalks to be operated, and the picking needle moves in the horizontal and vertical directions under the drive of the drive system and the needle tip contacts the rice stalks.

[0006] Furthermore, the strain carrier includes a strain carrier ring wall which is cylindrical and arranged along the circumference of the light hole, and a strain front end groove and a strain rear end groove are arranged at both ends along the radial direction on the upper bottom surface of the strain carrier ring wall. A needle picking groove is arranged on one side of the upper bottom surface of the strain carrier ring wall, and the depth of the needle picking groove is greater than the strain front end groove and the strain rear end groove.

[0007] Furthermore, the front end groove spacer is divided into a plurality of small grooves.

[0008] Furthermore, the driving system includes an active driving shaft, a longitudinal driving wheel, a transverse driving wheel, a longitudinal driven wheel combination, a belt, and a transverse rack; the longitudinal driving wheel and the transverse driving wheel are respectively arranged at both ends of the active driving shaft, the active driving shaft can move horizontally, the active driving shaft is not connected to the transverse driving wheel when connected to the longitudinal driving wheel, and the active driving shaft is not connected to the longitudinal driving wheel when connected to the transverse driving wheel, the longitudinal driven wheel combination cooperates with the longitudinal driving wheel to drive the belt to rotate, a clip is arranged on the belt and the clip is exposed from the driving box, and the transverse rack is fixed to the inner wall surface of the driving box and meshes with the transverse driving wheel.

[0009] Furthermore, the clip clamp includes an active clip, a passive clip, and a clip base; the clip base is fixed to a clamp support platform set on the belt, the active clip is connected to the clip base through a rotating shaft and a torsion spring, and the passive clip is connected to the clip base through a rotating shaft; the active clip drives the passive clip to separate or contact the bottom surface of the clamp support platform.

[0010] Furthermore, the active clip includes a clip button, an L-shaped clip connecting rod, and an active clip seat. One end of the L-shaped clip connecting rod fixes the clip button and the other end fixes the active clip seat. The side walls of the active clip seat on both sides of the L-shaped clip connecting rod are the first rotation pair; the passive clip includes a passive clip seat and a clip extension arm. The clip extension arm is arranged at the end of the passive clip seat and a second rotation pair with a groove is arranged; the L-shaped clip connecting rod passes under the end of the passive clip seat and the first rotation pair and the second rotation pair cooperate.

[0011] Furthermore, three positioning grooves are arranged on the active drive shaft along the circumferential direction, and a positioner is arranged in cooperation with the positioning grooves; the positioner is fixedly arranged, the positioner is hollow, and the inner diameter increases from the center to both sides, and the smallest inner diameter is the positioning tip, and the inner diameter of the positioning tip is smaller than the outer diameter of the active drive shaft; the positions of the three positioning grooves are as follows: (1) when the positioning tip is stuck in the first positioning groove close to the longitudinal drive, the active drive shaft is fixedly connected to the longitudinal drive wheel, and the active drive shaft is not fixedly connected to the transverse drive wheel; (2) when the positioning tip is stuck in the third positioning groove close to the longitudinal drive wheel, the active drive shaft is fixedly connected to the transverse drive wheel, and the active drive shaft is not fixedly connected to the longitudinal drive wheel; (3) when the positioning tip is stuck in the second positioning groove located between the first positioning groove and the third positioning groove, the active drive shaft is not fixedly connected to the longitudinal drive wheel and the transverse drive wheel.

[0012] Furthermore, it also includes a light hole turntable with a plurality of light holes with different inner diameters; the centers of all the light holes and the light holes on the operating table are on the same circle, and the light hole turntable is connected to the operating table via a rotating shaft.

[0013] Furthermore, the strain carriers include several sizes and are arranged on a strain carrier turntable; the axes of all strain carriers and the centers of the light holes are projected on the same circle. The strain carrier turntable is connected to the operating table via a rotating shaft.

