CN110923121B - Single spore separation device, single spore separation microscope and single spore separation method - Google Patents
Single spore separation device, single spore separation microscope and single spore separation method Download PDFInfo
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Abstract
The invention discloses a single spore separation device, a single spore separation microscope and a single spore separation method. The single spore separation device comprises a lifting regulator, a connecting rod, a needle-shaped object fastener and a separation needle; the lifting regulator is arranged at the position of the collecting lens lifting regulator on the microscope arm; the lifting hand wheel of the microscope is used for controlling the lifting regulator to ascend or descend; one end of the connecting rod is connected to the lifting adjuster, and the other end of the connecting rod is connected to the needle-shaped object fastener; one end of the separation needle is locked inside the needle-shaped object fastener. The invention has the advantages that: the accuracy of single spore separation is ensured; the height of the separation needle can be adjusted; the separation needle is not easy to break; frequent focusing is not required; the operation process is simple.
Description
Technical Field
The invention belongs to the technical field of optical microscopes.
Background
The fungal monospore isolation technology is the most basic key technology in the fungal research process. Identification of fungal diseases, pathogenicity analysis, fungal phylogeny and genetic variation analysis and the like all require that pathogenic bacteria are pure strains propagated from one spore, and single spore isolation is required.
To date, there have been many studies on techniques for isolating fungal monospores. Such as the early dilution separation method, agar plate cutting method, glass capillary separation method, and the later fixed insect needle picking method, eyebrow needle picking method, solid lens picking method, and the like, and further, there are methods which are improved on the basis of these methods, such as sporulation needle picking method (picking single spores with a self-made sporulation needle under a microscope), medium picking method (searching single spores which have not branched yet and have no other spores around just under a microscope, cutting the medium at the position of spores with the side surface of a needle, gently picking a small piece of medium with a needle), agar block cutting method (spreading a spore suspension on a medium, cutting agar blocks with a scalpel to transfer onto a slide, cutting off excess spores under a microscope, ensuring that the remaining agar blocks contain only one spore), medium streaking cutting method (drawing parallel lines on a medium plate, then, the prepared spore suspension is smeared on the wire by a pipette gun, a single colony which germinates after culture is dug away), and the agar block needle picking method (the agar block is cut and put on a sterile glass slide, the spores are smeared on the aseptic glass slide, the excess spores are removed by a picking needle, after only one spore is ensured on each agar block, the agar block is transferred to a flat plate or a slant culture medium for culture).
Unfortunately, the above described methods of monospore isolation all have some drawbacks. Some methods are highly technical and have high requirements on experimenters, and can be completed by skilled personnel after long-term training. For example, singlespores are picked up by hands under a microscope, the requirement on the stability of hand movements of experimenters is extremely high, even tiny shaking can cause great change under the enlarged visual field of the microscope, so that tiny spores are very difficult to pick, and the singlespores are difficult to ensure; some methods are complicated in tool manufacturing, for example, a small-size insect needle is fixed on an inoculation rod for fixing the inoculation needle, the required process is complicated, the tip of the insect needle needs to be knocked into a slightly flat wedge shape, and the insect needle needs to be ground into a sharp thin needle; most importantly, the methods cannot be finely operated, and the method cannot guarantee that the true single spore is picked. For example, when the culture medium is dug by hand under a microscope, tiny spores are easily trapped in gaps during the digging process and are ignored because the tiny spores cannot be seen, and the spores are mixed with hyphae of other spore germinants after germination, so that formed colonies are not single spore colonies. In addition, in the process of removing the redundant spores from the culture medium, the field of view is large, so that the spores in the field of view are easy to be overlooked in the cutting process, so that an operator mistakenly thinks that the picked spores are single spores, and as a result, the formed colonies are not single spore colonies actually.
Therefore, the existing single spore separation technology is difficult to really realize single spore separation and cannot meet the requirement of fungus research and development.
The monospore separating microscope disclosed in patent CN 205691850U comprises an existing microscope 1, a monospore separating filament 6 is fixed on a base 8 of the microscope 1, the lower end of the monospore separating filament 6 is fixed on a fixing ring 5, and the fixing ring 5 is sleeved on the outer wall of a light source 4.
Although the monospore isolation microscope disclosed in patent CN 205691850U is a great improvement in technology, it still has the disadvantages of simple structure and inconvenient operation. The angle and the length of the single spore isolation filament of the single spore isolation microscope are fixed, so that the single spore isolation microscope has the following defects:
1. the height of the single spore separating filament is not adjustable, and the single spore can not be ensured to be separated. The height of the single spore isolation wire of the single spore isolation microscope cannot be adjusted, and the structure determines that the top end of the single spore isolation wire is always contacted with the bottom of the culture medium in order to clearly see the needle tip and the spore under the magnified microscope field at the same time in the single spore isolation process. When the single spore isolation silk is scratched on the surface of the culture medium during the moving process of the object stage, other spores or impurities can be adhered to the top end of the single spore isolation silk, so that the single spore is not isolated or the isolated single spore is polluted.
2. The single spore isolation strand was easily broken. Since the top of the single spore isolation thread is always in contact with the bottom of the medium, but the surface of the medium is not a uniform horizontal plane, the slightly convex medium forms resistance during the movement of the stage, and the fragile single spore isolation thread is easily broken.
