CN109470760B - Device for preparing agarose gel, agarose gel electrophoresis method and application - Google Patents
Device for preparing agarose gel, agarose gel electrophoresis method and application Download PDFInfo
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- 239000011543 agarose gel Substances 0.000 title claims abstract description 127
- 238000000034 method Methods 0.000 title claims abstract description 49
- 238000000246 agarose gel electrophoresis Methods 0.000 title claims abstract description 48
- 239000000499 gel Substances 0.000 claims abstract description 136
- 239000003292 glue Substances 0.000 claims abstract description 60
- 238000001962 electrophoresis Methods 0.000 claims abstract description 40
- 210000001520 comb Anatomy 0.000 claims abstract description 28
- 239000007853 buffer solution Substances 0.000 claims abstract description 25
- 238000002360 preparation method Methods 0.000 claims abstract description 23
- 150000007523 nucleic acids Chemical class 0.000 claims description 33
- 102000039446 nucleic acids Human genes 0.000 claims description 31
- 108020004707 nucleic acids Proteins 0.000 claims description 31
- 239000000980 acid dye Substances 0.000 claims description 24
- 238000001502 gel electrophoresis Methods 0.000 claims description 12
- 238000007789 sealing Methods 0.000 claims description 11
- 238000004064 recycling Methods 0.000 abstract description 3
- 230000035945 sensitivity Effects 0.000 description 30
- 238000001514 detection method Methods 0.000 description 17
- 238000002474 experimental method Methods 0.000 description 11
- NOIIUHRQUVNIDD-UHFFFAOYSA-N 3-[[oxo(pyridin-4-yl)methyl]hydrazo]-N-(phenylmethyl)propanamide Chemical compound C=1C=CC=CC=1CNC(=O)CCNNC(=O)C1=CC=NC=C1 NOIIUHRQUVNIDD-UHFFFAOYSA-N 0.000 description 7
- 230000000694 effects Effects 0.000 description 6
- 229920000936 Agarose Polymers 0.000 description 3
- 230000015556 catabolic process Effects 0.000 description 2
- 238000006731 degradation reaction Methods 0.000 description 2
- 230000007774 longterm Effects 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 230000000149 penetrating effect Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 239000012945 sealing adhesive Substances 0.000 description 2
- 238000007711 solidification Methods 0.000 description 2
- 230000008023 solidification Effects 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 230000003321 amplification Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000004891 communication Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000002808 molecular sieve Substances 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 230000002035 prolonged effect Effects 0.000 description 1
- 239000012146 running buffer Substances 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- URGAHOPLAPQHLN-UHFFFAOYSA-N sodium aluminosilicate Chemical compound [Na+].[Al+3].[O-][Si]([O-])=O.[O-][Si]([O-])=O URGAHOPLAPQHLN-UHFFFAOYSA-N 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
- G01N27/416—Systems
- G01N27/447—Systems using electrophoresis
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
- G01N27/416—Systems
- G01N27/447—Systems using electrophoresis
- G01N27/44704—Details; Accessories
- G01N27/44747—Composition of gel or of carrier mixture
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- Electrochemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
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Abstract
The invention provides a device for preparing agarose gel, an agarose gel electrophoresis method and application, and relates to the technical field of electrophoresis. The device comprises a glue making platform, a gel tray, a sample adding comb and a glue storage container; the gel preparation platform is provided with at least one groove A for placing a gel tray, and two sample feeding combs are respectively arranged in parallel close to two opposite side walls of the groove A; at least one groove B for placing a gel tray and a space for storing buffer solution are arranged in the gel storage container so that agarose gel is immersed in the buffer solution; the device is matched with a gel storage container through the agarose gel with sample adding holes at two ends, so that the purpose of preparing the agarose gel for recycling is achieved. The agarose gel electrophoresis uses the reusable agarose gel prepared by the device for preparing agarose gel to carry out electrophoresis, can avoid gel preparation before electrophoresis, and simplifies experimental flow.
Description
Technical Field
The invention relates to the technical field of electrophoresis, in particular to a device for preparing agarose gel, an agarose gel electrophoresis method and application.