[0014] A method for rapidly isolating a single cell strain of rice blast fungus to achieve the purpose of the present invention comprises the following steps:

[0015] Step 1, cutting the diseased straw at the ear neck in the field into 5-10 cm;

[0016] Step 2, rinse the straw and place it in a test tube filled with ddH2O, and place it at 26°C in the dark for 24 hours;

[0017] Step 3, transfer the straw in step 2 into a test tube containing a wet cotton ball and place it at 26°C in the dark for 48 hours;

[0018] Step 4, placing the rice straw in step 3 on the corresponding groove of the strain carrier 400;

[0019] Step 5, adjust the coarse and fine focusing knobs until the conidia can be clearly seen in the eyepiece;

[0020] Step 6, actuating the driving system 500 to move the picking needle 300 toward the straw until the needle tip can be vaguely seen in the eyepiece;

[0021] Step 7, adjusting the driving system 500, moving the picking needle 300 up and down, so that the needle head is facing a conidia;

[0022] Step 8, adjusting the fine adjustment device to attach a single conidia to the needle;

[0023] Step 9, transferring the single conidia in step 8 to the slant of a PDA test tube to achieve separation of the single spore strain of rice blast fungus.

[0024] The invention solves the problems of the existing single spore separation of rice blast fungus, such as cumbersome process, high separation cost, long separation and purification cycle, easy contamination during separation, etc. By directly collecting the conidia of rice blast fungus from the diseased rice straw in the field, the time of single spore separation of rice blast fungus is greatly shortened, and the contamination of other bacteria is avoided to a great extent. The composition of the rice blast fungus population and the dynamic trend of the change of the avirulent gene of the physiological race in each rice-growing area are understood and mastered in real time and accurately, which provides a basis for the reasonable layout of disease-resistant rice varieties, and is also helpful for the screening and cultivation of disease-resistant germplasm resources and the exploration of local rice blast resistance resources.

[0025] The present invention will be further described below in conjunction with the accompanying drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 It is a schematic diagram of the structure of the present invention.

[0027] Figure 2 This is a schematic diagram of the drive box structure.

[0028] Figure 3 Schematic diagram of the strain carrier structure.

[0029] Figure 4 This is a schematic diagram of the drive system structure.

[0030] Figure 5 It is a schematic diagram of the active drive shaft structure.

[0031] Figure 6 It is a schematic diagram of the structure of the longitudinal drive wheel and the transverse drive wheel.

[0032] Figure 7 This is a schematic diagram of the positioner structure.

[0033] Figure 8 It is a schematic diagram of the limit locking position.

[0034] Fig. 9 Schematic diagram of the clip clamp structure.

[0035] Fig.10 Schematic diagram of the active clip structure.

[0036] Fig.11 Schematic diagram of the passive clip structure.

[0037] Fig.12 Schematic diagram of the combined structure of several light holes.

[0038] Fig.13 Schematic diagram of the combined structure of several strain carriers.

[0039] Among them, the operating table 100, the drive box 200, the picking needle 300, the strain carrier 400, the drive system 500, the clip clamp 600, the light hole 700, the drive box shell 201, the fixture exposure groove 202, the strain front end groove 401, the picking needle groove 402, the strain rear end groove 403, the strain carrier ring wall 404, the strain carrier turntable 405, the active drive shaft 510, the longitudinal drive wheel 511, the locking groove 5111, the transverse drive wheel 512, the locking piece 513, the positioner 514, the positioning tip 5141, the positioner bearing 5142, the rotating handle 515, the positioning groove 51 6. Bearing 517, first longitudinal driven wheel 520, second longitudinal driven wheel 521, third longitudinal driven wheel 522, tensioning wheel 523, belt 530, fixture support platform 531, transverse rack 540, active clip 610, clip button 611, clip connecting rod 612, active clip seat 613, first rotating pair 614, active clip shaft 615, passive clip 620, passive clip seat 621, clip extension arm 622, second rotating pair 623, active clip through hole 624, needle picking groove 625, passive clip shaft 626, clip base 630, light hole turntable 701. DETAILED DESCRIPTION

[0040] In the following description, many specific details are set forth to facilitate a full understanding of the present invention, but the present invention may also be implemented in other ways different from those described herein, and those skilled in the art may make similar generalizations without violating the connotation of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below.

[0041] Secondly, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The term "in one embodiment" that appears in different places in this specification does not necessarily refer to the same embodiment, nor does it refer to a separate or selective embodiment that is mutually exclusive with other embodiments.

[0042] Secondly, in the following description, all concepts related to direction (or orientation) such as up, down, left, right, front, back, counterclockwise, clockwise, etc. are all in reference to the position state of the figure being described, and are intended to facilitate public understanding. Therefore, they cannot be understood as special limitations on the technical solution provided by the present invention.

