CN219670495U - An experimental system for ballast water genetic sample preparation - Google Patents

Abstract

The utility model discloses an experimental system for preparing ballast water genetic samples, which includes a support base, a sample filtering mechanism, a sample enrichment mechanism, a sterile refrigerator and a layered placement rack; the bottom end of the sample filtering mechanism is detachable It is equipped with a sample enrichment mechanism. The sample filtration mechanism includes first-stage filtration and second-stage filtration. The first-stage filtration removes impurities visible to the naked eye, and the second-stage filtration removes plankton and plants larger than 100 μm. The sample enrichment mechanism can store the filtered of ballast water. This experimental system for ballast water genetic sample preparation is easy to disassemble, sample, clean, and operate. It can be used in the field or in the laboratory, and can be sterilized and reused. The multi-stage filtration setting prevents impurities in ballast water from clogging the filter membrane. , different filtrate can be used for different index tests respectively, achieving one-time operation and reducing resource waste and environmental pollution.

Description

An experimental system for ballast water genetic sample preparation

Technical field

The invention is suitable for research on the risk of invasion of new pollutants during the discharge of ship ballast water, especially the risk of the spread of antibiotic resistance genes in ballast water in the marine environment. It is a method for extracting antibiotic resistance gene samples from ballast water. Pretreatment experimental system.

Background technique

Antibiotics are widely used to protect the health of humans and animals or to increase the growth rate of animals. However, 70% of these antibiotics are excreted into the environment in the form of urine and feces, causing resistance genes to be generated in the environment and further passed through organisms. spread. The spread of antibiotic resistance genes (ARGs) has become a serious global problem in recent years. In 2006, ARGs were first proposed as a type of emerging environmental pollutants. Unlike other pollutants, ARGs can persist in organisms for a long time and be transferred to other organisms. Therefore, the persistence, migration and transformation of ARGs in the environment are more harmful to the ecological environment than the residues of antibiotics themselves.

In recent years, large amounts of ARGs have been detected in water environments in estuaries, ports, and coastal areas, and seawater containing ARGs may be pumped into ship ballast tanks and transferred to the global ocean. However, the existing documents related to the supervision of ship ballast water do not have corresponding control requirements for ARGs, and there is no corresponding sample preparation and detection method for ship ballast water ARGs, making ship ballast water a global spread of ARGs. important carrier. Systematically understanding the levels and composition of ARGs in ballast water will help to understand and control the potential adverse effects of ballast water on marine ecology and human health, and provide a basis for ballast water discharge management.

Utility model content

This utility model is suitable for research on the risk of invasion of new pollutants during the discharge of ballast water from ships, especially the risk of the spread of antibiotic resistance genes in ballast water in the marine environment. It is a method for extracting antibiotic resistance gene samples from ballast water. pretreatment experimental system.

In order to achieve the above purpose, the present utility model provides the following technical solutions: an experimental system for preparing ballast water genetic samples, including a support base, a sample filtering mechanism, a sample enrichment mechanism, a sterile refrigerator and a layered placement rack;

The bottom end of the sample filtering mechanism is detachably provided with a sample enrichment mechanism and can achieve sealing. The sample filtering mechanism includes first-stage filtration and second-stage filtration. The first-stage filtration removes impurities visible to the naked eye, and the second-stage filtration Remove phytoplankton and plants larger than 100μm, and the sample enrichment mechanism can store filtered ballast water;

The inner cavity of the sterile refrigerator is provided with a layered placement rack, and the petri dishes are placed in the sterile refrigerator and stored using the layered placement rack.

Preferably, in order to achieve two-stage filtration, a sample filtering mechanism is provided, in which the inner cavity of the glass body is detachably provided with a second-stage filter cartridge, and the inner cavity of the second-stage filter cartridge is detachably provided with a conical filter. The first stage filter of the structure.

Preferably, in order to store the filtered ballast water, a sample enrichment mechanism is provided, by detachably connecting the filter head to the main body of the glass, and using a sand core filter plate and a 0.22 μm nitrocellulose filter membrane to reprocess the water. Filter, so that the water enters the enrichment bottle that is detachably connected at the bottom of the filter head.