3. The requirements for single spore separating filaments are high, and the single spore separating filaments meeting the requirements can be manufactured only by skilled technicians. The top end of the single spore separating filament which can be clearly seen in the microscope magnifying visual field by utilizing the lifting hand wheel is a narrow focusing range, even a fixed point of the lifting hand wheel. Because the length and the angle of the single spore separating filament of the single spore separating microscope can not be adjusted, the single spore separating filament is longer or shorter, or the angle of the single spore separating filament deviates from a point, so that the needle tip of the single spore separating filament can not be clearly seen under the microscope magnifying visual field, or the needle tip can not be simultaneously and clearly seen with the spore on the culture medium. Therefore, it is difficult for beginners or ordinary operators to produce single spore isolation filaments with the correct length and angle.
4. The thickness of the single spore separation silk can not be adjusted: the single spore separating filament is fixed on the microscope base, the aperture of the single spore separating filament is fixed, so that only single spore separating filament with one specification can be placed, and single spore separating filament with too thick or too thin cannot be placed. However, the size of spores of different fungi is different greatly, large spores cannot be picked by fine single spore separating filaments, while small spores are easily polluted by coarse separating filaments, and in actual operation, single spore separating filaments with different specifications need to be used for working according to the size of the spores.
5. Frequent focusing is required, and the efficiency is low. In the existing single spore isolation microscope, a single spore isolation filament is contacted with a culture substrate part, and a lifting hand wheel and an objective table are required to be moved for many times to find out a single spore with ideal distribution, and focusing is required to be carried out during each movement. Frequent focusing tends to cause eye fatigue to the operator, thereby reducing work efficiency.
6. The operation is complicated, and redundant objective lenses need to be disassembled and assembled back and forth. The existing single spore separation microscope has the defects that the length of a single spore separation filament cannot be adjusted, so that in the single spore picking process, redundant objective lenses need to be disassembled, otherwise a culture dish cannot be placed on the microscope, and the single spore separation filament is broken if the microscope is placed forcibly. In order to restore the function of a common microscope after the single spore is separated, other objective lenses are required to be mounted, and the operation is complicated due to the fact that the objective lenses are dismounted and mounted back and forth.
Disclosure of Invention
The present invention is improved as follows against the disadvantages of the prior art.
The invention discloses a needle-shaped object fastener, which is provided with an external thread turnbuckle and an internal thread nut at the end part. The body of the external thread insert is a hollow cylinder with external threads; the body of the internal thread nut is a hollow cylinder with internal threads, the hollow cylinder is provided with the internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut and the external thread sleeve are mutually meshed. The fastener also has a hollow cone with a Y-shaped slot or a cross slot, the end of the hollow cone is horizontally truncated. The hollow cone is made of an elastic material. The needle-like object is locked within the hollow cone.
In one structure of the needle-like object fastener of the invention, the fastener has an externally threaded barrel and an internally threaded nut at its end. The body of the external thread insert is a hollow cylinder with external threads. The body of the internal thread nut is a hollow cylinder with internal threads. The threads of the internal thread nut and the external thread sleeve are mutually meshed. The fastener also has a hollow cone with a Y-shaped slot or a cross slot, the end of the hollow cone is horizontally truncated. The hollow cone and the external thread turnbuckle are connected into a whole. The hollow cone is made of an elastic material. The needle-like object is locked within the hollow cone.
In another structure of the needle-like object fastener of the invention, the fastener has an externally threaded barrel, a hollow cone, and an internally threaded nut at its end. The body of the external thread insert is a hollow cylinder with external threads. The body of the internal thread nut is a hollow cylinder with internal threads. The threads of the internal thread nut and the external thread sleeve are mutually meshed. The fastener also has a hollow cone with a Y-shaped slot or a cross slot, the end of the hollow cone is horizontally truncated. The hollow cone is a separate part located inside the internally threaded nut. The hollow cone is made of an elastic material. The needle-like object is locked within the hollow cone.
Further, in the needle-like object fastener of the present invention, the elastic material of which the hollow cone is made may be an elastic metal, such as 65 manganese steel.
The invention also discloses a single spore separation device which comprises a lifting regulator, a connecting rod, a needle-shaped object fastener and a separation needle. The lifting adjuster is arranged at the position of the condensing lens lifting adjuster on the microscope arm, and the lifting adjuster is controlled to ascend or descend by utilizing a lifting hand wheel of the microscope. One end of the connecting rod is connected to the lifting adjuster, and the other end of the connecting rod is connected to the needle-shaped object fastener. One end of the separation needle is locked inside the needle-shaped object fastener.
Further, in a single spore isolation apparatus of the present invention, the needle-like object fastening member has an externally threaded barrel and an internally threaded nut at its end portion. The body of the external thread insert is a hollow cylinder with external threads; the body of the internal thread nut is a hollow cylinder with internal threads, the hollow cylinder is provided with the internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut and the external thread sleeve are mutually meshed. The fastener is also provided with a hollow cone, the hollow cone is provided with a Y-shaped slot or a cross slot, and the end part of the hollow cone is horizontally cut off; the hollow cone and the external thread turnbuckle are connected into a whole. The hollow cone is made of an elastic material. The needle-like object is locked within the hollow cone.
Further, in another monospore separating apparatus of the present invention, the needle-like object fastening member has an externally threaded barrel and an internally threaded nut at its end portion. The body of the external thread insert is a hollow cylinder with external threads; the body of the internal thread nut is a hollow cylinder with internal threads, the hollow cylinder is provided with the internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut and the external thread sleeve are mutually meshed. The fastener is also provided with a hollow cone, the hollow cone is provided with a Y-shaped slot or a cross slot, and the end part of the hollow cone is horizontally cut off; the hollow cone is a separate part located inside the internally threaded nut. The hollow cone is made of an elastic material. The needle-like object is locked within the hollow cone.