[0043] Combination Figure 1, a device for quickly separating monospore strains of rice blast fungus, comprising an operating table 100, a drive box 200, a picking needle 300, and a strain carrier 400. Among them, a light hole 700 is arranged on the operating table 100. The drive box 200 is arranged on the operating table 100, and a drive system 500 is arranged therein. A clip clamp 600 is arranged on the drive system 500. The picking needle 300 is arranged on the clip clamp 600 and moves in a vertical or horizontal direction under the drive of the drive system 500. The strain carrier 400 is cylindrical, arranged at the light hole 700, and is used to carry processed rice straw. When separating the strains, under the drive of the drive system 500, the needle tip of the picking needle 300 moves horizontally between the center position of the strain carrier 400 and away from the center position of the strain carrier 400, and can also move in the vertical direction at the same time.

[0044] Combination Figure 2 The drive box 200 includes a drive box housing 201, and a clamp exposure groove 202 arranged on the side wall of the drive box housing 201 facing the strain carrier 400. A space for placing the drive system 500 is formed inside the drive box housing 201. The clamp exposure groove 202 is in a long strip shape, and the clip clamp 600 can extend out of the clamp exposure groove 202, which is convenient for operators to operate the clip clamp 600. A through hole is set on the other side wall of the drive box housing 201 for the rotating handle 515 in the drive system 500 to pass through, and the length of the through hole can make the tip of the pick needle 300 reach near the axis of the light hole 700.

[0045] Combination Figure 3 The strain carrier 400 includes a cylindrical strain carrier ring wall 404, which is arranged along the circumference of the light-through hole 700 to facilitate the light to enter from the light-through hole 700 to form a contrast with the strain. The strain front end groove 401, the strain rear end groove 403 and the pick needle groove 402 are arranged on the upper edge of the strain carrier ring wall 404. The strain front end groove 401 and the strain rear end groove 403 are symmetrically arranged along the axis of the strain carrier ring wall 404, so that the strain can be easily placed therein. The pick needle groove 402 is arranged on one side of the line connecting the strain front end groove 401 and the strain rear end groove 403, and the depth of the pick needle groove 402 is greater than the depth of the strain front end groove 401 and the strain rear end groove 403. The pick needle groove 402 allows the pick needle 300 to pass through.

[0046] Furthermore, the front end groove 401 of the strain can be divided into a plurality of small grooves by a partition, which are similar to the forked shape of the front end of the strain, so as to facilitate the stable placement of the strain.

[0047] Combination Figure 4The driving system 500 includes an active driving shaft 510, a longitudinal driving wheel 511, a transverse driving wheel 512, a longitudinal driven wheel assembly, a belt 530, and a transverse rack 540. The longitudinal driving wheel is fixed to one end of the active driving shaft 510. The transverse driving wheel 512 is fixed to the other end of the active driving shaft 512. The longitudinal driven wheel assembly cooperates with the longitudinal driving wheel 511 to drive the belt 530 to rotate. A clamp bearing platform 531 is provided on the belt 530. The transverse rack 540 is fixed to the bottom of the driving box housing 201 and meshes with the transverse driving wheel 512.

[0048] Further, the active drive shaft 510 can move in the axial direction. When the active drive shaft 510 moves in the direction of the longitudinal drive wheel 511, it can be fixedly connected to the longitudinal drive wheel 511 to drive the longitudinal drive wheel 511 to rotate. At this time, the active drive shaft 510 is not fixedly connected to the transverse drive wheel 512, that is, the active drive shaft 510 rotates and the transverse drive wheel 512 does not follow. On the contrary, when the active drive shaft 510 moves in the direction of the transverse drive wheel 512, it can be fixedly connected to the transverse drive wheel 512 to drive the transverse drive wheel 512 to rotate. At this time, the active drive shaft 510 is not fixedly connected to the longitudinal drive wheel 511, that is, the active drive shaft 510 rotates and the longitudinal drive wheel 511 does not follow.

