阅读说明：本技术 用于dna分子检测的小型昆虫浸制标本的简易长期保存方法 (Simple long-term preservation method of small insect impregnated specimen for DNA molecule detection ) 是由 刘霞 徐正会 于 2020-12-30 设计创作，主要内容包括：本发明公开一种用于DNA分子检测的小型昆虫浸制标本的简易长期保存方法,包括以下步骤：采集的新鲜昆虫标本放在装有无水乙醇的离心管中,加入无水乙醇至离心管的4/5高度,并书写标签投入离心管中；解剖镜下分类,同一采集地同种昆虫放入同一离心管；分类完毕后将同一采集地的离心管放入同一密闭容器,在容器中放入一片用透水材料包装的镁片,在容器中加无水乙醇至淹没镁片包装物；然后将该密闭容器避光保存,定期添加容器中的无水乙醇即可。本发明解决了因离心管密封性欠佳导致其中的保存液挥发的问题,同时可以降低外容器中的无水乙醇的含水量,使昆虫浸制标本得到长期或永久良好保存,并可根据标签标识快速查找。(The invention discloses a simple long-term preservation method of a small insect impregnated specimen for DNA molecule detection, which comprises the following steps: putting the collected fresh insect specimen into a centrifuge tube filled with absolute ethyl alcohol, adding the absolute ethyl alcohol to 4/5 height of the centrifuge tube, and putting a written label into the centrifuge tube; classifying under a dissecting mirror, and putting the same kind of insects in the same collection place into the same centrifugal tube; after classification, putting centrifuge tubes in the same collection place into the same closed container, putting a magnesium sheet packaged by a permeable material into the container, and adding absolute ethyl alcohol into the container until the magnesium sheet package is submerged; then storing the sealed container in dark place, and periodically adding anhydrous ethanol in the container. The invention solves the problem of volatilization of the preservation solution caused by poor sealing property of the centrifugal tube, and can reduce the water content of absolute ethyl alcohol in the outer container, so that the insect impregnated specimen can be preserved for a long time or permanently and can be quickly searched according to the label identification.)

1. A simple long-term preservation method of a small insect impregnated specimen for DNA molecule detection is characterized by comprising the following steps:

the method comprises the following steps: cleaning the collected fresh small insect specimen with absolute ethyl alcohol;

step two: putting the small insect specimen cleaned in the step one into a centrifuge tube, adding absolute ethyl alcohol to 4/5 height of the centrifuge tube, and putting a written label into the centrifuge tube;

step three: inserting the centrifuge tube obtained in the second step into a circular hole in a tray, wherein a plurality of circular holes for inserting the centrifuge tube are uniformly distributed in the tray, and the inner diameter of each circular hole is matched with the outer diameter of the inserted centrifuge tube; the tray is placed in a container with a cover, one or more steps for placing the tray are arranged on the inner wall of the container, the shape of the steps is matched with the shape of the periphery of the tray, a groove is formed in the center of the bottom of the container, a magnesium sheet package wrapped by a water-permeable material is placed in the groove, and absolute ethyl alcohol is added into the container until the magnesium sheet package is submerged;

step four: tightly sealing the container cover and storing in dark place;

step five: and (4) checking the volatilization condition of the absolute ethyl alcohol in the centrifugal tube and the container regularly, and if the volatilization of the absolute ethyl alcohol is completely or obviously reduced, adding the absolute ethyl alcohol again.

2. The simple long-term preservation method according to claim 1, wherein in the first step, the small insect specimen is placed in a centrifuge tube filled with absolute ethyl alcohol, preserved in a refrigerator at a low temperature of-20 ℃, taken out in batches, and cleaned with absolute ethyl alcohol.

3. The simple long-term preservation method according to claim 1, wherein in the second step, the cleaned small insect specimens are classified under a dissecting mirror, the insect specimens in the same centrifugal tube are the same kind of insects in the same collection place, and if the number of the same kind of insect specimens in the same collection place is large, the insect specimens are subpackaged into a plurality of centrifugal tubes; different insect specimens of different species are packed into different centrifuge tubes in the same collection place so as to be convenient for searching, but the insect specimens in the same nest are packed into the same centrifuge tube.

4. The simplified long-term storage method according to claim 1, wherein in step three, the inner diameter of the container is gradually reduced from top to bottom.

5. The simple long-term storage method according to claim 1, wherein in step three, the tray edge is further provided with symmetrical semicircular holes for taking and placing the tray when the centrifuge tube is filled and for flowing into the bottom of the container when the absolute ethyl alcohol is supplemented.

6. The simple long-term preservation method according to claim 1, wherein the centrifuge tube is 2 ml in size, and a label written with the contents of the collected place and the insect specimen is placed in the centrifuge tube.

7. The easy long-term preservation method according to claim 1 or 4, wherein a label is attached to the outside of the container.

Technical Field

The invention relates to the technical field of insect specimen preservation, in particular to a simple long-term preservation method of a small insect impregnated specimen for DNA molecule detection.

