CN110133166B - Device and method for simulating continuous insect invasion - Google Patents
Device and method for simulating continuous insect invasion Download PDFInfo
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- CN110133166B CN110133166B CN201910241569.XA CN201910241569A CN110133166B CN 110133166 B CN110133166 B CN 110133166B CN 201910241569 A CN201910241569 A CN 201910241569A CN 110133166 B CN110133166 B CN 110133166B
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New breeds of animals
- A01K67/033—Rearing or breeding invertebrates; New breeds of invertebrates
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
- C12Q1/6895—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for plants, fungi or algae
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/62—Detectors specially adapted therefor
- G01N30/72—Mass spectrometers
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/158—Expression markers
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N2030/022—Column chromatography characterised by the kind of separation mechanism
- G01N2030/027—Liquid chromatography
Abstract
The invention discloses a device and a method for simulating continuous insect infringement. The device comprises a door-shaped support, a needle part, a controller and a first stepping motor used for driving the needle part to move longitudinally, wherein the needle part is fixed at the output end of the first stepping motor, the first stepping motor is installed on the door-shaped support, and the controller is electrically connected with the first stepping motor. The invention can make the needle part penetrate into the blade at a fixed frequency, and the controller sends an instruction to regulate and control the first stepping motor, so as to better select and control the parameters of the penetrating frequency, time and the like, thereby truly simulating the invasion action of the insect on the blade. The invention avoids the error caused by manual operation in the experimental process, better simulates the feeding habit of insects and increases the reality of simulation.
Description
Technical Field
The invention relates to a device in the field of simulation experiments, in particular to a device for simulating continuous insect invasion, and further comprises a method for simulating continuous insect invasion.
Background
The HIPVs (plant volatile substances) generated by the insect invasion induced plant have important ecological functions and have important significance for the research of the HIPVs. When the HIPVs generated by the interaction of insects and plants are researched, the mechanical injury is usually caused to the plants by adopting a mechanical simulation method, so that the research is carried out. In most of the experimental studies at present, only a single mechanical injury is usually used to simulate the damage of insects, for example, a device for simulating insects to eat and pierce plants disclosed in chinese utility model CN201820922719.4, which only pierces the fixed position of the leaf blade by piercing, and the damage time and degree are greatly different from the damage caused by eating by insects, so that it is difficult to truly simulate the specific and fixed damage mode of phytophagous insects to eat, and the requirement of theoretical research cannot be well satisfied.
Disclosure of Invention
The invention aims to provide a device for simulating continuous insect invasion, which can cause continuous invasion to blades.
Also included is a method of simulating continuous infestation of an insect which better simulates infestation by feeding foliage of the insect.
The solution of the invention for solving the technical problem is as follows: the device comprises a door-shaped support, a needle part, a controller and a first stepping motor for driving the needle part to move longitudinally, wherein the needle part is fixed at the output end of the first stepping motor, the first stepping motor is installed on the door-shaped support, and the controller is electrically connected with the first stepping motor.
As a further improvement of the technical scheme, the first stepping motor is installed on the door-shaped support through a longitudinal adjusting mechanism, the longitudinal adjusting mechanism comprises a screw rod, a moving member matched with the screw rod and two auxiliary clamping plates, the two auxiliary clamping plates are respectively and oppositely fixed at the top and the bottom of the cross rod of the door-shaped support, two ends of the screw rod are connected with the two auxiliary clamping plates, and the first stepping motor is installed on the moving member.
As a further improvement of the technical scheme, the clamping device further comprises at least two connecting rods used for connecting the two auxiliary clamping plates, the connecting rods are respectively positioned on two sides of the cross rod of the door-shaped support, the moving member is provided with a vertical through hole, and the connecting rods penetrate through the vertical through hole.
As a further improvement of the technical scheme, a rolling belt and two rollers are arranged on a cross rod of the door-shaped support, the auxiliary clamping plate is fixedly connected with the outer side of the rolling belt, the rolling belt is installed on the two rollers, the rollers are driven by a second stepping motor, and the controller is electrically connected with the second stepping motor through a coordinate encoder.