With the needle-shaped object fastener having the structure of the present invention, since the hollow cone is made of an elastic material, the male threaded barrel can be locked with the female threaded nut, so that the hollow cone inside the female threaded nut locks the separation needle. The separating needles with different specifications, namely different thicknesses, can be locked corresponding to different locking degrees. The hollow aperture of the hollow cone of the needle-shaped object fastener can be adjusted by utilizing the locking degrees of the internal thread nut and the external thread screw sleeve, the aperture corresponding to the low locking degree is large, and the aperture corresponding to the high locking degree is small, so that the separation needles with different specifications can be selected according to the size of the separated fungal spores to be locked.
Further, in the single spore isolation apparatus of the present invention, the isolation needle may be made of a material such as metal, alloy, polymer fiber, or glass fiber.
Further, in the single spore isolation apparatus of the present invention, the end of the isolation needle may be straight or bent.
Further, in the single spore isolation apparatus of the present invention, the end of the isolation needle may have a sharp shape.
Further, in the single spore isolation apparatus having the above-described structure, the connection structure of the connection rod and the needle-like object fastener may be a steerable connection structure.
Further, in the single spore isolation apparatus having the above-described structure, the connection structure of the connection rod and the needle-shaped object fastener may be connected by a steering joint or by a ball pivot joint.
The invention also discloses the application of the monospore separation device in an optical microscope. The monospore isolation microscope of the present invention is an optical microscope, and may be an upright microscope or an inverted microscope.
The invention also discloses a connecting method of the monospore separating device, wherein one end of the connecting rod is connected with the needle-shaped object fastener; the other end of the connecting rod is connected to a lifting regulator on a microscope arm, and the lifting regulator is controlled to ascend or descend by utilizing a lifting hand wheel of a microscope; one end of the separation needle is locked inside the needle-shaped object fastener.
Further, in the above-described connecting method, one end of the connecting rod is connected to the needle-like object fastener through a steering joint or a ball pivot joint. The connecting rod and the needle-shaped object fastener form an included angle, the angle is variable, and the needle-shaped object fastener can rotate relative to the connecting rod as required.
The invention also discloses an optical microscope, which comprises a microscope base, a microscope arm, a lens cone, an ocular, an objective lens, a lifting hand wheel and an object stage, wherein the object stage is provided with a longitudinal adjuster and a transverse adjuster, and the microscope arm of the single-spore separation microscope is provided with the single-spore separation device. The microscope arm is provided with a lifting regulator of the single spore separation device, and the lifting regulator is controlled to ascend or descend by a lifting hand wheel of the microscope. Here, the elevation adjuster on the mirror arm is installed at the position of the collector stand elevation adjuster, and the elevation adjuster is connected to the mirror arm in the same manner as the collector stand elevation adjuster. The collecting lens support connected to the collecting lens support lifting adjuster, the collecting lens and the iris diaphragm connected to the collecting lens support need to be dismounted, and the lifting adjuster is installed at the position of the collecting lens support lifting adjuster.
The invention also discloses a single spore separation method, which comprises the following steps:
(1) smearing conidia on the surface of a transparent culture medium laid at the bottom of a culture dish;
(2) fixing the culture dish on a microscope objective table, and enabling the surface of the culture medium to face the separation needle;
(3) rotating a longitudinal regulator and/or a transverse regulator of a microscope objective table to find a target monospore under a microscope visual field;
(4) controlling a lifting regulator by using a lifting hand wheel to enable a separation needle to approach a target monospore;
(5) controlling a lifting regulator by using a lifting hand wheel to enable a separation needle to contact the culture medium;
(6) adjusting a longitudinal adjuster and/or a transverse adjuster of a microscope objective table to control the objective table, and picking the spores onto a separation needle;
(7) the lifting hand wheel is used for controlling the lifting regulator to move the separation needle, so that the separation needle is not contacted with the culture medium completely,
(8) spores were placed on clean medium.
The invention has the advantages that:
1. the accuracy of single spore separation is ensured. According to the invention, monospore is picked by controlling the longitudinal regulator and/or the transverse regulator, so that misoperation caused by hand shaking is effectively avoided; in addition, in the monospore separation process, the spores are picked from the surface of the culture medium and are placed on the new surface of the culture medium to be clearly visible, so that the true monospores are ensured to be separated.
2. The height of the separating needle can be adjusted. In the invention, the separation needle is fixed on the needle-shaped object fastener, in the process of separating the single spore, the needle-shaped object fastener is moved to enable the separation needle to be contacted with the surface of the culture medium after the target single spore is selected (namely, no other spores exist around the needle-shaped object fastener, or the other spores are far away from the needle-shaped object fastener, so that the operation is not interfered), the separation needle is moved to enable the separation needle not to be contacted with the culture medium after the single spore is picked, the opportunity that the separation needle is contacted with other spores is avoided, and the single spore is ensured to be separated.
3. The separation needle is not easy to break. In the invention, the lifting regulator is used for controlling the separation needle to move up and down, so that the contact between the separation needle and the surface of the culture medium in the moving process of the objective table can be effectively avoided, and the situation that the separation needle is broken due to the resistance from the uneven surface of the culture medium can not occur.