Background
The nucleic acid electrophoresis is an electrophoresis method with gel as a supporting electrophoresis medium and molecular sieve effect, has the advantages of high resolution, wide separation range and the like, and is an indispensable important experimental means in nucleic acid research. Among them, agarose gel electrophoresis is the most commonly used gel electrophoresis technique, which is suitable for nucleic acid electrophoresis and is a standard method for separating, identifying and purifying nucleic acid fragments.
The most commonly used gel electrophoresis in nucleic acid experiments is agarose gel electrophoresis, which is an important analysis method indispensable in molecular biology experiments such as sequence amplification, nucleic acid research and the like, and is widely applied to separating, identifying and purifying nucleic acid fragments at present. At present, agarose gel is prepared on site before each nucleic acid electrophoresis experiment is carried out, and most of the agarose gel is disposable, so that the experiment time and the experiment cost are wasted. In some laboratories, in order to save costs, the gel remaining after the primary test is often kept in a refrigerator at 4 ℃ for secondary use, but this can cause degradation of the nucleic acid dye, which reduces the sensitivity of the detection.
In view of this, the present invention has been made.
Disclosure of Invention
A first object of the present invention is to provide an apparatus for preparing agarose gel, which alleviates the technical problem that agarose gel cannot be reused in the prior art.
A second object of the present invention is to provide an agarose gel electrophoresis method for electrophoresis using the reusable agarose gel prepared by the apparatus for preparing agarose gel.
A third object of the present invention is to provide the use of the above-mentioned apparatus for preparing agarose gel or the above-mentioned agarose gel electrophoresis method in nucleic acid detection.
In order to solve the technical problems, the invention adopts the following technical scheme:
The present invention provides an apparatus for preparing agarose gel, the apparatus comprising: the device comprises a glue preparation platform, at least one gel tray, at least two sample feeding combs and a glue storage container;
The gel preparation platform is provided with at least one groove A for placing the gel tray, and the inner side wall of the groove A is contacted with flanges at two sides of the gel tray;
a plurality of first clamping grooves for detachably fixing two sample feeding combs are formed in the side wall of the groove A; the two sample feeding combs are respectively arranged in parallel near the two opposite side walls of the groove A;
At least one groove B for placing a gel tray is arranged in the gel storage container; the gel storage container is also provided with a space for storing buffer solution so that agarose gel is immersed in the buffer solution; the glue storage container further comprises a sealing device for sealing the glue storage container;
When in use, the gel tray is placed in the groove A of the gel preparation platform, then two sample feeding combs are fixed in the groove A, and the sample feeding combs are taken out after the gel is cooled; placing agarose gel subjected to gel electrophoresis or unused agarose gel into a gel tray, placing into a gel storage container, immersing into buffer solution, and sealing for storage.
Preferably, the glue storage container comprises a glue storage box body and a glue storage cover body; the gel storage box body is internally provided with at least one groove B for placing a gel tray, and the side wall of the groove B is lower than the side wall of the gel storage box body.
Preferably, a groove B is formed in the glue storage box body, second clamping grooves used for placing detachable baffles are formed in two opposite side walls of the groove B respectively, and the detachable baffles divide the groove B into a second groove B and a third groove B.
Preferably, the contact surface of the glue storage box body and the glue storage cover body is provided with a sealing rubber strip.
Preferably, the glue storage box body and the glue storage cover body are detachably linked or hinged.
Preferably, the ratio of the depth of the groove B to the height of the side wall of the glue storage box body is 1: (2-5).
The invention also provides an agarose gel electrophoresis method, which comprises the following steps: preparing an agarose gel using the apparatus for preparing an agarose gel of any one of claims 1 to 6; adding a sample to be detected into a plurality of sample application holes on one side of the agarose gel, then performing gel electrophoresis, and placing the agarose gel after electrophoresis into the gel storage container for storage; and adding the sample to be detected into a plurality of sample application holes on the other side for gel electrophoresis when the sample to be detected is used next time.
Preferably, after the agarose gel is used for 3-5 times, the nucleic acid dye and the sample to be detected are uniformly mixed and then added into the sample adding hole.
Preferably, the volume ratio of the nucleic acid dye to the sample to be tested is 1: (5-10); the nucleic acid dye is nucleic acid dye with the volume percentage concentration of 0.08 percent.
The invention also provides an application of the device for preparing agarose gel or the agarose gel electrophoresis method in nucleic acid detection.
