Pear genotype detection kit and detection method

Abstract

The invention relates to the field of gene detection, and discloses a pear genotype detection kit and a detection method.The pear genotype detection kit comprises a kit body and a kit cover hinged to the kit body, multiple rows of folding frames arranged in parallel are installed in the kit body, and multiple PCR tubes are installed on each row of folding frames; grinding and extracting bins in one-to-one correspondence with the folding frames are arranged in the box body on one side of the folding frames, a plurality of net bags are arranged on the box cover, and quantitative suction pipes and injectors are placed in the net bags.

Description

Technical Field

[0001] The invention relates to the field of gene detection, and in particular to a kit and a method for detecting pear genotypes. Background Art

[0003] The existing methods for variety identification include traditional morphological characteristics identification methods and protein and isozyme electrophoresis pattern technology. However, these identification methods are greatly affected by the environment, time-consuming and labor-intensive to operate, not only inefficient, but also have certain errors. DNA molecular marker technology is a reduction technology that is not affected by environmental conditions and can be used for rapid, effective and accurate identification. More and more identifications are beginning to use DNA molecular marker technology for identification.

[0004] Although DNA molecular marker technology can be used for rapid and accurate identification, it is costly and requires a high level of professional expertise from the operator. In addition, DNA molecular markers are extremely sensitive, so the entire experimental conditions need to be strictly controlled. Therefore, multiple repeated experiments are generally performed to ensure the accuracy of the results. Summary of the invention

[0005] The purpose of the present invention is to solve the problems in the background technology and provide a pear genotype detection kit and a detection method. The detection kit can provide the environment required by DNA molecular marker technology. At the same time, the entire operation can be completed by just following the steps, which greatly improves the convenience of DNA molecular marker detection.

[0006] The objective of the present invention is achieved through the following technical solutions:

[0007] A pear genotype detection kit comprises a box body and a box cover hinged on the box body, wherein a plurality of rows of parallel folding racks are installed in the box body, a plurality of PCR tubes are installed on each row of the folding racks, a grinding and extraction bin corresponding to the folding racks is arranged in the box body on one side of the folding racks, a plurality of net bags are arranged on the box cover, and quantitative pipettes and syringes are placed in the net bags.

[0008] The folding frame includes a crossbeam and a support rod hinged at both ends of the crossbeam, a support seat fixedly connected to the box body is provided at the bottom of the support rod, a strip hole hinged to the support seat is processed at one end of the support rod, and a limiting groove adapted to the contour of the support rod is processed on the surface of the support seat. A plurality of rotatable clamping rings are installed on the crossbeam, and the PCR tube is conical and installed in the clamping ring.

[0009] The PCR tube is pre-installed with polymerase, a tin foil sealing sheet is installed on the top of the PCR tube, a closable cap is connected to the top side wall of the PCR tube, and a dropper is processed on the cap.

[0010] The grinding and extraction chamber includes a culture chamber, a support cover and a movable cover, wherein the support cover is installed on the culture chamber, and a slot is processed on the culture chamber to support and limit the support cover, a grinding slot is processed at the center of the surface of the support cover, and the movable cover is rotatably installed on the surface of the support cover, a grinding rod is processed on the movable cover to extend into the grinding slot, a drainage tube is processed at the bottom of the grinding slot to extend into the culture chamber, and a plurality of reinforcing ribs are processed on the bottom surface of the support cover outside the drainage tube, and one of the reinforcing ribs extends downward and engages with the slot.

[0011] The surfaces of the grinding rod and the grinding groove are both processed with grinding teeth.

[0012] A filter cavity is processed on the side wall of the culture bin, and filter cotton is installed on the side wall of the filter cavity.

[0013] A method for detecting pear genotypes, comprising the following steps:

[0014] (1) Sample collection: Open the box cover, take out the grinding and extraction chamber, then take fresh leaves or fruit tissues and place them in the grinding groove on the support cover, then cover the movable cover and rotate the movable cover so that the grinding rod on the movable cover cooperates with the grinding groove to assemble and grind the fresh leaves or fruits into a paste;

[0015] (2) DNA extraction: Take out the syringe from the net bag, draw up the slow-release solution, and push it into the grinding groove of the support cover. During this process, the movable cover is continuously turned to allow the sample tissue to flow into the culture chamber along the drainage tube. The culture chamber is then placed in an environment of 0 to 5°C for 30 minutes to wait for the DNA in the sample tissue to be released.