[0049] As an embodiment, three positioning grooves 516 are provided on the active drive shaft 510. Figure 5 A positioner 514 is provided to cooperate with the positioning groove, and the positioner 514 covers three positioning grooves 516 along the circumference of the active drive shaft 510. The cross-sectional view of the positioner 514 is shown in FIG. Figure 7 As shown. The locator 514 is hollow and spindle-shaped, that is, the inner diameter in the middle is smaller than the inner diameters on both sides. The inner diameter in the middle is slightly smaller than the outer diameter of the active drive shaft 510, and the middle tip is the positioning tip 5141, which can just be stuck in the positioning groove 516. Locator bearings 5142 are set at both ends of the locator 514 to keep the locator 514 fixed. The locator 514 has a certain elasticity, and can be made of a physical form of setting an opening to make it elastic, or the locator 514 can be made of a deformable material. The active drive shaft 510 can move in the axial direction, and the positioning tip 5141 is stuck in or out of the three positioning grooves 516 when the active drive shaft 510 moves. The position setting scheme of the three positioning grooves 516 is as follows:

[0050] (1) a first positioning groove close to the longitudinal driving wheel 511, when the positioning tip 5141 is located in the positioning groove, the active driving shaft 510 is fixedly connected to the longitudinal driving wheel 511, and the active driving shaft 510 is not fixedly connected to the transverse driving wheel 512;

[0051] (2) a third positioning groove close to the longitudinal driving wheel 511, when the positioning tip 5141 is located in the positioning groove, the active driving shaft 510 is fixedly connected to the transverse driving wheel 512, and the active driving shaft 510 is not fixedly connected to the longitudinal driving wheel 511;

[0052] (3) The second positioning groove is located between the first positioning groove and the third positioning groove. When the positioning tip 5141 is located in the second positioning groove, the active driving shaft 510 is not fixedly connected to the longitudinal driving wheel 511 and the transverse driving wheel 512.

[0053] As an embodiment, the active drive shaft 510 is fixedly connected to the longitudinal drive gear 511 and the transverse drive gear 512 respectively as follows: Figure 6 As shown. The longitudinal drive gear 511 and the transverse drive gear 512 are provided with locking grooves 5111 on the opposite surfaces. Locking members 513 are respectively provided at both ends of the active drive shaft 510 to match the locking grooves. When the locking member 513 and the locking groove 5111 cooperate, the active drive shaft 510 can be locked with the longitudinal drive gear 511 or the transverse drive gear 512 in the rotation direction. Preferably, the locking groove 5111 is a polygonal groove, and the locking member 513 is a matching polygonal prism. The active drive shaft 510 is rotated so that the locking member 513 can be inserted into the locking groove 5111.

[0054] Further, a drive box body is arranged inside the drive box housing 201. An active drive shaft 512 is fixed to the inner wall of the drive box body through a sliding bearing, and the active drive shaft passes through the drive box body and a rotating handle 515 is arranged at one end. A longitudinal drive wheel 511 is arranged in the drive box body through a rotating bearing, and a transverse drive wheel 512 is arranged on the outer wall of the drive box body through a bearing and meshes with a transverse rack 540, and the transverse rack 540 is arranged on the inner bottom surface of the drive box housing 201. A longitudinal driven wheel assembly is arranged on the inner wall of the drive box body through a rotating shaft and a bearing. The drive box body can be an unclosed box body, at least with a through hole for exposing the fixture exposure groove 202 through a common fixture bearing platform 531. A guide rail is arranged on the inner wall of the drive box housing 201, and the drive box body moves in a horizontal direction along the guide rail. The above-mentioned positioner bearing 5142 is arranged on the inner wall of the drive box body.

[0055] As an embodiment, the longitudinal driven wheel assembly includes a first longitudinal driven wheel 520, a second longitudinal driven wheel 521, a third longitudinal driven wheel 522, and a tensioning wheel 523. The first longitudinal driven wheel 520 is meshed with the longitudinal driving wheel 511, and the center of the first longitudinal driven wheel 520 is connected to the center of the second longitudinal driven wheel 521 through a rotating shaft, and the rotating shaft is connected to the inner wall of the driving box body through a rotating bearing. The third longitudinal driven wheel 522 is arranged above the second longitudinal driven wheel 521, and is connected to the inner wall of the driving box body through a rotating shaft and a bearing. The tensioning wheel 523 is located between the second and third longitudinal driven wheels and deviates from the connecting line of the second and third longitudinal driven wheels. The tensioning wheel 523 is connected to the inner wall of the driving box body through a rotating shaft and a bearing. The belt passes around the second driven wheel 521, the tensioning wheel 523 and the third driven wheel 522, and rotates under the drive of the longitudinal driven wheel assembly.