Compared with the prior art, the invention has the following beneficial effects:
according to the device for preparing agarose gel, provided by the invention, the purpose of preparing agarose gel for recycling is achieved by preparing the agarose gel with the sample adding holes at two ends and matching with the gel storage container. The agarose gel prepared by the device can be repeatedly utilized and stored for a plurality of times, and the time and the experiment cost of an experimenter are saved under the condition of ensuring a good electrophoresis result.
According to the agarose gel electrophoresis method provided by the invention, the reusable agarose gel prepared by the device for preparing the agarose gel is used for electrophoresis, gel preparation can be avoided before electrophoresis, and the reusable agarose gel has similar detection sensitivity with the agarose gel prepared at present. Therefore, the agarose gel electrophoresis method provided by the invention optimizes the experimental flow and improves the experimental efficiency.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are needed in the description of the embodiments or the prior art will be briefly described, and it is obvious that the drawings in the description below are some embodiments of the present invention, and other drawings can be obtained according to the drawings without inventive effort for a person skilled in the art.
Fig. 1 is a schematic side view of a glue making platform provided in embodiment 1;
Fig. 2 is a schematic diagram of one side of the glue making platform provided in embodiment 1;
fig. 3 is a schematic view of the other side of the glue producing platform provided in embodiment 1;
FIG. 4 is a schematic side view of the gel storage container according to example 1 in which agarose gel is stored;
FIG. 5 is a schematic view of a glue reservoir according to example 1;
FIG. 6 is a schematic view of another glue reservoir provided in example 1;
FIG. 7-A is the sensitivity of agarose gel at the first use in the agarose gel electrophoresis method provided in example 2;
FIG. 7-B is the sensitivity of agarose gel at the third use in the agarose gel electrophoresis method provided in example 2;
FIG. 7-C shows the sensitivity of agarose gel in the fifth use in the agarose gel electrophoresis method provided in example 2;
FIG. 7-D is the sensitivity of agarose gel at the sixth use in the agarose gel electrophoresis method provided in example 2;
FIG. 7-E is the sensitivity of agarose gel at the seventh use in the agarose gel electrophoresis method provided in example 2;
FIG. 8-A is the sensitivity of agarose gel at the first use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-B is the sensitivity of agarose gel at the third use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-C shows the sensitivity of agarose gel in the fifth use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-D is the sensitivity of agarose gel at the seventh use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-E is the sensitivity of agarose gel at ninth use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-F shows the sensitivity of agarose gel at the tenth use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-G shows the sensitivity of agarose gel at the tenth use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-H shows the sensitivity of agarose gel in the twelfth use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-I shows the sensitivity of agarose gel at the tenth use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-J is the sensitivity of agarose gel at the sixteenth use in the agarose gel electrophoresis method provided in example 3;
FIG. 8-K shows the sensitivity of agarose gel at the eighteenth use in the agarose gel electrophoresis method provided in example 3.
Icon: 100-a glue preparation platform; 111-a first groove a; 122-a second groove a; 123-third groove a; 130-a first card slot; 200-gel tray; 300-sample adding comb; 400-glue storage container; 410-groove B; 422-second groove B; 423-a third groove B; 430-a second card slot; 440-a glue storage box body; 450-a glue storage cover body; 460-removable baffle.
Detailed Description
The following description of the embodiments of the present invention will be made apparent and fully in view of the accompanying drawings, in which some, but not all embodiments of the invention are shown. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
In the description of the present invention, it should be noted that, if terms such as "center", "upper", "lower", "left", "right", "vertical", "horizontal", "inner", "outer", and the like are used, the indicated orientation or positional relationship is based on the orientation or positional relationship shown in the drawings, only for convenience of describing the present invention and simplifying the description, and does not indicate or imply that the indicated apparatus or element must have a specific orientation, be configured and operated in a specific orientation, and thus should not be construed as limiting the present invention. Furthermore, the terms "first," "second," "third," and the like, as used herein, are used for descriptive purposes only and are not to be construed as indicating or implying any relative importance.