4. There is no particular requirement for the separation needle gauge. In the invention, the separating needle is fixed on the needle-shaped object fastener, not only can move up and down, but also the needle-shaped object fastener can rotate under the condition that the connecting rod is connected with the needle-shaped object fastener through the steering joint or the spherical rotating joint, so that the length and the angle of the separating needle are not particularly high.
5. The separating needle with different specifications can be selected according to the size of the spore. According to the invention, the hollow aperture of the hollow cone of the needle-shaped object fastener can be adjusted, and the separation needles with different specifications can be selected for fixing according to the size of the picked fungal spores, so that the fungal spores with different sizes can be subjected to single spore separation.
6. Frequent focusing is not needed, and the working efficiency is greatly improved. In the invention, after the single spore is found to be well focused, the next process does not need to be focused, and the visual fatigue of operators caused by frequent focusing can not be caused, thereby improving the working efficiency.
7. Unnecessary objective lens is not required to be disassembled and assembled. In the invention, the position of the separating needle can be adjusted, and when the culture dish needs to be placed, the separating needle and the objective table are moved away from the objective lens, so that the objective lens does not obstruct the placing of the culture dish, and the redundant objective lens does not need to be disassembled and assembled.
8. The operation process is simple. The monospore separation process of the invention is simple, and even a beginner can master the technology quickly.
9. The monospore separation process can be performed by demonstration, video recording and photography: the biological microscope provided with the single spore separation device can be selectively provided with the camera, so that the whole single spore separation process can be demonstrated on a computer or a large screen, the demonstration of a single spore separation technology is facilitated, and image data can be left in the separation process for storage.
Drawings
Fig. 1 is an exploded view of a needle-like object fastener according to an embodiment of the present invention.
FIG. 2 is a schematic front view of a single spore isolation microscope equipped with a single spore isolation apparatus according to an embodiment of the present invention.
FIG. 3 is a schematic side view of a single spore isolation microscope equipped with a single spore isolation apparatus according to an embodiment of the present invention.
Fig. 4 is a schematic view illustrating the angle adjustment of a needle-shaped object fastener of the single spore isolation apparatus according to an embodiment of the present invention.
Detailed Description
Specifically, the present invention is as follows:
1. a needle-like object fastener characterized by: the needle-shaped object fastener is provided with an external thread turnbuckle and an internal thread nut at the end part; the body of the external thread turnbuckle is a hollow cylinder; the body of the internal thread nut is a hollow cylinder, the hollow cylinder is provided with internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut and the external thread sleeve are mutually meshed; the needle-shaped object fastener is also provided with a hollow cone, the hollow cone is provided with a Y-shaped slot or a cross slot, and the end part of the hollow cone is horizontally cut off; the hollow cone and the external thread threaded sleeve are connected into a whole; the hollow cone is made of an elastic material; the needle-like object is locked within the hollow cone.
2. A needle-like object fastener characterized by: the needle-shaped object fastener is provided with an external thread turnbuckle and an internal thread nut at the end part; the body of the external thread turnbuckle is a hollow cylinder; the body of the internal thread nut is a hollow cylinder, the hollow cylinder is provided with internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut and the external thread sleeve are mutually meshed; the needle-shaped object fastener is also provided with a hollow cone, the hollow cone is provided with a Y-shaped slot or a cross slot, and the end part of the hollow cone is horizontally cut off; the hollow cone is an independent part and is positioned on the inner side of the internal thread nut; the hollow cone is made of an elastic material; the needle-like object is locked within the hollow cone.
3. The needle-like object fastener according to claim 1 or 2, characterized in that: the elastic material is elastic metal.
4. The needle-like object fastener according to claim 1 or 2, characterized in that: the elastic material is 65 manganese steel.
5. A single spore separation device comprises a lifting adjuster, a connecting rod, a needle-shaped object fastener and a separation needle; the lifting regulator is arranged at the position of the condensing lens lifting regulator on the microscope arm, and the lifting regulator is controlled to ascend or descend by utilizing a lifting hand wheel of the microscope; one end of the connecting rod is connected to the lifting adjuster, and the other end of the connecting rod is connected to the needle-shaped object fastener; one end of the separation needle is locked inside the needle-shaped object fastener.
6. A single spore separation device comprises a lifting adjuster, a connecting rod, a needle-shaped object fastener and a separation needle; the lifting regulator is arranged at the position of the condensing lens lifting regulator on the microscope arm, and the lifting regulator is controlled to ascend or descend by utilizing a lifting hand wheel of the microscope; one end of the connecting rod is connected to the lifting adjuster, and the other end of the connecting rod is connected to the needle-shaped object fastener; one end of the separation needle is locked inside the needle-shaped object fastener; the needle-shaped object fastener is provided with an external thread turnbuckle and an internal thread nut at the end part; the body of the external thread turnbuckle is a hollow cylinder; the body of the internal thread nut is a hollow cylinder, the hollow cylinder is provided with internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut and the external thread sleeve are mutually meshed; the needle-shaped object fastener is also provided with a hollow cone, the hollow cone is provided with a Y-shaped slot or a cross slot, and the end part of the hollow cone is horizontally cut off; the hollow cone and the external thread threaded sleeve are connected into a whole; the hollow cone is made of an elastic material.