[0056] As an embodiment, in order to prevent the active drive shaft 510 from malfunctioning when cooperating with the drive gear, a limit device is provided on the drive shaft. Figure 8 As shown. The two rotating shafts at A and B pass through the reserved through holes of the drive box housing 201, and are then locked by the limiting device. After locking, the longitudinal driven wheel assembly or the first longitudinal action wheel 520, and the gear corresponding to the rotating shaft at B cannot rotate, that is, the clip 600 cannot rotate in the vertical direction and the horizontal direction. When the active drive shaft 510 rotates and cooperates with the longitudinal drive wheel 511 or the transverse drive wheel 512, the limiting device at A and / or B is opened, and the clip 600 moves in the vertical direction or the horizontal direction under the drive of the drive system 500. The gear corresponding to the rotating shaft at B is set in the drive box housing 201 outside the above-mentioned drive box body.

[0057] Combination Fig. 9 The clip clamp 600 includes an active clip 610, a passive clip 620 and a clip base 630. Two clip bases 630 are provided and fixed on the fixture bearing platform 531. The active clip 610 is provided between the two clip bases 630 through a rotating shaft, and the passive clip 620 is provided between the two clip bases 630 through a rotating shaft. The active clip 610 is also connected to one of the clip bases 630 through a torsion spring. The rotating shaft of the active clip 610 is located in front of the rotating shaft of the passive clip 620. The active clip 610 can drive the passive clip to separate and contact with the fixture bearing platform 531.

[0058] Furthermore, the clip base 630 is fixed on the fixture support platform 531 by glue or screws.

[0059] Furthermore, the active clip 610 includes a clip button 611, a clip connecting rod 612, and an active clip seat 613. Fig.10As shown, the clip button 611 is fixedly connected to the active clip seat 613 through the clip connecting rod 612. The clip connecting rod 612 is an L-shaped connecting rod, that is, the middle part of the connecting rod is bent. The clip button 611, the clip connecting rod 612, and the active clip seat 613 are designed as an integrated whole. The side walls of the active clip seat 613 on both sides of the clip connecting rod 612 serve as the first rotation pair 614. The passive clip 620 includes a passive clip seat 621 and a clip extension arm 622. Fig.11 As shown, a pin picking groove 625 is provided at the front end of the lower bottom of the passive clip seat 621. Two clip extension arms 622 are provided and fixed to the rear end of the passive clip seat 621, and an active clip through hole 624 is formed between the two clip extension arms 622. A groove-shaped second rotation pair 623 is provided on each clip extension arm 622. During assembly, the clip connecting rod 612 passes through the active clip through hole 624, the first rotation pair 614 is provided in the second rotation pair 623, the active clip 610 is connected to the clip base 630 through a rotating shaft and a torsion spring, and the passive clip 620 is connected to the clip base 630 through a rotating shaft. When the clip button 611 rotates counterclockwise around the shaft, the two rotating pairs interact with each other to drive the passive clip seat 621 to move clockwise along the shaft, so that the passive clip seat 621 is separated from the fixture support platform 531; conversely, the passive clip seat 621 moves counterclockwise toward the fixture support platform 531, and the pick-up needle clamping groove 625 presses the pick-up needle 300. The pick-up needle clamping groove 625 can be made of rubber material, which has a certain elasticity and friction. The pick-up needle clamping groove 625 is a cuboid and a groove is set along the long axis of the lower bottom surface for placing the pick-up needle 300.

[0060] Furthermore, in order to allow different amounts of light to enter the light holes 700, a plurality of light holes 700 with different inner diameters are arranged on the light hole rotating disk 701, such as Fig.12 In order to adapt to rice straws of different sizes, the strain carrier 400 is provided with various inner diameters and heights and is placed on the strain carrier turntable 405, such as Fig.13 As shown. Preferably, the light-through hole turntable 701 is arranged below the operating table 100, and the strain carrier turntable 405 is arranged above the operating table 100. Ratchets with opposite rotation directions are respectively arranged on the light-through hole turntable 701 and the strain carrier turntable 405, and a rotating shaft passes through the ratchets of the two turntables and is arranged on the pawls matching the ratchets. When the rotating shaft rotates clockwise, the strain carrier turntable 405 rotates under the push of the corresponding pawls, and the corresponding pawls of the light-through hole turntable 701 slide on the corresponding ratchet tooth backs; when the rotating shaft rotates counterclockwise, the light-through hole turntable 701 rotates under the push of the corresponding pawls, and the corresponding pawls of the strain carrier turntable 405 slide on the corresponding ratchet tooth backs. At this time, the corresponding light hole is arranged on the operating table 100 to correspond to the through hole 700.