In the description of the present invention, it should be noted that unless explicitly stated and limited otherwise, the terms "mounted," "connected," and "connected" should be construed broadly, and may be, for example, fixedly connected, detachably connected, or integrally connected; can be mechanically or electrically connected; can be directly connected or indirectly connected through an intermediate medium, and can be communication between two elements. The specific meaning of the above terms in the present invention will be understood in specific cases by those of ordinary skill in the art.
The present invention provides an apparatus for preparing agarose gel, the apparatus comprising: the device comprises a glue preparation platform, at least one gel tray, at least two sample feeding combs and a glue storage container;
The gel preparation platform is provided with at least one groove A for placing the gel tray, and the inner side wall of the groove A is contacted with flanges at two sides of the gel tray so as to prevent molten gel from penetrating into a gap between the gel preparation platform and the gel tray;
A plurality of first clamping grooves for detachably fixing two sample feeding combs are formed in the side wall of the groove A; the two sample feeding combs are respectively arranged in parallel near the two opposite side walls of the groove A;
At least one groove B for placing a gel tray is arranged in the gel storage container; the gel storage container is also provided with a space for storing buffer solution so that agarose gel is immersed in the buffer solution; the glue storage container further comprises a sealing device for sealing the glue storage container;
According to the invention, the agarose gel with sample adding holes at two ends is prepared to be matched with the gel storage container, so that the purpose of preparing the agarose gel for recycling is achieved.
When the gel tray is used, the gel tray is placed in the groove A of the gel preparation platform, then two sample feeding combs are fixed in the groove A, agarose with certain concentration is melted and then poured into the gel preparation platform, and the sample feeding combs are taken out after the gel is cooled. Because the two ends of the groove A are respectively inserted with a sample adding comb, a row of sample adding holes can be respectively manufactured at the upper end and the lower end of the agarose gel. When in use, firstly, a sample to be electrophoresed is added into a sample adding hole at one side for electrophoresis, after electrophoresis, agarose gel is placed in a gel tray and put into a gel storage container, and the gel storage container is immersed in buffer solution and then sealed for storage. And when the sample is used next time, adding the sample to be detected into the unused sample adding hole at the last time, and then carrying out electrophoresis.
Because the directions of the same agarose gel in the electrophoresis tank are opposite in the two electrophoresis processes, namely, the sample adding hole spotted in the first electrophoresis is close to the negative electrode, and the sample moves towards the center of the agarose gel; in the second electrophoresis, the sample adding hole is close to the positive electrode, so that the last sample remained in the agarose gel moves outwards of the agarose gel until the last sample moves outwards of the agarose gel, and the sample adding hole spotted in the last electrophoresis can be used continuously for the next time.
Meanwhile, the agarose gel is not easy to store in the repeated electrophoresis and storage processes, so that the device is provided with the gel storage container, and the agarose gel subjected to gel electrophoresis or unused agarose gel can be immersed in the buffer solution, so that the purpose of long-term storage is achieved.
The agarose gel prepared by the device can be repeatedly used, is easy to store, and saves time and experiment cost of experimenters under the condition of ensuring good electrophoresis results.
In some alternative embodiments, the glue reservoir comprises a glue reservoir cartridge and a glue reservoir cap; the gel storage box body is internally provided with at least one groove B for placing a gel tray, and the side wall of the groove B is lower than the side wall of the gel storage box body. When the agarose gel placed in the gel tray is placed in the groove B, enough space can be reserved above the gel storage box body to pour the buffer solution, so that the agarose gel can be fully immersed in the buffer solution, and the agarose gel is prevented from breaking due to drying, so that the service life is prolonged.
In some optional embodiments, a groove B is provided in the glue storage box, two opposite side walls of the groove B are respectively provided with a second clamping groove for placing a detachable baffle, and the detachable baffle divides the groove B into a second groove B and a third groove B for storing gel trays with various sizes and agarose gels contained therein.
In some preferred embodiments, the glue storage box body and the glue storage cover body are detachably linked or hinged, and a sealing adhesive tape is preferably arranged on the contact surface of the glue storage box body and the glue storage cover body, so that the sealing effect is enhanced, the leakage of the buffer solution is prevented, and the volatilization of the buffer solution is reduced.