Background

At present, insect impregnated specimens are mainly preserved by using commercially available 2 ml or 5 ml centrifuge tubes, and preservation solutions are 75% alcohol solutions or 95% alcohol solutions, but for DNA molecule detection, the aqueous solutions cause DNA molecule hydrolysis, and the detection results are affected, so that absolute ethyl alcohol should be used as the preservation solution. However, the sealing performance of the centrifugal tube sold in the market at present is poor, ethanol is a solvent which is very easy to volatilize, and the fact that the preservation solution in 2 ml of centrifugal tube is obviously reduced or volatilized completely after 1 year is found in practice, the cover needs to be opened again to add the preservation solution, time and labor are wasted, and the specimen is easily damaged; in addition, the number of centrifuge tubes stored in the container is large, so that the sample is difficult to search; if a completely sealed centrifuge tube is developed, the cost is high. In addition, owing to put and need open the container when holding the centrifuging tube of sample, the anhydrous alcohol in the moisture in the air can get into the container, and then probably influences the purity of anhydrous alcohol in the centrifuging tube.

In summary, the existing preservation problems of insect soaking specimens are as follows: 75% alcohol solution or 95% alcohol solution can not be applied to the preparation of specimens in DNA molecular detection occasions; the insect-impregnated specimen is stored in 2 ml and 5 ml centrifuge tubes sold in the market at present, and the sealing performance of the centrifuge tubes is poor, so that the preservation solution (absolute ethyl alcohol) in the centrifuge tubes is easy to volatilize, and the moisture enters the ethyl alcohol in the specimen taking and placing process, so that the concentration of the ethyl alcohol is reduced, the specimen cannot be stored for a long time or permanently, and the specimen is difficult to search.

Disclosure of Invention

Aiming at the problems in the prior art, the invention provides a simple long-term preservation method of a small insect impregnated specimen for DNA molecule detection, which comprises the following steps:

the method comprises the following steps: cleaning the collected fresh small insect specimen with absolute ethyl alcohol;

step two: putting the small insect specimen cleaned in the step one into a centrifuge tube, adding absolute ethyl alcohol to 4/5 height of the centrifuge tube, and putting a written label into the centrifuge tube;

step three: inserting the centrifuge tube obtained in the second step into a circular hole in a tray, wherein a plurality of circular holes for inserting the centrifuge tube are uniformly distributed in the tray, and the inner diameter of each circular hole is matched with the outer diameter of the inserted centrifuge tube; the tray is placed in a container with a cover, one or more steps for placing the tray are arranged on the inner wall of the container, the shape of the steps is matched with the shape of the periphery of the tray, a groove is formed in the center of the bottom in the container and used for placing a magnesium sheet package wrapped by a water-permeable material, and absolute ethyl alcohol is added into the container until the magnesium sheet package is submerged;

step four: tightly sealing the container cover and storing in dark place;

step five: and (4) checking the volatilization condition of the absolute ethyl alcohol in the centrifugal tube and the container regularly, and if the volatilization of the absolute ethyl alcohol is completely or obviously reduced, adding the absolute ethyl alcohol again.

Further, in the first step, the small insect specimen is placed in a centrifuge tube filled with absolute ethyl alcohol, stored in a refrigerator at the low temperature of minus 20 ℃, taken out in batches and cleaned by absolute ethyl alcohol.

Further, in the second step, the cleaned small insect specimens are classified under an anatomical lens, the insect specimens in the same centrifugal tube are the same type of insects in the same collection place, and if the number of the same type of insect specimens in the same collection place is large, the insect specimens are subpackaged into a plurality of centrifugal tubes; different insect specimens of different species are packed into different centrifuge tubes in the same collection place so as to be convenient for searching, but the insect specimens in the same nest are packed into the same centrifuge tube.

Further, in the third step, the inner diameter of the container is gradually reduced from top to bottom; the tray edge still system has symmetrical semicircle orifice for get when putting the centrifuging tube and put the tray to and flow into the container bottom when supplementing anhydrous alcohol.

Furthermore, the centrifuge tube is selected to be 2 ml in specification, and a label written with the contents of the collection place and the insect specimen is placed in the centrifuge tube. The container is externally pasted with a label.

The invention utilizes the principle of evaporation, solves the problem of volatilization of the preservation solution in the centrifugal tube due to poor sealing property of the centrifugal tube, and can reduce the water content of absolute ethyl alcohol in the outer container, so that the insect-impregnated specimen can be well preserved for a long time or permanently; according to the label identification, the method can be used for quickly searching and greatly saving time compared with the existing method.