As a further improvement of the technical scheme, the door-shaped support is characterized by further comprising a base, and the door-shaped support is installed on the base through a moving mechanism.
As a further improvement of the above technical solution, the moving mechanism includes two belts respectively disposed on two sides of the base and a third stepping motor for driving the belts to rotate, the two supports of the door-shaped support are respectively fixed on the two belts, the moving direction of the door-shaped support is perpendicular to the cross bar of the door-shaped support, and the controller is electrically connected to the third stepping motor through a coordinate encoder.
A method for simulating continuous damage of insect includes such steps as putting the blade under the needle, fixing, moving the needle up and down by controller, and penetrating the blade by the needle at fixed frequency.
As a further improvement of the technical proposal, the needle part reciprocates along a linear track in the horizontal direction, and the needle part pierces the blade once when moving 1mm horizontally.
As a further improvement of the technical proposal, the needle part reciprocates along a rectangular track in the horizontal direction, and the needle part pierces the blade once when moving 1mm horizontally.
As a further improvement of the technical scheme, after the needle part penetrates into the blade for 10-30 times, the needle part stops moving for 30-90 s and then continues moving.
The invention has the beneficial effects that: the invention can make the needle part penetrate into the blade at a fixed frequency, and the controller sends an instruction to regulate and control the first stepping motor, so as to better select and control the parameters of the penetrating frequency, time and the like, thereby truly simulating the invasion action of the insect on the blade. The invention avoids the error caused by manual operation in the experimental process, better simulates the feeding habit of insects and increases the reality of simulation.
Drawings
In order to more clearly illustrate the technical solution in the embodiments of the present invention, the drawings used in the description of the embodiments will be briefly described below. It is clear that the described figures are only some embodiments of the invention, not all embodiments, and that a person skilled in the art can also derive other designs and figures from them without inventive effort.
FIG. 1 is a schematic view of the overall structure of the present invention;
FIG. 2 is a schematic view of a longitudinal adjustment mechanism of the present invention;
FIG. 3 is a schematic view of a prior art treated blade;
FIG. 4 is a schematic view of a blade after treatment according to a third embodiment of the invention;
FIG. 5 is a graph comparing the amounts of linalool, synthetic gene one, and synthetic gene two in the control group and the cicada infestation group;
FIG. 6 is a graph comparing the amounts of linalool, synthetic gene one, and synthetic gene two in the control group, single lesion group, and consecutive lesion group;
fig. 7 is a graph comparing the geraniol content in the control group, the single lesion group, the cicada-affected group, and the continuous lesion group.
Detailed Description
The conception, the specific structure, and the technical effects produced by the present invention will be clearly and completely described below in conjunction with the embodiments and the accompanying drawings to fully understand the objects, the features, and the effects of the present invention. It is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments, and those skilled in the art can obtain other embodiments without inventive effort based on the embodiments of the present invention, and all embodiments are within the protection scope of the present invention. In addition, all the coupling/connection relationships mentioned herein do not mean that the components are directly connected, but mean that a better coupling structure can be formed by adding or reducing coupling accessories according to specific implementation conditions.
Referring to fig. 1 to 2, the device for simulating continuous insect infestation comprises a door-shaped support 2, a needle part 1, a controller and a first stepping motor 3 for driving the needle part 1 to move longitudinally, wherein the needle part 1 is fixed at the output end of the first stepping motor 3, the first stepping motor 3 is installed on the door-shaped support 2, and the controller is electrically connected with the first stepping motor 3. The controller comprises a high-frequency conversion device, a lubricator, a cooling device, an encoder and the like. After setting the puncturing frequency and the movement time of the needle part 1 on the controller, the controller sends a command to the first stepping motor 3 to move the needle part 1 longitudinally at a fixed frequency to perform puncturing operation on the blade. The needle portion 1 repeatedly penetrates a fixed position of the blade. In addition, the output end of the first stepping motor 3 is provided with an external thread, and the needle part 1 is connected with the external thread through a connecting part with an internal thread, so that the needle part with different diameters can be replaced to better simulate the invasion of insects, and the diameter of the needle part 1 is preferably 0.5 mm.