In some preferred embodiments, the ratio of the depth of the groove B to the height of the side wall of the glue storage box is 1: (2-5), for example, but not limited to, 1:2, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5, or 1:5, preferably 1: (3-4), more preferably 1:4. If the distance between the upper edge of the side wall of the groove B in the glue storage box and the upper edge of the glue storage box body is too large, the space and the buffer solution are wasted; if the upper edge of the side wall of the groove B in the glue storage box is too small in distance from the upper edge of the glue storage box body, the space reserved for the buffer solution is too small, so that on one hand, the volume of the buffer solution for immersing the agarose gel is too small, the agarose gel is easy to dry and crack, and on the other hand, the buffer solution is easy to overflow. Therefore, the preservation effect of the agarose gel can be improved by selecting a proper ratio of the depth of the groove B to the height of the side wall of the gel storage box body.
In a preferred embodiment of the present invention, the agarose gel is prepared and stored with better effect when the size parameters of the apparatus for preparing agarose gel are as follows:
The inner diameter of the glue storage box body is 150mm multiplied by 150mm, and the height of the glue storage box body is 20mm; the inner diameter of the groove B is 135cm multiplied by 135cm, and the depth is 5mm.
The inner diameter of the glue making platform is 120mm multiplied by 120mm, and the depth of the groove A is 10mm.
The dimensions of the gel tray include 60mm by 120mm and/or 120mm by 120mm.
The sample adding comb is fixed parallel to the side of the gel tray, the length of which is 120 mm; the sample feeding comb comprises 11-hole sample feeding combs with 1 mm/tooth and 25-hole sample feeding combs with 1 mm/tooth, wherein the thickness of each 1 mm/tooth of the sample feeding comb is 1mm.
The invention also provides an agarose gel electrophoresis method, which comprises the steps of preparing agarose gel by using the device for preparing agarose gel; adding a sample to be detected into a plurality of sample application holes on one side of the agarose gel, then performing gel electrophoresis, and placing the agarose gel after electrophoresis into the gel storage container for storage; and adding the sample to be detected into a plurality of sample application holes on the other side for gel electrophoresis when the sample to be detected is used next time.
According to the agarose gel electrophoresis method provided by the invention, the reusable agarose gel prepared by the device for preparing the agarose gel is used for electrophoresis, gel preparation can be avoided before electrophoresis, and the reusable agarose gel has similar detection sensitivity with the agarose gel prepared at present. Therefore, the agarose gel electrophoresis method provided by the invention simplifies the experimental flow and improves the experimental efficiency.
In some preferred embodiments, the agarose gel electrophoresis method provided by the invention is implemented according to the following steps of:
(a) When agarose gel is prepared for the first time, adding nucleic acid dye (added according to the proportion in the specification of the nucleic acid dye) before gel solidification, pouring the gel into a gel preparation platform, respectively inserting two sample feeding combs into first clamping grooves at two ends of the gel preparation platform, and carrying out nucleic acid electrophoresis after the gel is cooled and solidified.
(B) After the nucleic acid electrophoresis is finished and shooting is carried out, or agarose gel which is not prepared for use is placed in a gel tray and placed in a gel storage container, electrophoresis buffer solution is added into the gel storage container, the gel storage container is sealed, and the gel storage container is placed in a refrigerator at 4 ℃ for storage and is used next time.
(C) And then, when the sample is repeatedly used, the sample is directly added into the sample adding hole at the other end opposite to the previous one, so that the sample to be detected moves from the negative electrode to the positive electrode, and the sample existing in the gel in the previous experiment can migrate out of the gel from the other side, thereby avoiding the influence of the sample to be detected. After each experiment is finished, the gel is placed in a gel storage container, added with electrophoresis buffer solution and stored in a refrigerator at 4 ℃, and can be repeatedly used for four times.
In some preferred embodiments, after 3-5 times of repeated use of agarose gel, the added sample is mixed with the nucleic acid dye at each detection and then added in order to prevent the decrease of detection sensitivity due to the small degradation of the nucleic acid dye.