[0061] Preferably, the centers of the plurality of light-through holes 700 are located on the same circle, and the axes of the plurality of bacterial strain carriers 400 intersect with the same circle.

[0062] Preferably, the height of the different strain carrier 400 is 1 to 3 cm, and the inner diameter is 3 to 5 cm.

[0063] Preferably, the diameter of the needle tip of the picking needle 300 is 30-80 μm.

[0064] Furthermore, a fine adjustment device is provided on the driving box 200 to finely adjust the driving box 200 in the horizontal direction. Or the base circle diameter of the transverse driving wheel 512 is relatively large, and the teeth of the transverse driving wheel 512 and the transverse rack 540 are relatively more and dense, so that the horizontal movement step of the picking needle 300 is small.

[0065] A method for rapidly isolating a single cell strain of rice blast fungus using the above device comprises the following steps:

[0066] Step 1, cutting the diseased straw at the ear neck in the field into 5-10 cm;

[0067] Step 2, rinse the straw and place it in a test tube filled with ddH2O, and place it at 26°C in the dark for 24 hours;

[0068] Step 3, transfer the straw in step 2 into a test tube containing a wet cotton ball and place it at 26°C in the dark for 48 hours;

[0069] Step 4, placing the rice straw in step 3 on the corresponding groove of the strain carrier 400;

[0070] Step 5, adjust the coarse and fine focusing knobs until the conidia can be clearly seen in the eyepiece;

[0071] Step 6, actuating the driving system 500 to move the picking needle 300 toward the straw until the needle tip can be vaguely seen in the eyepiece;

[0072] Step 7, adjusting the driving system 500, moving the picking needle 300 up and down, so that the needle head is facing a conidia;

[0073] Step 8, adjusting the fine adjustment device to attach a single conidia to the needle;

[0074] Step 9, transferring the single conidia in step 8 to the slant of a PDA test tube to achieve separation of the single spore strain of rice blast fungus.

[0075] Before isolating the strain, move the movable ruler of the microscope to the farthest point of the objective lens so that the needle tip cannot touch the strain carrier 400, light the inoculation needle dipped in alcohol, burn the needle tip fixed on the microscope to red for sterilization, and apply liquid PDA to the needle tip.

[0076] In step 9, when the single spore is adsorbed on the needle tip, the drive system 500 is adjusted so that the needle tip is close to the rice straw. A small piece of PDA culture medium is picked up with the inoculation needle, touching the needle tip, and the single spore is transferred to the culture medium, and then transferred to the slope of the PDA test tube for cultivation.

Claims

1. A device for rapidly isolating a monospore strain of rice blast fungus, characterized in that: It comprises an operating table (100), a drive box (200), a picking needle (300), and a strain carrying table (400); wherein A light hole is provided on the operating table (100). A drive system (500) is arranged in the drive box (200). The strain carrying platform (400) is arranged along the circumference of the light hole and is used to carry the rice straw to be operated. The picking needle (300) moves in horizontal and vertical directions under the drive of the driving system (500) and the needle tip contacts the rice stalk; The driving system comprises a driving shaft (510), a longitudinal driving wheel (511), a transverse driving wheel (512), a longitudinal driven wheel assembly, a belt (530), and a transverse rack (540); The longitudinal driving wheel (511) and the transverse driving wheel (512) are respectively arranged at two ends of the active driving shaft (510). The active drive shaft (510) can move horizontally. When the active drive shaft (510) is connected to the longitudinal drive wheel (511), it is not connected to the transverse drive wheel (512). When the active drive shaft (510) is connected to the transverse drive wheel (512), it is not connected to the longitudinal drive wheel (511). The longitudinal driven wheel assembly cooperates with the longitudinal driving wheel (511) to drive the belt (530) to rotate. A clip clip (600) is provided on the belt (530) and the clip clip (600) is exposed from the drive box (200). The transverse rack (540) is fixed to the inner wall surface of the drive box (300) and meshes with the transverse drive wheel (512). The strain carrier platform (400) comprises a strain carrier ring wall (404) which is cylindrical and arranged along the circumference of the light hole. The upper bottom surface of the annular wall (404) of the strain carrier is provided with a strain front end groove (401) and a strain rear end groove (402) at both ends along the radial direction. A needle picking groove (402) is provided on one side of the bottom surface of the annular wall (404) of the strain carrier. The depth of the needle picking groove (402) is greater than the front end groove (401) and the rear end groove (402) of the strain. The front end groove (401) is divided into a number of small grooves by a spacer.