During the operation, the two-stage filtration system is set up and placed in the main body of the glass. The main body of the glass is then connected to the enrichment system (sealed through fixed clips or frosted). The ballast water samples first pass through the two-stage filtration, and then filter The mechanism can be taken out, and the water sample inside the glass body 21 can be passed through a 0.22um filter membrane through suction. The filter membrane is used for gene extraction, and the water sample can be used for other measurements.

Compared with the existing technology, the beneficial effects of this utility model are: the experimental system for preparing ballast water genetic samples is easy to disassemble, sample, and clean, and is easy to operate. It can be used in the field or in the laboratory, and can be sterilized and repeated Use multi-stage filtration settings to avoid impurities in ballast water clogging the filter membrane. Different filtrate can be used for different index tests to achieve one-time operation and reduce resource waste and environmental pollution.

Description of drawings

Figure 1 is a schematic structural diagram of the utility model;

Figure 2 is a schematic diagram of the combined structure of the sample filtering mechanism and sample enrichment mechanism of the present invention;

Figure 3 is a schematic structural diagram of the sample filtering mechanism of the present utility model;

Figure 4 is a schematic structural diagram of the sample enrichment mechanism of the present invention.

In the picture: 1. Support seat, 2. Sample filtration mechanism, 21. Glass body, 22. Outlet faucet, 23. 100μm filter membrane structure, 24. Second-stage filter cartridge, 25. Snap ring two, 26. First Stage filter, 27. Snap ring one, 3. Sample enrichment mechanism, 31. Enrichment bottle, 32. Suction filter port, 33. Filter head, 34. Sand core filter plate, 35. 0.22μm nitrocellulose filter membrane, 4. Sterile refrigerator, 5. Layered racks.

Detailed ways

The technical solutions in the embodiments of the present utility model will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present utility model. Obviously, the described embodiments are only part of the embodiments of the present utility model, not all implementations. example. Based on the embodiments of the present utility model, all other embodiments obtained by those of ordinary skill in the art without creative efforts fall within the scope of protection of the present utility model.

Please refer to Figures 1-4. The present utility model provides a technical solution: an experimental system for preparing ballast water genetic samples, including a support base 1, a sample filtering mechanism 2, a sample enrichment mechanism 3, a sterile refrigerator 4 and The rack 5 is placed in layers, and the top of the support base 1 is movably provided with a sample filtering mechanism 2, a sample enrichment mechanism 3 and a sterile refrigerator 4 from left to right. The sample filtering mechanism 2 is detachably disposed on the sample. The top end of the enrichment mechanism 3 and the inner cavity of the sterile refrigerator 4 are provided with a layered placement rack 5 .

More specifically, the sample filtering mechanism 2 includes: a glass body 21, a spout 22, a 100 μm filter membrane structure 23, a second-stage filter cartridge 24, a second snap ring 25, a first-stage filter 26, and a snap ring 27;

The glass body 21 is hollow from top to bottom and is made of high borosilicate with a volume of 500ml. The diameter of the bottom edge of the glass body 21 is larger than the hollow diameter and the edge is made of frosted material. The second-stage filter cartridge 24 is detachably arranged on the glass. In the inner cavity of the cup body 21, the diameter of the second-stage filter cylinder 24 is smaller than the diameter of the glass cup body 21, and the top of the second-stage filter cylinder 24 is fixed with a snap ring 25. The snap ring 25 can be placed on the glass body. The top of 21 is used to fix the second-stage filter cartridge 24 in the glass body 21. The 100 μm filter membrane structure 23 is threadedly connected to the bottom end of the second-stage filter cartridge 24 to filter and remove zooplankton and plants larger than 100 μm. The second stage The space between the filter cylinder 24 and the glass body 21 is the ballast water storage space 2. The first-stage filter 26 is detachably arranged in the inner cavity of the second-stage filter cylinder 24, and the top of the first-stage filter 26 is fixed. A snap ring 27 is provided, and the snap ring 27 can be placed on the top of the snap ring 22 25 to fix the first-stage filter 26 in the second-stage filter cartridge 24;

In this embodiment, the first-stage filter 26 has a conical structure and is a large-pore filter to remove impurities visible to the naked eye;

The space between the first-stage filter 26 and the second-stage filter cartridge 24 is the ballast water storage space 1, and the outlet faucet 22 is provided on the outer wall of the glass body 21;

The lower surfaces of snap ring one 27 and snap ring two 25 are both provided with annular slots, so that snap ring two 25 can be stuck on the glass body 21 by using the annular slot, and snap ring one 27 can be stuck on the snap ring by using the annular slot. 2:25a.