In some preferred embodiments, a nucleic acid dye is used at a concentration of 0.08% by volume, with a volume ratio to the sample to be tested of 1: (5-10) mixing, which may be, for example, but not limited to, 1:5, 1:6, 1:7, 1:8, 1:9, or 1:10; when the volume percent concentration of the nucleic acid dye is 0.08%, the nucleic acid dye and the sample are mixed with 1:6, the optimal electrophoresis effect and detection sensitivity can be achieved by electrophoresis after mixing, the DNA content of about 0.5ng can be detected, and the sensitivity is equivalent to that of the traditional agarose gel which is prepared at present. The nucleic acid dye according to the present embodiment may be a commercially available common nucleic acid dye, and may be, for example, but not limited to, EB, gold view, GENEGREEN, SYBR safe, ecoDye, or the like. The agarose gel can be repeatedly used for twelve times within two weeks by optimizing the proportion of the nucleic acid dye and the sample to be tested, and the sensitivity of the agarose gel is equivalent to that of the agarose gel which is prepared in the prior art.
The invention also provides an application of the device for preparing agarose gel or the agarose gel electrophoresis method in nucleic acid detection, which can be applied to agarose gel electrophoresis detection of nucleic acid samples, such as detection of DNA extracted from tissues or cells or detection of PCR products.
Example 1
This example provides an apparatus for preparing agarose gel as shown in fig. 1-6. The device comprises a glue making platform 100, a gel tray 200, two sets of sample feeding combs 300 and a glue storage container 400.
The two surfaces of the glue making platform 100 are provided with one or a plurality of grooves A, and the depth of each groove A is 10mm. The inner side walls of the grooves a are in contact with the flanges on both sides of the gel tray 200 to prevent melted gel from penetrating into the gap between the gel-making stage 100 and the gel tray 200. The side wall of the groove A is provided with a plurality of first clamping grooves 130 for detachably fixing the two sample application combs 300, and when the sample application combs are used, the two sample application combs 300 are respectively close to the two opposite side walls of the groove A and are arranged in parallel.
One face of the glue making platform 100 is provided with a first recess a 110 for placing a gel tray 200 of dimensions 120mm x 120mm, as shown in fig. 2. The other surface of the glue making platform 100 is provided with two grooves a, namely a second groove a 122 and a third groove a 123, as shown in fig. 3, the second groove a 122 and the third groove a 123 are formed by equally dividing raised strips parallel to the side wall of the groove a in the groove a, and are used for placing a gel tray 200 with the size of 60mm×120 mm.
The apparatus for preparing agarose gel comprises gel trays 200 having dimensions of 120mm by 120mm and 120mm by 60mm, respectively; two sets of loading combs 300, each set of loading combs 300 comprising an 11 hole 1 mm/tooth loading comb 300 and a 25 hole 1 mm/tooth loading comb 300, wherein 1 mm/tooth means that the loading comb 300 has a tooth thickness of 1mm.
The device for preparing agarose gel further comprises a glue storage container 400, which consists of a glue storage box body 440 and a glue storage cover body 450, wherein the glue storage box body 440 and the glue storage cover body 450 are hinged, and the contact surface is provided with a sealing adhesive tape. The inner diameter of the glue storage box 440 is 150mm by 150mm and the height is 20mm. The glue storage box body 440 is provided with a groove B410, the depth of the groove B410 is 5mm, the inner diameter of the groove B410 is 135cm multiplied by 135cm, the groove B410 is used for placing the gel tray 200 with the size of 120mm multiplied by 120mm, and the inner diameter of the groove B410 is slightly larger than the size of the gel tray 200, so that the gel tray 200 can be conveniently taken.
The two opposite sidewalls of the groove B410 are respectively provided with a second clamping groove 430, which is respectively disposed in the middle of the two opposite sidewalls of the groove B410, and is used for placing the detachable baffle 460. The detachable baffle 460 is parallel to the sidewall of the groove B410, dividing the groove B into the second groove B422 and the third groove B423 on average. The second groove B422 and the third groove B423 may place the gel tray 200 having dimensions of 120mm×60mm, respectively.
In use, the gel tray 200 is placed in the groove a of the gel making platform 100, and then two sample application combs 300 are fixed in the groove a, so that the sample application combs 300 with the same or different hole numbers can be fixed, as shown in fig. 1. After agarose with a certain concentration is melted, pouring the melted agarose into the gel preparation platform 100, and taking out the sample adding comb 300 after the gel is cooled. Since one sample-adding comb 300 is inserted into each of both ends of the groove A, a row of sample-adding holes can be formed at each of the upper and lower ends of the agarose gel. When in use, the sample to be electrophoresed is firstly added into the sample adding hole at one side for electrophoresis.