[0016] (3) Gene amplification: Based on step (2), the culture chamber is deflected to make the slow-release liquid gather toward the filter cavity. The deflected state is maintained to allow the slow-release liquid to pass through the filter cotton and be placed in the filter cavity. At this time, the folding frame is unfolded, and the tin foil sealing sheet on the corresponding PCR tube is torn off. Then, the quantitative pipette is taken out and the DNA in the filter cavity is dripped into the PCR tube in turn. Then, the cap is covered and the PCR tube is placed in the PCR instrument for circulation, and the number of DNA fragments is waited to increase;

[0017] (4) Product detection: Turn the PCR tube over, separate the product in the PCR tube by electrophoresis on agarose gel, stain with nucleic acid dye, and observe the results using a gel imaging system;

[0018] (5) Data analysis: The pear gene can be identified based on the electrophoresis pattern and the results of the previous analysis.

[0019] The sustained-release solution is prepared by uniformly mixing 1-3 ml of sodium alkyl sulfonate, 1-2 ml of sodium fatty alcohol ether sulfate, 1-2 g of sodium chloride, 50 ml of pure water and 3-5 ml of alcohol in parts by weight.

[0020] The pear genotype detection kit and detection method provided by the present invention have the following beneficial effects:

[0021] (1) The DNA data of pears can be obtained quickly and accurately, and the pear varieties can be quickly identified. At the same time, the identification process is simple to operate, which reduces the technical requirements for operators;

[0022] (2) The kit enables fresh pear tissue to be collected on site, thereby improving the accuracy of detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for use in the embodiments are briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without creative work.

[0024] Figure 1 A schematic diagram of the structure of a kit provided in an embodiment of the present invention.

[0025] Figure 2 A schematic structural diagram of a folding frame provided in an embodiment of the present invention.

[0026] Figure 3 A schematic diagram of the structure of a PCR tube provided in an embodiment of the present invention.

[0027] Figure 4 A schematic diagram of the structure of a grinding and extraction chamber provided in an embodiment of the present invention.

[0028] Figure numerals: 1. box body; 2. box cover; 3. folding frame; 31. support rod; 32. support seat; 33. strip hole; 34. limit groove; 35. clamping ring; 36. crossbeam; 4. PCR tube; 41. cap; 42. tin foil sealing sheet; 44. dropper; 5. grinding and extraction chamber; 51. culture chamber; 52. support cover; 53. movable cover; 54. clamping groove; 55. grinding groove; 56. grinding rod; 57. drainage tube; 58. reinforcing rib; 59. filter chamber; 510. filter cotton; 6. net bag; 61. quantitative pipette; 62. syringe. DETAILED DESCRIPTION

[0029] Example

[0030] like Figure 1 to Figure 4As shown, the pear genotype detection kit provided in this embodiment includes a box body 1 and a box cover 2 hinged on the box body 1, wherein the box body 1 is provided with multiple rows of parallel folding frames 3, which can be folded up when the box body 1 is covered to reduce the floor space, and can be unfolded when the pear gene needs to be extracted to facilitate the operation of the staff, and multiple PCR tubes 4 are installed on each row of the folding frames 3, and the multiple PCR tubes 4 can be adjusted forward and reverse at the same time, so that multiple data can be obtained at the same time, and the box body 1 on one side of the folding frame 3 is provided with a grinding and extraction chamber 5 corresponding to the folding frame 3 one by one, and the grinding and extraction chamber 5 can conveniently crush and extract the pear tissue, and can greatly improve the crushing efficiency compared with the existing complex extraction process, and the box cover 2 is provided with multiple net bags 6, and the net bags 6 are used to store the required consumables, and quantitative pipettes 61 and syringes 62 are placed in the net bags 6, and the quantitative pipettes 61 and syringes 62 can conveniently transfer liquids without the use of pipettes.

[0031] In order to facilitate gene amplification operations, Figure 2 , Figure 3 As shown, the folding frame 3 includes a crossbeam 36 and a support rod 31 hinged at both ends of the crossbeam 36. The bottom of the support rod 31 is provided with a support seat 32 fixedly connected to the box body 1. One end of the support rod 31 is processed with a strip hole 33 hinged to the support seat 32. The surface of the support seat 32 is processed with a limiting groove 34 adapted to the contour of the support rod 31. Through the strip hole 33, the support rod 31 can be inserted into the limiting groove 34 after the folding frame 3 is unfolded, thereby ensuring the unfolded state of the folding frame 3 and effectively improving the stability. A plurality of rotatable clamping rings 35 are installed on the crossbeam 36. The PCR tube 4 is conical The PCR tube 4 is installed in the clamp ring 35. In order to simplify the gene amplification operation, the PCR tube 4 is pre-installed with polymerase. A tin foil sealing sheet 42 is installed on the top of the PCR tube 4. When in use, the tin foil sealing sheet 42 on the PCR tube 4 is directly torn off to directly perform the liquid transfer operation without the need to configure the polymerase. The top side wall of the PCR tube 4 is connected with a coverable cap 41. The cap 41 can prevent the polymerase from spilling out after the PCR tube 4 is placed in the PCR machine. A dropper 44 is processed on the cap 41. Under the action of the dropper 44, the polymerase can be conveniently and accurately dropped into the designated place.