2. The device according to claim 1, characterized in that The clip clamp (600) comprises an active clip (610), a passive clip (620), and a clip base (630); The clip base (630) is fixed on a fixture support platform (531) provided on the belt (530). The active clip (610) is connected to the clip base (630) via a rotating shaft and a torsion spring. The passive clip (620) is connected to the clip base (630) via a rotating shaft; The active clamping piece (610) drives the passive clamping piece (620) to separate from or contact the upper bottom surface of the clamp bearing platform (531).

3. The device according to claim 2, characterized in that The active clip (610) comprises a clip button (611), an L-shaped clip connecting rod (612), and an active clip seat (613). One end of the L-shaped clip link (612) fixes the clip button (611) and the other end fixes the active clip seat (613). The side walls of the active clip seat (613) on both sides of the L-shaped clip connecting rod (612) are the first rotation pair (614); The passive clip (620) comprises a passive clip seat (621) and a clip extension arm (622). The clip extension arm (622) is arranged at the end of the passive clip seat (621) and is provided with a second rotation pair (623) with a groove; The L-shaped clip connecting rod (612) passes under the end of the passive clip seat (621) and The first rotating pair (614) and the second rotating pair (623) cooperate with each other.

4. The device according to claim 1, characterized in that Three positioning grooves (516) are arranged on the active drive shaft (510) along the circumferential direction, and a positioner (514) is arranged in cooperation with the positioning grooves; The positioner (514) is fixedly set, The positioner (514) is hollow and its inner diameter increases from the center to both sides, and the smallest inner diameter is the positioning tip (5141). The inner diameter of the positioning tip (5141) is smaller than the outer diameter of the active drive shaft (510); The positions of the three positioning grooves are as follows: (1) When the positioning tip (5141) is stuck in the first positioning groove close to the longitudinal driving wheel (511), the active driving shaft (510) is fixedly connected to the longitudinal driving wheel (511), and the active driving shaft (510) is not fixedly connected to the transverse driving wheel (512); (2) When the positioning tip (5141) is stuck in the third positioning groove close to the longitudinal drive wheel (511), the active drive shaft (510) is fixedly connected to the transverse drive wheel (512), while the active drive shaft (510) is not fixedly connected to the longitudinal drive wheel (511); (3) When the positioning tip (5141) is stuck in the second positioning groove located between the first positioning groove and the third positioning groove, the active drive shaft (510) is not fixedly connected to the longitudinal drive wheel (511) and the transverse drive wheel (512).

5. The device according to claim 1, characterized in that It also includes a light-through hole rotating disk (701) provided with a plurality of light-through holes (700) having different inner diameters; The centers of all the light holes (700) and the light holes on the operating table (100) are on the same circle. The light-through hole rotating disk (701) is connected to the operating table (100) via a rotating shaft.

6. The device according to claim 1 or 2, characterized in that: The strain carrier platform (400) includes several sizes and is arranged on the strain carrier turntable (405); The axes of all strain carriers (400) and the centers of the light holes are projected onto the same circle; The bacterial strain carrier turntable (405) is connected to the operating table (100) via a rotating shaft.

7. A method for rapidly isolating a monospore strain of rice blast fungus using the device according to any one of claims 1 to 6, characterized in that: The following steps are involved: Step 1, cutting the diseased straw at the ear neck in the field into 5-10 cm; Step 2, rinse the straw and place it in a test tube filled with ddH2O, and place it at 26°C in the dark for 24 hours; Step 3, transfer the straw in step 2 into a test tube containing a wet cotton ball and place it at 26°C in the dark for 48 hours; Step 4, placing the rice straw in step 3 on the corresponding groove of the strain supporting platform (400); Step 5, adjust the coarse and fine focusing knobs until the conidia can be clearly seen in the eyepiece; Step 6, actuating the driving system (500) to move the picking needle (300) toward the straw until the needle tip can be vaguely seen in the eyepiece; Step 7, adjusting the driving system (500) to move the picking needle (300) up and down so that the needle head is facing a conidia; Step 8, adjusting the fine adjustment device to attach a single conidia to the needle; Step 9, transferring the single conidia in step 8 to the slant of a PDA test tube to separate the single spore strain of rice blast fungus.

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