Further as a preferred embodiment, the first stepping motor 3 is installed on the door-shaped support 2 through a longitudinal adjusting mechanism, the longitudinal adjusting mechanism includes a screw rod 51, a moving member 52 matched with the screw rod 51, and two auxiliary clamping plates 53, the two auxiliary clamping plates 53 are respectively fixed on the top and the bottom of the cross bar of the door-shaped support 2 relatively, two ends of the screw rod 51 are connected with the two auxiliary clamping plates 53, and the first stepping motor 3 is installed on the moving member 52. In order to adjust the distance between the needle part 1 and the blade to achieve a better penetration effect, the vertical height of the needle part 1 can be adjusted by a longitudinal adjusting mechanism. When the screw rod 51 is rotated, the moving block 52 moves in the longitudinal direction and drives the first stepping motor 3 to move, thereby adjusting the height of the needle portion 1. The screw rod 51 can be driven by manual or power device, preferably, a fourth stepping motor is connected with one end of the screw rod 51, the fourth stepping motor is electrically connected with the controller, and the user can control the height of the first stepping motor 3 by debugging on the controller, thereby adjusting the relative position of the needle part 1 and the blade.
Further as a preferred embodiment, the device further comprises at least two connecting rods 54 for connecting the two auxiliary clamping plates 53, the connecting rods 54 are respectively located at two sides of the cross bar of the door-shaped support 2, the moving member 52 is provided with a vertical through hole, and the connecting rods 54 penetrate through the vertical through hole. Since the connecting rod 54 penetrates the vertical through hole, the moving member 52 is restricted in freedom by the connecting rod 54 to move only in the longitudinal direction.
Further as a preferred embodiment, a rolling belt and two rollers are arranged on the cross bar of the door-shaped support 2, the auxiliary clamping plate 53 is fixedly connected with the outer side of the rolling belt, the rolling belt is installed on the two rollers, the rollers are driven by the second stepping motor 4, and the controller is electrically connected with the second stepping motor 4 through a coordinate encoder. The second stepping motor 4 drives the roller to rotate, the rolling belt rolls along with the roller, and the longitudinal adjusting mechanism fixed on the rolling belt is driven to move transversely, so that the needle part 1 moves transversely.
Further, as a preferable embodiment, the door-shaped bracket 2 further comprises a base, and the door-shaped bracket is installed on the base through a moving mechanism. A base is provided for the blade to be placed on a more even surface for piercing.
As a preferred embodiment, the moving mechanism includes two belts respectively disposed on two sides of the base and a third stepping motor for driving the belts to rotate, the two supports of the door-shaped support 2 are respectively fixed on the two belts, the moving direction of the door-shaped support 2 is perpendicular to the cross bar of the door-shaped support 2, and the controller is electrically connected to the third stepping motor through a coordinate encoder. And a third step motor drives the belt to rotate under the instruction of the controller, so that the whole door-shaped bracket 2 is driven to move back and forth, and the needle part 1 moves back and forth. By inputting a simple assembler to the controller and the coordinate encoder, the needle 1 can be moved to the target coordinate along a predetermined trajectory.
A method for simulating continuous infestation by insects is described in detail below with reference to three preferred embodiments which simulate infestation of tea plant leaves by lesser leafhoppers.
The first embodiment is as follows:
the blade is placed under the needle part 1, and the position of the blade is fixed by means of transparent adhesive or the like. The longitudinal movement pattern of the needle portion 1 is set by the controller as follows: the needle part 1 is set to penetrate into the blade at the frequency of 20 times/min, the needle part 1 stops pausing for 1min every time the needle part continuously penetrates into the blade for 1min, then the previous penetrating operation is repeated, and the like, and the whole process lasts for 12 h. In addition, in order to better simulate the invasion habits of insects with different habits, the puncturing frequency of the needle part 1 can be selected to be 10 times/min or 30 times/min, the pause time between two operations can be selected to be 30s or 90s, and the duration time of the whole process is selected to be 2 h-24 h.