[0032] In order to conveniently extract DNA from pear tissues, Figure 4As shown, the grinding and extraction chamber 5 includes a culture chamber 51, a support cover 52 and a movable cover 53. The support cover 52 is installed on the culture chamber 51. The culture chamber 51 is processed with a card slot 54 for supporting and limiting the support cover 52. The center of the surface of the support cover 52 is processed with a grinding groove 55. The movable cover 53 is rotatably installed on the surface of the support cover 52. The movable cover 53 is processed with a grinding rod 56 extending into the grinding groove 55. In order to improve the grinding efficiency, the surfaces of the grinding rod 56 and the grinding groove 55 are processed with grinding teeth. The bottom of the grinding groove 55 is processed with a grinding rod 56 extending into the culture chamber 51. A drainage tube 57 is provided, and the drainage tube 57 can introduce the ground juice into the culture chamber 51. A plurality of reinforcing ribs 58 are processed on the bottom surface of the support cover 52 outside the drainage tube 57, and one of the reinforcing ribs 58 extends downward and engages with the card slot 54. The reinforcing ribs 58 can improve the stability and structural strength of the support cover 52. A filter cavity 59 is processed on the side wall of the culture chamber 51, and a filter cotton 510 is installed on the side wall of the filter cavity 59. Under the action of the filter cotton 510, the tissue fluid flowing into the filter cavity 59 can filter out the solid tissue, which can prevent the tissue from clogging the channel during the subsequent extraction process.

[0033] A method for detecting pear genotypes, comprising the following steps:

[0034] (1) Sample collection: Open the box cover 2, take out the grinding and extraction chamber 5, then take fresh leaves or fruit tissues and place them in the grinding groove 55 on the support cover 52, then cover the movable cover 53, and rotate the movable cover 53 so that the grinding rod 56 on the movable cover 53 cooperates with the grinding groove 55, and the fresh leaves or fruits are assembled and ground into a paste;

[0035] (2) DNA extraction: Take out the syringe 62 from the net bag 6, and draw a slow-release solution. The slow-release solution is uniformly mixed with 1-3 ml of sodium alkyl sulfonate, 1-2 ml of sodium fatty alcohol ether sulfate, 1-2 g of sodium chloride, 50 ml of pure water, and 3-5 ml of alcohol by weight. The sodium alkyl sulfonate and sodium fatty alcohol ether sulfate components in the slow-release solution can fuse with the phospholipids in the tissue, thereby causing the DNA of the tissue to seep into the slow-release solution. At the same time, under the action of sodium chloride, the dissolution of DNA can be accelerated and impurities can be precipitated. The taken-out slow-release solution is pushed into the grinding groove 55 of the support cover 52. During this process, the movable cover 53 is continuously rotated. The slow-release solution can not only increase the contact area between the slow-release solution and the pear tissue, but also enhance the fluidity of the pear tissue, so that the sample tissue flows into the culture chamber 51 along the drainage tube 57, and the culture chamber 51 is placed in an environment of 0-5°C for 30 minutes to wait for the DNA in the sample tissue to be released;

[0036] (3) Gene amplification: Based on step (2), the culture chamber 51 is deflected to make the slow-release liquid gather toward the filter chamber 59, and the deflected state is maintained so that the slow-release liquid passes through the filter cotton 510 and is placed in the filter chamber 59. At this time, the folding frame 3 is unfolded, and the tin foil sealing sheet 42 on the corresponding PCR tube 4 is torn off, and then the quantitative pipette 61 is taken out, and the DNA in the filter chamber 59 is dripped into the PCR tube 4 in sequence, and then the cap 41 is covered, and the PCR tube 4 is placed in the PCR instrument for circulation, waiting for the number of DNA fragments to increase, and using the PCR instrument to increase the DNA fragments belongs to the prior art, so it is not described here in detail;

[0037] (4) Product detection: Turn over PCR tube 4, separate the product in PCR tube 4 by electrophoresis on agarose gel, stain with nucleic acid dye, and observe the result by gel imaging system;

[0038] (5) Data analysis: The pear gene can be identified based on the electrophoresis pattern and the results of the previous analysis.

[0039] The above is only a preferred embodiment of the present invention, but the protection scope of the present invention is not limited thereto, and any modification and replacement based on the technical solution and inventive concept provided by the present invention shall be included in the protection scope of the present invention. It should be noted that the structures or components illustrated in the drawings are not necessarily drawn to scale, and the present invention omits the description of known components and processing technologies and processes to avoid unnecessary limitation of the present invention.