The ballast water sample should first pass through the sample filtering mechanism 2 to remove impurities and phytoplankton. The ballast water sample passing through the first-stage filter 26 enters the ballast water storage space 1. This part of the ballast water can continue to be used for ballast water. In compliance with the D-2 standard of the Convention on Carrying Water, the ballast water sample that undergoes second-stage filtration through the 100 μm filter membrane structure 23 enters the ballast water storage space 2. In order to control the volume of suction filtration, the outer wall of the glass body 21 is There is a scale, and the side wall of the ballast water storage space 2 is provided with an outlet faucet 22, which is located at the 200ml volume mark of the glass to facilitate the control of the volume of ballast water entering the next step, and the ballast water flowing out through the outlet faucet 22 can continue to pass through Procedures such as sample extraction, elution, drying, and reconstitution are used to detect antibiotic content in ballast water.

More specifically, the sample enrichment mechanism 3 includes: an enrichment bottle 31, a suction filter port 32, a filter head 33, a sand core filter plate 34 and a 0.22 μm nitrocellulose filter membrane 35;

The enrichment bottle 31 is a wide-mouth glass bottle in the shape of high borosilicate glass with a volume of 1L. The inside and outside of the mouth are frosted and equipped with a frosted stopper. The outside of the bottle mouth is frosted to facilitate sealing with the filter head 33. The bottle body There is a suction filter port 32 5cm outside the bottom, which is used to connect the suction filter hose and the motor. The thread of the filter head 33 is set on the top of the enrichment bottle 31. The inside of the filter head 33 is frosted. The top of the filter head 33 is set in a concentric circle. The sand core filter plate 34 is threaded in the inner cavity of the filter head 33, and the 0.22 μm nitrocellulose filter membrane 35 is laid on the top of the sand core filter plate 34. The outer diameter of the filter head 33 is equal to the outer edge diameter of the glass body 21, which is convenient for Assemble seal;

In order to strengthen the sealing between the filter head 33 and the glass body 21, the glass body 21 and the filter head 33 can be connected by threads, and a sealing ring is used between the glass body 21 and the filter head 33 to strengthen the overlapping and fix the seal. Effect;

Combine the sample filtration mechanism 2 and the sample enrichment mechanism 3. Place a 0.22 μm nitrocellulose filter membrane 35 on the sand core filter plate 34 of the filter head 33 of the sample enrichment structure 3. First pass the ballast water sample through the filter device to filter the sample. After the volume exceeds 200ml, remove the two-stage filtration structure and filter the sample through the motor. After the 200ml sample is filtered, dismantle the system. The 0.22μm nitrocellulose filter membrane 35 is used for the detection of ARGs in ballast water. The mouth of the wide-mouth bottle And the suction filter port is plugged and stored, and the water sample can be used for basic water quality testing.

The sterile refrigerator 4 is used for freezing and storing the 0.22 μm nitrocellulose filter membrane 35 after suction filtration, and the temperature is set to -20°C. A layered placement rack 5 for layered storage of petri dishes is provided inside, and the 0.22 μm nitrocellulose filter membrane 35 is placed. In a sterilized Petri dish, the Petri dish is placed in a sterile refrigerator and brought back to the laboratory, where DNA is extracted and used for detection of ARGs.

The detailed connection means are well-known technologies in this field. The following mainly introduces the working principle and process. The specific work is as follows.

The tightness of the equipment is tested by filtering sterile water, and the effectiveness of the equipment is tested by filtering surface water sampled in the field. Three types of samples are set up.

