An environmental DNA extraction method and device

Abstract

The application provides an environmental DNA extraction method and device. The environmental DNA extraction device comprises an upper seat body, a lower seat body and a detachable membrane assembly. The upper seat body is a column with a sealed upper end and an open lower end. Tangential flow water inlets and tangential flow water outlets are arranged on the two side walls of the upper seat body. The upper end of the lower seat body is open, and the lower end of the lower seat body is provided with a filtrate outlet. The lower end of the upper seat body is connected to the upper end of the detachable membrane assembly, and the upper end of the lower seat body is connected to the lower end of the detachable membrane assembly. The detachable membrane assembly is arranged between the upper seat body and the lower seat body. Environmental water flows into the detachable membrane assembly through the tangential flow water inlets. The detachable membrane assembly filters and adsorbs the environmental water to adsorb environmental DNA in the environmental water on the detachable membrane assembly, and the remaining filtrate flows out through the filtrate outlet of the lower seat body. The extraction process can be simplified, the time consumption can be reduced, the risk of DNA degradation and sample pollution can be avoided, and the DNA extraction quality can be improved.

Description

Technical Field

[0001] This invention relates to the field of environmental monitoring technology, and more specifically, to a method and apparatus for extracting environmental DNA. Background Technology

[0002] Environmental DNA extraction technology has wide applications in ecological monitoring, species detection, and biodiversity assessment. Traditional environmental DNA extraction methods include filtration, centrifugation, and adsorption column methods. However, these methods typically rely on complex reagents and expensive equipment in laboratories, resulting in complex and time-consuming extraction processes, as well as risks of DNA degradation and sample contamination.

[0003] In addition, existing environmental DNA extraction methods are not very efficient at extracting low concentrations of environmental DNA, especially in aquatic environments. Low concentrations of environmental DNA are more easily interfered with by inhibitors such as humic acid and metal ions in the environment, resulting in poor quality of extracted environmental DNA and affecting subsequent detection and analysis.

[0004] Therefore, developing a simple, efficient, and field-suitable method for environmental DNA extraction is a pressing issue that this application aims to address. Summary of the Invention

[0005] In view of this, the present invention provides an environmental DNA extraction method and apparatus that simplifies the extraction process, reduces time consumption, avoids the risk of DNA degradation and sample contamination, and improves the quality of DNA extraction.

[0006] The first aspect of this application provides an environmental DNA extraction device, which includes an upper seat, a lower seat, and a detachable membrane assembly;

[0007] The upper body is a column with a closed upper end and an open lower end. Tangential flow inlet and tangential flow outlet are respectively provided on the two side walls of the upper body.

[0008] The lower seat has an opening at its upper end, and a filtrate outlet is provided at its lower end.

[0009] The lower end of the upper seat is connected to the upper end of the detachable membrane assembly, and the upper end of the lower seat is connected to the lower end of the detachable membrane assembly, so that the detachable membrane assembly is disposed between the upper seat and the lower seat.

[0010] Optionally, the removable membrane assembly includes a ceramic filter membrane and an adsorption assembly;

[0011] The ceramic filter membrane is connected to the lower end of the upper body as the upper end of the detachable membrane assembly;

[0012] The adsorption component is connected to the upper end of the lower seat as the lower end of the detachable membrane component; wherein the adsorption component contains salt-activated broken glass fiber particles.

[0013] Optionally, the adsorption component includes a polyester fiber mesh wrapping layer and a salt-activated glass fiber adsorption layer;

[0014] The polyester fiber mesh wrapping layer is used to wrap the salt-activated glass fiber adsorption layer; wherein, the salt-activated glass fiber adsorption layer is composed of salt-activated broken glass fiber particles;

[0015] The polyester fiber mesh wrapping layer serves as the lower end of the detachable membrane assembly and is connected to the upper end of the lower seat.

[0016] Optionally, the environmental DNA extraction device further includes a ring clamp fixing assembly;

[0017] The annular clamp fixing assembly is disposed in the gap between the upper seat and the lower seat, and is used to fix the detachable membrane assembly between the upper seat and the lower seat.

[0018] Optionally, the upper seat and the lower seat have their tube openings flush and fit together, and the outer and inner diameters of the upper seat are the same as those of the lower seat.

[0019] Optionally, a pump interface is also integrated on the side wall of the upper body;

[0020] The peristaltic pump interface is connected to an external power supply and pump to integrate the power supply and peristaltic pump into a single device.

[0021] Optionally, the pore size of the ceramic filter membrane is within a first preset range.

[0022] Optionally, the polyester fiber mesh wrapping layer is a sealed interlayer formed by two layers of polyester mesh.

[0023] Optionally, the area of ​​the salt-activated glass fiber adsorption layer is smaller than the area of ​​the polyester fiber mesh wrapping layer, and the ratio of the area of ​​the salt-activated glass fiber adsorption layer to the area of ​​the polyester fiber mesh wrapping layer is within a second preset range.