7. A single spore separation device comprises a lifting adjuster, a connecting rod, a needle-shaped object fastener and a separation needle; the lifting regulator is arranged at the position of the condensing lens lifting regulator on the microscope arm, and the lifting regulator is controlled to ascend or descend by utilizing a lifting hand wheel of the microscope; one end of the connecting rod is connected to the lifting adjuster, and the other end of the connecting rod is connected to the needle-shaped object fastener; one end of the separation needle is locked inside the needle-shaped object fastener; the needle-shaped object fastener is provided with an external thread turnbuckle and an internal thread nut at the end part; the body of the external thread turnbuckle is a hollow cylinder; the body of the internal thread nut is a hollow cylinder, the hollow cylinder is provided with internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut and the external thread sleeve are mutually meshed; the needle-shaped object fastener is also provided with a hollow cone, the hollow cone is provided with a Y-shaped slot or a cross slot, and the end part of the hollow cone is horizontally cut off; the hollow cone is an independent part and is positioned on the inner side of the internal thread nut; the hollow cone is made of an elastic material.
8. The monospore separation apparatus according to item 5, wherein: the separating needle is made of metal, alloy, polymer fiber or glass fiber.
9. The monospore separation apparatus according to item 5, wherein: the ends of the separating needles are straight or bent.
10. The monospore separation apparatus according to item 5, wherein: the tip of the separation needle has a sharp shape.
11. The monospore separation apparatus according to item 6, wherein: the separating needle is made of metal, alloy, polymer fiber or glass fiber.
12. The monospore separation apparatus according to item 6, wherein: the ends of the separating needles are straight or bent.
13. The monospore separation apparatus according to item 6, wherein: the tip of the separation needle has a sharp shape.
14. The monospore separation apparatus according to item 7, wherein: the separating needle is made of metal, alloy, polymer fiber or glass fiber.
15. The monospore separation apparatus according to item 7, wherein: the ends of the separating needles are straight or bent.
16. The monospore separation apparatus according to item 7, wherein: the tip of the separation needle has a sharp shape.
17. The monospore separation apparatus according to item 8, wherein: the ends of the separating needles are straight or bent.
18. The monospore separation apparatus according to item 8, wherein: the tip of the separation needle has a sharp shape.
19. The monospore separation apparatus according to claim 9, wherein: the tip of the separation needle has a sharp shape.
20. The monospore separation apparatus according to claim 11, wherein: the ends of the separating needles are straight or bent.
21. The monospore separation apparatus according to claim 11, wherein: the tip of the separation needle has a sharp shape.
22. The monospore separation apparatus according to claim 12, wherein: the tip of the separation needle has a sharp shape.
23. The monospore separation apparatus according to claim 14, wherein: the ends of the separating needles are straight or bent.
24. The monospore separation apparatus according to claim 14, wherein: the tip of the separation needle has a sharp shape.
25. The monospore separation apparatus according to claim 15, wherein: the tip of the separation needle has a sharp shape.
26. The monospore separation apparatus according to any one of claims 5 to 25, wherein: the connecting rod is connected with the needle-shaped object fastener through a steering joint.
27. The monospore separation apparatus according to any one of claims 5 to 25, wherein: the connecting rod is connected with the needle-shaped object fastener through a sphere rotating joint.
28. The utility model provides an optical microscope, includes microscope base, microscope arm, lens cone, eyepiece, objective, lifting hand wheel, objective table, has vertical regulator and horizontal regulator on the objective table, its characterized in that: the optical microscope is equipped with the monospore separation apparatus described in any one of items 5 to 27.
29. Use of a monospore separation apparatus according to any one of claims 5 to 27 in an optical microscope.
30. The method for connecting a single spore isolation apparatus according to any one of claims 5 to 27, wherein the method comprises the steps of: one end of the connecting rod is connected with the needle-shaped object fastener; the other end of the connecting rod is connected to the lifting adjuster on the microscope arm, and the lifting adjuster is controlled to ascend or descend by a lifting hand wheel of a microscope; one end of the separation needle is locked inside the needle-shaped object fastener.
31. The method of connecting a single spore isolation apparatus according to item 30, wherein the method comprises the steps of: one end of the connecting rod is connected with the needle-shaped object fastener through a steering joint or a ball rotating joint.
32. A monospore isolation method comprises the following steps:
(1) smearing conidia on the surface of a transparent culture medium laid at the bottom of a culture dish;
(2) fixing the culture dish on a microscope objective table, and enabling the surface of the culture medium to face the separation needle;
(3) rotating a longitudinal regulator and/or a transverse regulator of a microscope objective table to find a target monospore under a microscope visual field;
(4) controlling a lifting regulator by using a lifting hand wheel to enable a separation needle to approach a target monospore;
(5) controlling a lifting regulator by using a lifting hand wheel to enable a separation needle to contact the culture medium;
(6) adjusting a longitudinal adjuster and/or a transverse adjuster of a microscope objective table to control the objective table, and picking the spores onto a separation needle;
(7) the lifting hand wheel is used for controlling the lifting regulator to move the separation needle, so that the separation needle is not contacted with the culture medium completely,
(8) spores were placed on clean medium.
The present invention will be described with reference to the accompanying drawings.
For a better understanding of the present invention, reference will now be made to the following descriptions taken in conjunction with the accompanying drawings. However, the specific embodiments set forth herein are merely illustrative of the invention and are not to be construed as limiting the invention. The scope of the invention is defined only by the claims of the invention and not by the specific embodiments. Any modification and variation of the present invention, and any equivalent substitution, are within the scope of the present invention without departing from the spirit of the present invention.