Example two:
in the first embodiment, the needle part 1 is reciprocated along a linear trajectory in the horizontal direction while being moved longitudinally to pierce the blade. The longitudinal movement mode of the needle part 1 is the same as that of the first embodiment, and the movement steps of the needle part 1 in the horizontal direction are as follows:
a. the needle part 1 moves 1mm to the left after each penetration;
b. when the needle part 1 moves leftwards for 20mm, the needle part 1 changes the direction of horizontal movement, and moves rightwards for 1mm every time when the needle part is penetrated;
c. when the needle part 1 returns to the original position, the steps a and b are repeated in sequence, and the whole process lasts for 12 h.
Example three:
on the basis of the first embodiment, the needle part 1 is reciprocated along a rectangular trajectory in the horizontal direction while being moved longitudinally to pierce the blade. The longitudinal movement mode of the needle part 1 is the same as that of the first embodiment, and the movement steps of the needle part 1 in the horizontal direction are as follows:
a. the needle part 1 moves 1mm to the left after each penetration;
b. when the needle part 1 moves leftwards for 20mm, the needle part 1 changes the direction of horizontal movement and moves forwards for 1mm every time the needle part is penetrated;
c. when the forward movement of the needle part 1 is accumulated to 20mm, the needle part 1 changes the direction of horizontal movement and moves 1mm to the left once it is pierced;
d. when the needle part 1 moves leftwards for 20mm, the needle part 1 changes the direction of horizontal movement and moves backwards for 1mm every time the needle part is penetrated;
e. when the needle part 1 returns to the original position, the steps a to d are repeated in order, and the whole process lasts for 12 hours.
The invasive effect of the invention on the blade is shown from two angles:
firstly, phenotypic change:
in the above examples, particularly after the treatments of the second and third examples, referring to fig. 4, the insect infestation damage present on the leaf as a sample is different from the insect infestation damage simulated by the prior art. Specifically, the samples in the first to third examples were continuous irregular-shaped lesions, and the lesions had both a portion penetrating the leaf completely and a portion piercing only the leaf, thus more truly simulating the leaf lesions after being attacked by insects; whereas with the prior art, with reference to fig. 3, only a single circular lesion hole may be made to the blade.
Secondly, detecting volatile substances and gene expression thereof:
groups of consecutive lesions: 15 tea leaves treated in example three;
control group: 15 leaves of tea tree which had not been subjected to any treatment;
single lesion group: 15 tea leaves which are directly punctured by a needle to cause single injury;
cicada invasion group: 15 healthy single-bud trefoil leaves were infested with 30 lesser leafhoppers for 3 days.
The volatile substances are determined and analyzed by a gas chromatography-mass spectrometer (GC-MS). The relative amount of linalool in the sample is the ratio of the area of the peak of linalool (m/z 71) to the area of the peak of the ethyl decanoate of the internal standard (m/z 88). The relative amount of geraniol in a sample is the ratio of the peak area of geraniol (m/z 69) to the peak area of the internal standard ethyl decanoate (m/z 88).
The total RNA of tea leaves is rapidly extracted by a Quick RNA isolation kit (Huayueyang biotechnology (Beijing) Co., Ltd., Beijing, China). Using iTaqTM Universal
Green Supermix (BioRad, Herrax, USA) was reacted in a 20. mu.L system containing 10. mu.L of iTaqTM
Green Supermix (2X), 0.4. mu.M forward and reverse primers. The template was diluted 20-fold and 4. mu.L was added to 20. mu.L of PCR reaction. Real-time quantitative PCR (qRT-PCR) was performed by Roche LightCycle 480(Roche Applied Science, Mannheim, Germany) at 95 ℃ for 30s for one cycle, 95 ℃ for 5s for 40 cycles, and 60 ℃ for 30 s. The specificity of the PCR product was verified at the end of the reaction according to the melting curve. Beta-actin was used as an internal control and the change in mRNA levels for each treatment was normalized to beta-actin.