Claims

1. A kit for detecting pear genotypes, characterized in that: The invention comprises a box body (1) and a box cover (2) hinged on the box body (1); a plurality of rows of folding racks (3) arranged in parallel are installed in the box body (1); a plurality of PCR tubes (4) are installed on each row of the folding racks (3); a grinding and extraction chamber (5) corresponding to the folding racks (3) is arranged in the box body (1) on one side of the folding racks (3); a plurality of net bags (6) are arranged on the box cover (2); quantitative pipettes (61) and syringes (62) are placed in the net bags (6).

2. The pear genotype detection kit according to claim 1, characterized in that: The folding frame (3) comprises a crossbeam (36) and a support rod (31) hinged at both ends of the crossbeam (36); a support seat (32) fixedly connected to the box body (1) is provided at the bottom of the support rod (31); a strip hole (33) hinged to the support seat (32) is processed at one end of the support rod (31); a limiting groove (34) adapted to the profile of the support rod (31) is processed on the surface of the support seat (32); a plurality of rotatable clamping rings (35) are installed on the crossbeam (36); the PCR tube (4) is conical and is installed in the clamping ring (35).

3. The pear genotype detection kit according to claim 1, characterized in that: The PCR tube (4) is pre-installed with polymerase, a tin foil sealing sheet (42) is installed on the top of the PCR tube (4), a closable cap (41) is connected to the top side wall of the PCR tube (4), and a dropper (44) is processed on the cap (41).

4. The pear genotype detection kit according to claim 1, characterized in that: The grinding and extraction chamber (5) comprises a culture chamber (51), a support cover (52) and a movable cover (53); the support cover (52) is mounted on the culture chamber (51); a clamping groove (54) for supporting and limiting the support cover (52) is processed on the culture chamber (51); a grinding groove (55) is processed at the center of the surface of the support cover (52); the movable cover (53) is rotatably mounted on the surface of the support cover (52); a grinding rod (56) extending into the grinding groove (55) is processed on the movable cover (53); a drainage tube (57) extending into the culture chamber (51) is processed at the bottom of the grinding groove (55); a plurality of reinforcing ribs (58) are processed on the bottom surface of the support cover (52) outside the drainage tube (57); one of the reinforcing ribs (58) extends downward and engages with the clamping groove (54).

5. The pear genotype detection kit according to claim 4, characterized in that: The surfaces of the grinding rod (56) and the grinding groove (55) are both processed with grinding teeth.

6. The pear genotype detection kit according to claim 4, characterized in that: A filter cavity (59) is processed on the side wall of the culture chamber (51), and filter cotton (510) is installed on the side wall of the filter cavity (59).

7. A method for detecting pear genotypes, characterized in that: The following steps are involved: (1) Sample collection: Open the box cover (2), take out the grinding and extraction chamber (5), then take fresh leaves or fruit tissues and place them in the grinding groove (55) on the support cover (52), then cover the movable cover (53), and rotate the movable cover (53) so that the grinding rod (56) on the movable cover (53) and the grinding groove (55) cooperate with each other, and grind the fresh leaves or fruits into a paste; (2) DNA extraction: Take out the syringe (62) from the net bag (6), suck up the slow-release liquid, and push it into the grinding groove (55) of the support cover (52). During this process, the movable cover (53) is continuously rotated to allow the sample tissue to flow into the culture chamber (51) along the drainage tube (57), and the culture chamber (51) is placed in an environment of 0 to 5° C. and allowed to stand for 30 minutes to wait for the DNA in the sample tissue to be released; (3) Gene amplification: Based on step (2), the culture chamber (51) is deflected to make the slow-release liquid gather toward the filter chamber (59), and the deflected state is maintained so that the slow-release liquid passes through the filter cotton (510) and is placed in the filter chamber (59). At this time, the folding frame (3) is unfolded, and the tin foil sealing sheet (42) on the corresponding PCR tube (4) is torn off, and then the quantitative pipette (61) is taken out, and the DNA in the filter chamber (59) is dripped into the PCR tube (4) in turn, and then the cap (41) is covered, and the PCR tube (4) is placed in the PCR instrument for circulation, and the number of DNA fragments is waited to increase; (4) Product detection: Turn the PCR tube (4) over, separate the product in the PCR tube (4) by electrophoresis on an agarose gel, stain with a nucleic acid dye, and observe the result using a gel imaging system; (5) Data analysis: The pear gene can be identified based on the electrophoresis pattern and the results of the previous analysis.

8. The method for detecting pear genotype according to claim 7, characterized in that: The sustained-release solution is prepared by uniformly mixing 1-3 ml of sodium alkyl sulfonate, 1-2 ml of sodium fatty alcohol ether sulfate, 1-2 g of sodium chloride, 50 ml of pure water and 3-5 ml of alcohol in parts by weight.

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