Fig. 1 is an exploded view of a needle-like object fastener according to an embodiment of the present invention. The needle-like object fastener 5 of one embodiment of the present invention has the following structure: the needle-like object fastener 5 has an externally threaded barrel 12 and an internally threaded nut 7 at its end; the body of the external thread insert 12 is a hollow cylinder with external threads; the body of the internal thread nut 7 is a hollow cylinder with internal threads, the hollow cylinder is provided with the internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut 7 and the external thread insert 12 are mutually meshed; the fastener 5 also has a hollow cone 6, which is a separate part inside the internally threaded nut; the hollow cone 6 is provided with a Y-shaped slot, and the end part of the hollow cone 6 is horizontally cut off; the hollow cone 6 is made of an elastic material; the needle-like object is locked within the hollow cone.
In the structure of the needle-like object fastener of another embodiment of the present invention, the needle-like object fastener 5 has an externally threaded barrel 12, an internally threaded nut 7 at its end; the body of the external thread insert 12 is a hollow cylinder with external threads; the body of the internal thread nut 7 is a hollow cylinder with internal threads, the hollow cylinder is provided with the internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off; the threads of the internal thread nut 7 and the external thread insert 12 are mutually meshed; the fastener 5 is also provided with a hollow cone 6 which is connected with the external thread turnbuckle into a whole; the hollow cone 6 is provided with a Y-shaped slot, and the end part of the hollow cone 6 is horizontally cut off; the hollow cone 6 is made of an elastic material; the needle-like object is locked within the hollow cone.
FIG. 2 is a schematic front view of a single spore isolation microscope equipped with a single spore isolation apparatus according to an embodiment of the present invention. FIG. 3 is a schematic side view of a single spore isolation microscope equipped with a single spore isolation apparatus according to an embodiment of the present invention.
The monospore separating device comprises a lifting adjuster 4, a connecting rod 3, a needle-shaped object fastener 5 and a separating needle 8. The lifting adjuster 4 is square, and the lifting adjuster 4 is arranged at the position of the condensing lens lifting adjuster on the microscope arm 2. The lifting adjuster 4 is connected with a condenser lifting adjuster on the microscope arm in the same way, and the lifting adjuster 4 is controlled to ascend or descend by a lifting hand wheel 10 of the microscope. One end of the connecting rod 3 is connected to the lifting adjuster 4, and the other end of the connecting rod 3 is connected to the needle-shaped object fastener 5. The connecting rod 3 is connected to the lifting adjuster 4. One end of the separation needle 8 is locked inside the needle-like object fastener 5.
In the structure of the single spore isolation apparatus according to one embodiment of the present invention, as shown in fig. 1, the needle-like object fastening piece 5 has an externally threaded insert 12, a hollow cone 6 and an internally threaded nut 7 at its end. The body of the externally threaded insert 12 is a hollow cylinder with external threads. The body of the internal thread nut 7 is a hollow cylinder with internal threads, the hollow cylinder is provided with internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off. The threads of the internally threaded nut 7 and the externally threaded barrel 12 are engaged with each other. The hollow cone is an independent part and is positioned on the inner side of the internal thread nut, the hollow cone is provided with a Y-shaped groove, and the end part of the hollow cone is horizontally cut off. The separation needle 8 is locked within the hollow cone 6 by locking the externally threaded barrel 12 using the internally threaded nut 7. The hollow cone 6 is made of an elastic material, which may be, for example, an elastic metal, such as 65 manganese steel.
In the structure of the single spore isolation apparatus according to another embodiment of the present invention, the needle-like object fastening member 5 has an externally threaded barrel 12 and an internally threaded nut 7 at its end. The body of the externally threaded insert 12 is a hollow cylinder with external threads. The body of the internally threaded nut 7 is a hollow cylinder with internal threads. The threads of the internally threaded nut 7 and the externally threaded barrel 12 are engaged with each other. The needle-shaped object fastener 5 is also provided with a hollow cone 6, the hollow cone 6 is connected with the external thread screw sleeve 12 into a whole, the hollow cone 6 is provided with a Y-shaped slot, and the end part of the hollow cone is horizontally cut off. The separation needle 8 is locked within the hollow cone 6 by locking the externally threaded barrel 12 using the internally threaded nut 7. The hollow cone 6 is made of an elastic material, which may be, for example, an elastic metal, such as 65 manganese steel.
In the structure of the monospore separation apparatus of the present invention, the needle-like object fastening member 5 has an externally threaded insert 12, a hollow cone 6 and an internally threaded nut 7 at its end. Because the hollow cone 6 at the end part of the needle-shaped object fastener 5 is of a slotted structure, the internal thread nut 7 can lock the external thread turnbuckle 12, so that the hollow cone 6 tightly clamps the slotted conical end part of the internal thread nut 7 to fix the separating needles 8 with different thicknesses.
In the structure of the single spore separation device, the hollow aperture of the hollow cone of the needle-shaped object fastener 5 can be adjusted by utilizing different locking degrees of the internal thread nut and the external thread screw sleeve 12, the aperture corresponding to low locking degree is large, and the aperture corresponding to high locking degree is small, so that separation needles with different specifications can be selected for locking according to the size of the separated fungal spores.
Fig. 4 is a schematic view illustrating the angle adjustment of a needle-shaped object fastener of the single spore isolation apparatus according to an embodiment of the present invention.
In the configuration of the single spore isolation apparatus according to one embodiment of the present invention, as shown in fig. 4, the needle-like object fastening piece 5 and the connecting rod 3 have a steering joint 11 therebetween, which can be rotated.