Drawings

FIG. 1 is a class-numbered object of an insect specimen centrifuge tube according to a first embodiment of the present invention;

FIG. 2 is a view showing an insect specimen as a plastic mineral water bottle in accordance with an embodiment of the present invention;

FIG. 3 is a physical example of an insect specimen storage organizer according to an embodiment of the present invention;

FIG. 4 is a schematic view of an insect specimen preservation device according to the principles of the present invention;

FIG. 5 is a schematic view of the container of FIG. 4;

FIG. 6 is a schematic view of the container lid of FIG. 4;

FIG. 7 is a schematic view of the tray of FIG. 4;

the labels in the figures are: 1-centrifuge tube, 3-container, 2-cover, 4-tray, 5-groove, used for placing packaging material wrapping magnesium sheet; 6-step for placing the tray.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail with reference to the following embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention. The examples do not specify particular techniques or conditions, and are performed according to the techniques or conditions described in the literature in the art or according to the product specifications. The raw materials, reagents or instruments used are not indicated by manufacturers, and are all conventional products which can be obtained by purchase.

Example one, the simple long-term preservation method of the small insect impregnated specimen for DNA molecule detection is performed according to the following steps, as shown in FIGS. 1 to 3:

the method comprises the following steps: instantly putting the collected fresh ant specimen into a 2 ml centrifugal tube with absolute ethyl alcohol (more than or equal to 99.5 percent) to be brought back to a laboratory, putting the fresh ant specimen into a low-temperature refrigerator at the temperature of-20 ℃ for storage, and then taking out the fresh ant specimen in batches to be cleaned by the absolute ethyl alcohol;

step two: classifying the cleaned ant specimens under a dissecting mirror, putting the same kind of ant specimens collected from the same place into the same centrifugal tube 1, adding absolute ethyl alcohol to the height of 4/5 degrees of the centrifugal tube, independently putting the ant specimens in the same nest into the same centrifugal tube, and writing labels into the same centrifugal tube, wherein the number records are shown in figure 1;

step three: collecting the waste plastic mineral water bottles, and cleaning the waste plastic mineral water bottles with clear water for later use; putting the centrifuge tube marked with the same collection place into a plastic mineral water bottle, placing a packaging object wrapping a magnesium sheet with a water permeable material in the plastic mineral water bottle, then adding absolute ethyl alcohol until submerging all the centrifuge tubes, writing a label and putting the label into the centrifuge tube or sticking the label outside the centrifuge tube, numbering and recording so as to be convenient for finding, and covering the cover tightly, see fig. 2.

Step four: the tightly covered plastic mineral water bottles are placed in order in a storage box to be stored in a dark place, and writing labels are pasted outside, as shown in figure 3.

Step five: checking the volatilization condition of the absolute ethyl alcohol in the centrifuge tube and the plastic mineral water bottle every 3-5 years, and if the absolute ethyl alcohol is completely volatilized or obviously reduced, only adding the absolute ethyl alcohol again, and storing for a long time or permanently.

Example two, a simple long-term preservation method of a small insect-impregnated specimen for DNA molecule detection is performed by the following steps, using a preservation apparatus as shown in fig. 4 to 7:

the method comprises the following steps: instantly putting the collected fresh ant specimen into a 2 ml centrifugal tube with absolute ethyl alcohol (more than or equal to 99.5 percent) to be brought back to a laboratory, putting the fresh ant specimen into a low-temperature refrigerator at the temperature of-20 ℃ for storage, and then taking out the fresh ant specimen in batches to be cleaned by the absolute ethyl alcohol;

step two: classifying the cleaned ant specimens under a dissecting mirror, putting the same kind of ant specimens collected from the same place into the same centrifugal tube 1, adding absolute ethyl alcohol to the height of 4/5 degrees of the centrifugal tube, independently putting the ant specimens in the same nest into the same centrifugal tube, and writing labels into the same centrifugal tube, wherein the labels are put into the same centrifugal tube;

step three: during inserting the round hole on tray 4 with the centrifuging tube external diameter of inserting, the internal diameter of round hole cooperatees, and tray 4 is put into the container 3 of taking container lid 2, container inner wall system have one or more be used for laying tray 4 step 6, and the shape of step and the peripheral shape cooperation of tray, the internal diameter of container from top to bottom reduces gradually. A groove 5 is formed in the center of the bottom in the container and used for placing a packaging material wrapping the magnesium sheet by a water-permeable material, absolute ethyl alcohol is added until the packaging material wrapping the magnesium sheet is submerged, and the writing label is externally attached to the container;

the tray on evenly distributed have a plurality of be used for inserting the round hole of putting the centrifuging tube, the semicircle orifice of symmetry is in addition in the tray edge for get from step 6 when putting the centrifuging tube full and put the tray, and let alcohol flow in the container bottom when supplementing anhydrous alcohol.

Step four: and (4) tightly covering the container in the third step with a cover 2, sealing, and storing in dark place.

Step five: checking the volatilization condition of the absolute ethyl alcohol in the centrifuge tube and the container regularly every 3-5 years, and if the volatilization of the absolute ethyl alcohol is almost exhausted or obviously reduced, only adding the absolute ethyl alcohol again to store for a long time or permanently.

The first embodiment or the second embodiment of the invention can solve the problem of volatilization of the preservation solution caused by poor sealing property of the centrifugal tube, so that the insect impregnated specimen can be well preserved for a long time or permanently; and can be searched fast according to the label record, and can save time greatly compared with the existing method.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.