Place the first-stage filter 26 in the second-stage filter cartridge 24, and place the second-stage filter cartridge 24 in the glass body 21 of the filter structure;

Place the 0.22 μm nitrocellulose filter membrane 35 on the sand core filter plate 34 of the filter head 33 and moisten it with water;

Assemble and connect the filter head 33 and the enrichment bottle 31 to achieve sealing through the frosted structure;

Connect the glass body 21 to the filter head 33 connected to the enrichment bottle 31, and achieve sealing through a frosted structure. In order to strengthen the sealing between the filter head 33 and the glass body 21, the glass body 21 and the filter head 33 can be Threaded connection, and a sealing ring is used between the glass body 21 and the filter head 33 to enhance the fixed sealing effect after overlapping;

Connect the suction filter port 32 of the enrichment bottle 31 to the motor through a hose;

Sealing test of connected devices with sterile water;

After filtering through three different surface water samples, a DNA extraction kit was used to extract the DNA samples on the 0.22 μm nitrocellulose filter membrane 35 and test the concentration of sul1. The test results showed that the absolute concentrations of sul1 in the three samples were 2.71×10 ⁵ copies/mL, 2.18×10 ⁴ copies/mL and 2.16×10 ⁴ copies/mL respectively.

Although the embodiments of the present invention have been shown and described, those of ordinary skill in the art will understand that various changes and modifications can be made to these embodiments without departing from the principles and spirit of the present invention. , substitutions and modifications, the scope of the present invention is defined by the appended claims and their equivalents.

Claims (6)

1. The utility model provides an experimental system of ballast water gene sample preparation, its characterized in that, including supporting seat (1), sample filtering mechanism (2), sample enrichment mechanism (3), aseptic fridge (4) and layering rack (5), the top of supporting seat (1) is provided with sample filtering mechanism (2), sample enrichment mechanism (3) and aseptic fridge (4) from left to right movable in proper order, the setting that sample filtering mechanism (2) can dismantle is on the top of sample enrichment mechanism (3) to can reach sealedly, the inner chamber of aseptic fridge (4) is provided with layering rack (5).

2. An experimental system for ballast water gene sample preparation according to claim 1, wherein: the sample filtration mechanism (2) comprises:

a glass main body (21) made of high boron silicon and hollow at the upper and lower sides, wherein the diameter of the bottom edge of the glass main body (21) is larger than the hollow diameter;

the second-stage filter cylinder (24) is detachably arranged in the inner cavity of the glass main body (21), a second clamping ring (25) is fixedly arranged at the top end of the second-stage filter cylinder (24), and the second clamping ring (25) can be sleeved at the top end of the glass main body (21) and is used for fixing the second-stage filter cylinder (24) in the glass main body (21);

a 100 mu m filter membrane structure (23) in threaded connection with the bottom end of the second-stage filter cylinder (24);

the detachable first-stage filter screen (26) is arranged in the inner cavity of the second-stage filter cylinder (24), a first clamping ring (27) is fixedly arranged at the top end of the first-stage filter screen (26), and the first clamping ring (27) can be sleeved at the top end of a second clamping ring (25) and is used for fixing the first-stage filter screen (26) in the second-stage filter cylinder (24);

and a water outlet tap (22) arranged on the outer wall of the glass main body (21).

3. An experimental system for ballast water gene sample preparation according to claim 2, wherein: the diameter of the second stage filter cartridge (24) is smaller than the diameter of the glass body (21).

4. An experimental system for ballast water gene sample preparation according to claim 3, wherein: the first stage filter screen (26) is of a conical structure.

5. The experimental system for preparing a ballast water gene sample according to claim 4, wherein: the sample enrichment mechanism (3) comprises:

the wide-mouth glass bottle is an enrichment bottle (31) made of high borosilicate glass;

a suction filtration port (32) arranged on the outer wall of the enrichment bottle (31) and used for connecting a suction filtration hose and a motor;

a filter head (33) which is arranged at the top end of the enrichment bottle (31) in a threaded manner;

the sand core filter plate (34) is arranged in the inner cavity of the filter head (33) in a threaded manner, and the 0.22 mu m nitrocellulose filter membrane (35) is paved at the top end of the sand core filter plate (34).

6. The experimental system for preparing a ballast water gene sample according to claim 5, wherein: the diameter of the outer circle of the filter head (33) is equal to the diameter of the outer edge of the glass main body (21), so that the assembly and the sealing are convenient.