In the structure of the single spore isolation apparatus according to an embodiment of the present invention, the needle-shaped object fastening member 5 and the connecting rod 3 have a spherical rotation joint therebetween, and can rotate in a three-dimensional space.
The separation needle 8 of the monospore separation apparatus of the present invention may be made of a material such as metal, alloy, polymer fiber or glass fiber.
The end part of the separating needle 8 of the single spore separating device can be straight or bent, so that the included angle between the needle tip and the base part of the separating needle is an obtuse angle, such as an angle of 135 degrees, and single spores can be picked conveniently.
The end of the separating needle 8 of the single spore separating device of the invention can be in a sharp shape to ensure that the needle tip is thin enough and small single spores can be picked.
The single spore separating device can be arranged on a common optical microscope, and can be an upright microscope or an inverted microscope.
In one embodiment of the connecting method of the present invention, the needle-like object fastener 5 is connected to the lift adjuster 4 through the connecting rod 3; one end of the connecting rod 3 is connected to a lifting regulator 4 on a microscope arm, and a lifting hand wheel 10 controls the lifting regulator 4 to ascend or descend; the other end of the connecting rod 3 is connected with the needle-shaped object fastener 5 through the steering joint 11, the connecting rod 3 and the needle-shaped object fastener 5 form an included angle and the angle is variable, and the needle-shaped object fastener 5 can rotate relative to the connecting rod 3 as required.
In another embodiment of the attaching method of the present invention, the needle-like object fastener 5 is attached to the elevation adjuster 4 through the connecting rod 3; one end of the connecting rod 3 is connected to a lifting regulator 4 on a microscope arm, and a lifting hand wheel 10 controls the lifting regulator 4 to ascend or descend; the other end of connecting rod 3 is connected with needle-like object fastener 5 through spheroid rotary joint, and connecting rod 3 forms contained angle and angle with needle-like object fastener 5 is variable, and as required, needle-like object fastener 5 can rotate for connecting rod 3.
In one embodiment of the connection method of the present invention, the connecting rod 3 of the monospore separating apparatus is fixed to the middle-lower portion of the elevation adjuster 4 by a screw.
In another embodiment of the coupling method of the present invention, the coupling rod 3 of the single spore isolation apparatus is fixed to the lower middle portion of the microscope lifting regulator 4 by welding.
The single spore isolation microscope of the present invention is a microscope equipped with the single spore isolation apparatus of the present invention. The single spore separating device can be arranged on a common optical microscope, and can be an upright microscope or an inverted microscope.
FIG. 2 is a schematic front view of a single spore isolation microscope according to an embodiment of the present invention. FIG. 3 is a schematic side view of a single spore isolation microscope according to one embodiment of the present invention.
The single spore isolation microscope comprises a microscope base 1, a microscope arm 2, a lens cone, an ocular, an objective lens, a lifting hand wheel 10 and an object stage 9, wherein the object stage 9 is provided with a longitudinal adjuster and a transverse adjuster, and the single spore isolation microscope is provided with the single spore isolation device.
In order to mount the single spore isolation apparatus of the present invention on a microscope, it is necessary to remove the condenser lens holder elevation adjuster, the condenser lens holder connected to the condenser lens holder elevation adjuster, and the condenser lens and the iris diaphragm connected to the condenser lens holder, and mount the elevation adjuster 4 at the position of the condenser lens holder elevation adjuster.
In one embodiment of the monospore separating microscope of the present invention, the monospore separating apparatus is fixed to the arm of the microscope at the position of the collector stand elevation adjuster, and the elevation adjuster 4 adjusts the elevation or the lowering in the same manner as the collector stand elevation adjuster.
In one embodiment of the monospore isolation microscope of the present invention, the elevation and the descent of the elevation adjuster 4 are adjusted by an elevating handwheel 10.
In one embodiment of the single spore isolation microscope of the present invention, one end of a connecting rod 3 of the single spore isolation apparatus is connected to a lifting adjuster 4, the other end of the connecting rod 3 is connected to a needle-like object fastening member 5, and the connecting rod 3 is positioned in a horizontal direction or a direction approximately horizontal.
In one embodiment of the single spore isolation microscope of the present invention, the connection of the connecting rod 3 and the needle-like object fastening member 5 is connected by the steering joint 11, and the needle-like object fastening member 8 can be rotated with respect to the connecting rod 3, so that the operable area for picking up single spores is large, and the single spore isolation operation is easy.
In another embodiment of the single spore isolation microscope of the present invention, the connection of the connecting rod 3 and the needle-like object fastener 5 is connected by a ball pivot joint, and the needle-like object fastener 8 can pivot in a three-dimensional space with respect to the connecting rod 3, so that the operable area for picking up single spores is further increased, and the single spore isolation operation is easier to perform.
One embodiment of the monospore isolation method of the present invention comprises the steps of:
(1) preparing a culture dish with a transparent culture medium laid at the bottom, and smearing conidia of the strain to be separated on the surface of the culture medium;
(2) the culture dish is fixed on a microscope stage 9 with the surface of the culture medium facing the separation needle 8;
(3) rotating a longitudinal regulator and/or a transverse regulator of the microscope objective table 9 to find a target monospore under a microscope visual field;
(4) controlling the lifting regulator 4 by using a lifting hand wheel 10 and/or regulating the needle-shaped object fastener 5 to enable the separation needle 8 to approach the target monospore;
(5) the lifting hand wheel 10 is used for controlling the lifting regulator 4 to enable the separation needle 8 to contact the culture medium;
(6) adjusting a longitudinal adjuster and/or a transverse adjuster of the microscope objective table 9 to control the objective table 9, and picking the spores onto the separation needle 8;
(7) the lifting hand wheel 10 is used for controlling the lifting regulator 4 to move the separation needle 8, so that the separation needle 8 is not contacted with the culture medium at all.