After electrophoresis, agarose gel is placed in a gel tray 200 and placed in a gel storage container 400, and is immersed in a buffer solution and then stored in a sealed manner. When the gel tray 200 with the size of 120mm multiplied by 120mm and agarose gel contained therein are stored, the gel tray 200 is directly placed into the groove B410 of the gel storage container 400; when storing the gel tray 200 with the size of 120mm×60mm and the agarose gel contained therein, the detachable baffle 460 is inserted into the second clamping groove 430, the groove B410 is divided into the second groove B422 and the third groove B423, and then the gel tray 200 with the size of 120mm×60mm and the agarose gel contained therein are placed into the second groove B422 or the third groove B423, or the agarose gel can be stored in the second groove B422 or the third groove B423 at the same time, as shown in fig. 4 to 6.
And when the sample is used next time, adding the sample to be detected into the unused sample adding hole at the last time, and then carrying out electrophoresis. Because the directions of the same agarose gel in the electrophoresis tank are opposite in the two electrophoresis processes, namely, the sample adding hole spotted in the first electrophoresis is close to the negative electrode, and the sample moves towards the center of the agarose gel; the sample adding hole for the last sample adding is close to the positive electrode in the second electrophoresis, so that the residual last sample in the agarose gel moves towards the outside of the agarose gel until the last sample moves to the outside of the agarose gel, and the sample adding hole spotted in the first electrophoresis can be used continuously for the next time.
Meanwhile, the agarose gel is not easy to store in the repeated electrophoresis and storage processes, so the device is also provided with the gel storage container 400, and the agarose gel subjected to gel electrophoresis or unused agarose gel can be immersed in the buffer solution, so that the purpose of long-term storage is achieved.
Example 2
This example provides a agarose gel electrophoresis method, using the agarose gel apparatus for preparing agarose gel provided in example 1, comprising the steps of:
(1) In the first preparation of agarose gel, nucleic acid dye (added according to the proportion in the description of the nucleic acid dye) is added before gel solidification, the gel is poured into the gel preparation platform 100, two sample application combs 300 are respectively inserted into the first clamping grooves 130 at two ends of the gel preparation platform 100, and after the gel is cooled and solidified, the nucleic acid electrophoresis is carried out.
(2) After the nucleic acid electrophoresis is finished, the gel is placed in a gel tray 200 and placed in a gel storage container 400, electrophoresis buffer solution is added into the gel storage container 400, and the gel is sealed and then placed in a refrigerator at 4 ℃ for storage, and is used next time.
(3) And then, when the sample is repeatedly used, the sample is directly added into the sample adding hole at the other end opposite to the previous one, so that the sample to be detected moves from the negative electrode to the positive electrode, and the sample existing in the gel in the previous experiment can migrate out of the gel from the other side, thereby avoiding the influence of the sample to be detected. After each experiment, the gel was stored in a 4℃refrigerator with running buffer in the gel storage container 400.
As shown in FIGS. 7-A to 7-E, the agarose gel electrophoresis method provided by the embodiment can reuse the agarose gel at least four times, and the detection sensitivity is reduced when the agarose gel is reused for the fifth time, as shown in the figures, the agarose gel electrophoresis results are shown in FIGS. 7-A to 7-E, wherein the agarose gel sensitivity is the sensitivity of agarose gel during the first, third, fifth, sixth and seventh repeated use.
Example 3
This example provides an agarose gel electrophoresis method, which differs from example 2 in that: after four times of repeated use, the nucleic acid dye was degraded slightly, and in order to prevent the decrease in detection sensitivity, the added sample was mixed with the nucleic acid dye at each detection and then added. The nucleic acid dye with the volume percent concentration of 0.08% in this example was used with the sample loading at 1:6, as shown in fig. 8-a to 8-K, the optimal electrophoresis effect and detection sensitivity can be achieved, and the DNA content of about 0.5ng can be detected, and the agarose gel electrophoresis method provided in this embodiment is repeatedly used for twelve times within two weeks, and the sensitivity is equivalent to that of the conventional agarose gel for the current preparation. However, after the agarose gel is stored for two weeks or repeatedly used for twelve times, the agarose gel has the phenomena of reduced hardness, easy breakage, reduced sensitivity of detection results, scattered tailing and the like, and is not suggested to be used in the later period.
Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and not for limiting the same; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some or all of the technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit of the invention.
Claims (8)
1. An agarose gel electrophoresis method, comprising: preparing an agarose gel using an apparatus for preparing an agarose gel; adding a sample to be detected into a plurality of sample application holes on one side of the agarose gel, then performing gel electrophoresis, and placing the agarose gel after electrophoresis into a gel storage container for storage; when the sample to be measured is used next time, adding the sample to be measured into a plurality of sample application holes on the other side for gel electrophoresis;
the apparatus for preparing agarose gel includes: the device comprises a glue preparation platform, at least one gel tray, at least two sample feeding combs and a glue storage container;
The gel preparation platform is provided with at least one groove A for placing the gel tray, and the inner side wall of the groove A is contacted with flanges at two sides of the gel tray;
a plurality of first clamping grooves for detachably fixing two sample feeding combs are formed in the side wall of the groove A; the two sample feeding combs are respectively arranged in parallel near the two opposite side walls of the groove A;
At least one groove B for placing a gel tray is arranged in the gel storage container; the gel storage container is also provided with a space for storing buffer solution so that agarose gel is immersed in the buffer solution; the glue storage container further comprises a sealing device for sealing the glue storage container;
When in use, the gel tray is placed in the groove A of the gel preparation platform, then two sample feeding combs are fixed in the groove A, and the sample feeding combs are taken out after the gel is cooled; placing agarose gel subjected to gel electrophoresis or unused agarose gel into a gel tray, placing into a gel storage container, immersing into buffer solution, and sealing for storage.
2. The agarose gel electrophoresis method of claim 1, wherein the gel storage container comprises a gel storage box and a gel storage cover; the gel storage box body is internally provided with at least one groove B for placing a gel tray, and the side wall of the groove B is lower than the side wall of the gel storage box body.
3. The agarose gel electrophoresis method according to claim 2, wherein a groove B is provided in the gel storage case, and two opposite sidewalls of the groove B are respectively provided with a second clamping groove for placing a detachable baffle plate, and the detachable baffle plate divides the groove B into a second groove B and a third groove B.
4. The agarose gel electrophoresis method according to claim 2 or 3, wherein a sealing rubber strip is arranged on the contact surface of the gel storage box body and the gel storage cover body.
5. A method of agarose gel electrophoresis according to claim 2 or 3 wherein the gel storage cassette and the gel storage cap are detachably linked or hinged.
6. The agarose gel electrophoresis method according to claim 2, wherein the ratio of the depth of the groove B to the height of the side wall of the gel storage case is 1: (2-5).
7. The agarose gel electrophoresis method of claim 1, wherein the agarose gel is applied 3-5 times, and the nucleic acid dye is mixed with the sample to be measured before being added into the sample addition well.
8. The agarose gel electrophoresis method of claim 7, wherein the volume ratio of the nucleic acid dye to the sample to be tested is 1: (5-10); the nucleic acid dye is nucleic acid dye with the volume percentage concentration of 0.08 percent.
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| CN205333647U (en) * | 2015-12-11 | 2016-06-22 | 中国农业科学院特产研究所 | Counter immunoelectrophoresis square hole agarose gel's preparation facilities |
| CN206057247U (en) * | 2016-08-31 | 2017-03-29 | 武汉爱博泰克生物科技有限公司 | Multiple rows of sample well agarose gel preparation facilitiess |
| CN209764787U (en) * | 2018-12-26 | 2019-12-10 | 天康生物股份有限公司 | Device for preparing agarose gel and gel electrophoresis device |
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| CN203798772U (en) * | 2014-04-25 | 2014-08-27 | 中国农业科学院兰州畜牧与兽药研究所 | Sepharose gel making device |
| CN205333647U (en) * | 2015-12-11 | 2016-06-22 | 中国农业科学院特产研究所 | Counter immunoelectrophoresis square hole agarose gel's preparation facilities |
| CN206057247U (en) * | 2016-08-31 | 2017-03-29 | 武汉爱博泰克生物科技有限公司 | Multiple rows of sample well agarose gel preparation facilitiess |
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