Referring to fig. 5 to 7, it can be seen that: the content of linalool and geraniol in leaves in the continuous injury group is obviously increased, and the expression of related synthetic gene I and synthetic gene II is also obviously increased, which is consistent with that after the tea leaves are invaded by lesser leafhoppers in a natural state. The insect invasion is simulated by using a single mechanical injury in the prior art, namely the rising degree of linalool and geraniol of the leaves of a single injury group is not so obvious, the rising curve is relatively gentle, and the expression of a synthetic gene I and a synthetic gene II is greatly different from that of a cicada invasion group.
While the preferred embodiments of the present invention have been illustrated and described, it will be understood by those skilled in the art that the present invention is not limited to the details of the embodiments shown and described, but is capable of numerous equivalents and substitutions without departing from the spirit of the invention as set forth in the claims appended hereto.
Claims (9)
1. A device for simulating continuous insect infestation, comprising: comprises a door-shaped bracket (2), a needle part (1), a controller and a first stepping motor (3) for driving the needle part (1) to move longitudinally, the needle part (1) is fixed at the output end of a first stepping motor (3), the first stepping motor (3) is arranged on a door-shaped bracket (2), the controller is electrically connected with a first stepping motor (3), the first stepping motor (3) is arranged on the door-shaped bracket (2) through a longitudinal adjusting mechanism, the longitudinal adjusting mechanism comprises a screw rod (51), a moving piece (52) matched with the screw rod (51) and two auxiliary splints (53), the two auxiliary splints (53) are respectively and relatively fixed at the top and the bottom of the cross bar of the door-shaped bracket (2), two ends of the screw rod (51) are connected with two auxiliary clamping plates (53), and the first stepping motor (3) is arranged on the moving part (52).
2. The device for simulating continuous infestation of insects of claim 1, wherein: the clamping device is characterized by further comprising at least two connecting rods (54) used for connecting the two auxiliary clamping plates (53), wherein the connecting rods (54) are respectively located on two sides of the cross rod of the door-shaped support (2), vertical through holes are formed in the moving member (52), and the connecting rods (54) penetrate through the vertical through holes.
3. The device for simulating continuous infestation of insects of claim 2, wherein: the rolling belt and the two rollers are arranged on the cross rod of the door-shaped support (2), the auxiliary clamping plate (53) is fixedly connected with the outer side of the rolling belt, the rolling belt is installed on the two rollers, the rollers are driven by the second stepping motor (4), and the controller is electrically connected with the second stepping motor (4) through a coordinate encoder.
4. The device for simulating continuous infestation of insects of claim 1, wherein: the door-shaped support is characterized by further comprising a base, and the door-shaped support (2) is installed on the base through a moving mechanism.
5. The device of claim 4, wherein the device further comprises: the moving mechanism comprises two belts arranged on two sides of the base respectively and a third stepping motor used for driving the belts to rotate, the two supports of the door-shaped support (2) are fixed on the two belts respectively, the moving direction of the door-shaped support (2) is perpendicular to the cross rod of the door-shaped support (2), and the controller is electrically connected with the third stepping motor through a coordinate encoder.
6. A method of simulating continuous infestation of an insect, comprising: a device for simulating continuous insect infestation according to any of claims 1 to 5 wherein the blade is placed under the needle (1) and fixed, the first stepping motor (3) drives the needle (1) up and down and the needle (1) penetrates the blade at a fixed frequency.
7. The method of simulating continuous infestation of insects of claim 6, wherein: the needle part (1) reciprocates along a linear track in the horizontal direction, and the needle part (1) penetrates the blade once every 1mm of horizontal movement.
8. The method of simulating continuous infestation of insects of claim 6, wherein: the needle part (1) reciprocates along a rectangular track in the horizontal direction, and the needle part (1) penetrates the blade once every 1mm of horizontal movement.
9. A method of simulating continuous infestation by an insect according to any one of claims 6 to 8 wherein: after the needle part (1) penetrates into the blade for 10-30 times, the needle part (1) stops moving for 30-90 s and continues moving.
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