(8) Rotating the longitudinal regulator and/or the transverse regulator of the object stage 9 to move the spores to the vicinity of the clean culture medium;
(9) the lifting regulator 4 is controlled to move the separating needle 8 to place the spores on a clean culture medium.
Reference numerals:
1. a lens base;
2. a scope arm;
3. a connecting rod;
4. a lift adjuster;
5. a needle-like object fastener;
6. a hollow cone;
7. an internally threaded nut;
8. a separation needle;
9. an object stage;
10. a lifting hand wheel;
11. a steering joint;
12. an external thread insert.
Claims (14)
1. A single spore separation device comprises a lifting adjuster, a connecting rod, a needle-shaped object fastener and a separation needle;
the lifting regulator is arranged at the position of the condensing lens lifting regulator on the microscope arm, and the lifting regulator is controlled to ascend or descend by utilizing a lifting hand wheel of the microscope;
one end of the connecting rod is connected to the lifting adjuster, and the other end of the connecting rod is connected to the needle-shaped object fastener;
one end of the separation needle is locked inside the needle-shaped object fastener;
the needle-shaped object fastener is provided with an external thread turnbuckle and an internal thread nut at the end part;
the body of the external thread turnbuckle is a hollow cylinder;
the body of the internal thread nut is a hollow cylinder, the hollow cylinder is provided with internal threads, the end part of the internal thread nut is a hollow cone, and the end part of the hollow cone is horizontally cut off;
the threads of the internal thread nut and the external thread sleeve are mutually meshed;
the needle-shaped object fastener is also provided with a hollow cone, the hollow cone is provided with a Y-shaped slot or a cross slot, and the end part of the hollow cone is horizontally cut off;
the hollow cone is an independent part and is positioned on the inner side of the internal thread nut;
the hollow cone is made of elastic material 65 manganese steel;
the needle-like object is locked within the hollow cone.
2. The monospore separation apparatus according to claim 1, wherein: the separating needle is made of metal, alloy, polymer fiber or glass fiber.
3. The monospore separation apparatus according to claim 1, wherein: the ends of the separating needles are straight or bent.
4. The monospore separation apparatus according to claim 1, wherein: the tip of the separation needle has a sharp shape.
5. The monospore separation apparatus according to claim 2, wherein: the ends of the separating needles are straight or bent.
6. The monospore separation apparatus according to claim 2, wherein: the tip of the separation needle has a sharp shape.
7. A monospore separation apparatus according to claim 3, wherein: the tip of the separation needle has a sharp shape.
8. The monospore isolation apparatus according to claim 5, wherein: the tip of the separation needle has a sharp shape.
9. The monospore separation apparatus according to any one of claims 1 to 8, wherein: the connecting rod is connected with the needle-shaped object fastener through a steering joint.
10. The monospore separation apparatus according to any one of claims 1 to 8, wherein: the connecting rod is connected with the needle-shaped object fastener through a sphere rotating joint.
11. The utility model provides an optical microscope, includes microscope base, microscope arm, lens cone, eyepiece, objective, lifting hand wheel and objective table, has vertical regulator and horizontal regulator on the objective table, its characterized in that: a single spore isolation apparatus according to any one of claims 1 to 10 mounted on the arm of the optical microscope.
12. Use of a monospore separation apparatus according to any one of claims 1 to 10 in an optical microscope.
13. The method for connecting a monospore separation apparatus according to any one of claims 1 to 10, wherein: one end of the connecting rod is connected with the needle-shaped object fastener; the other end of the connecting rod is connected to the lifting adjuster on the microscope arm, and the lifting adjuster is controlled to ascend or descend by a lifting hand wheel of a microscope; one end of the separation needle is locked inside the needle-shaped object fastener.
14. The method of connecting a single spore isolation apparatus according to claim 13, wherein: one end of the connecting rod is connected with the needle-shaped object fastener through a steering joint or a ball rotating joint.
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| EP0714730A2 (en) * | 1994-12-02 | 1996-06-05 | Makita Corporation | Blade mounting device in cutting tool |
| CN202322845U (en) * | 2011-12-03 | 2012-07-11 | 云南农业大学 | Simple and rapid quantitative microbe sampler |
| CN105255705A (en) * | 2015-10-27 | 2016-01-20 | 乔建国 | Fungus single-spore picker and method of separating and purifying plant pathogenic fungi through fungus single-spore picker |
| CN209452943U (en) * | 2019-01-24 | 2019-10-01 | 淮阴师范学院 | A guide screw frame for circular tube parts |
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| EP0714730A2 (en) * | 1994-12-02 | 1996-06-05 | Makita Corporation | Blade mounting device in cutting tool |
| CN202322845U (en) * | 2011-12-03 | 2012-07-11 | 云南农业大学 | Simple and rapid quantitative microbe sampler |
| CN105255705A (en) * | 2015-10-27 | 2016-01-20 | 乔建国 | Fungus single-spore picker and method of separating and purifying plant pathogenic fungi through fungus single-spore picker |
| CN209452943U (en) * | 2019-01-24 | 2019-10-01 | 淮阴师范学院 | A guide screw frame for circular tube parts |
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