[0024] A second aspect of this application provides an environmental DNA extraction method, applied to the environmental DNA extraction apparatus provided in the first aspect of this application, the method comprising:

[0025] Ambient water is tangentially flowed into the removable membrane assembly through the tangential inlet of the upper body;

[0026] The environmental water is filtered and adsorbed through the detachable membrane assembly to adsorb environmental DNA from the environmental water onto the detachable module, and the remaining filtrate flows out through the filtrate outlet of the lower body.

[0027] This application provides an environmental DNA extraction method and apparatus. The environmental DNA extraction apparatus includes an upper body, a lower body, and a detachable membrane assembly. The upper body is a cylindrical body with a sealed upper end and an open lower end. A tangential inlet and a tangential outlet are respectively provided on the two side walls of the upper body. The lower body has an open upper end and a filtrate outlet at its lower end. The lower end of the upper body is connected to the upper end of the detachable membrane assembly, and the upper end of the lower body is connected to the lower end of the detachable membrane assembly, so that the detachable membrane assembly is disposed between the upper body and the lower body. Therefore, this application utilizes an upper body with a tangential inlet to allow environmental water to flow into a detachable membrane module, effectively preventing membrane clogging by utilizing shear force. The detachable membrane module filters and adsorbs the incoming environmental water, adsorbing environmental DNA onto the detachable module, and the remaining filtrate flows out through the filtrate outlet of the lower body. This eliminates the need for complex reagents and expensive equipment in the laboratory, simplifying the extraction process, reducing time consumption, and avoiding the risks of DNA degradation and sample contamination. Furthermore, the environmental DNA extraction device provided by this application can efficiently extract low-concentration environmental DNA, ensuring the quality of the extracted environmental DNA and preventing any impact on subsequent detection and analysis. Attached Figure Description

[0028] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0029] Figure 1 This is a schematic diagram of the external structure of an environmental DNA extraction device provided in an embodiment of this application;

[0030] Figure 2 This is a cross-sectional structural diagram of an environmental DNA extraction device provided in an embodiment of this application;

[0031] Figure 3 This is a schematic diagram of the external structure of another environmental DNA extraction device provided in an embodiment of this application;

[0032] Figure 4 This is a schematic diagram of the external structure of another environmental DNA extraction device provided in an embodiment of this application;

[0033] Figure 5This is a schematic diagram of the external structure of another environmental DNA extraction device provided in an embodiment of this application;

[0034] Figure 6 A schematic flowchart of an environmental DNA extraction method provided in this application embodiment;

[0035] Figure 7 A schematic diagram of a process for extracting DNA from tap water using an environmental DNA extraction device, provided for an embodiment of this application;

[0036] Figure 8 This is a schematic diagram illustrating the extraction of DNA from natural lake water using an environmental DNA extraction device, as provided in an embodiment of this application. Detailed Implementation

[0037] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0038] In this application, relational terms such as "first" and "second" are used merely to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes the element.

[0039] See Figure 1 This application illustrates an environmental DNA extraction device provided in an embodiment of the present application. The environmental DNA extraction device includes an upper seat 1, a lower seat 2, and a detachable membrane assembly 3 that also has filtering and adsorption functions.

[0040] The upper body 1 is a column with a closed upper end and an open lower end. Tangential flow inlet 4 and tangential flow outlet 5 are respectively provided on the two side walls of the upper body.

[0041] The lower body 2 has an opening at the upper end and a filtrate outlet 6 at the lower end.

[0042] The lower end of the upper seat 1 is connected to the upper end of the detachable membrane assembly 3, and the upper end of the lower seat 2 is connected to the lower end of the detachable membrane assembly 3, so that the detachable membrane assembly 3 is disposed between the upper seat 1 and the lower seat 2.

[0043] It should be noted that, in order to better understand the internal structure of the environmental DNA extraction device provided in this application, the embodiments of this application also provide a cross-sectional view of the environmental DNA extraction device, such as... Figure 2 As shown.

[0044] In this embodiment, the tangential inlet of the upper body allows ambient water to flow tangentially into the detachable membrane module. The detachable membrane module filters and adsorbs the ambient water to adsorb environmental DNA from the ambient water onto the detachable module, and the remaining filtrate flows out through the filtrate outlet of the lower body. Therefore, this application utilizes an upper body with a tangential inlet to allow ambient water to flow into the detachable membrane module, effectively using shear force to prevent membrane clogging. Filtering and adsorbing the incoming ambient water through the detachable membrane module eliminates the need for complex reagents and expensive equipment in a laboratory, simplifying the extraction process, reducing time consumption, and avoiding the risks of DNA degradation and sample contamination. Furthermore, the environmental DNA extraction device provided in this application can efficiently extract low-concentration environmental DNA, preventing low-quality extracted environmental DNA and thus avoiding impacts on subsequent detection and analysis.

[0045] In some embodiments, the upper body is a cylindrical tube with a sealed upper end and an open lower end, or a rectangular column with a sealed upper end and an open lower end, or a normal column with a sealed upper end and an open lower end; the lower body is a cylindrical tube with an outlet at the lower end of the upper body opening.

[0046] It should be noted that the water inlet configured on the side wall of the upper body is designed as a tangential flow in order to allow ambient water to flow into the removable membrane module through a tangential flow after passing through the tangential flow inlet, thereby reducing particle deposition by utilizing shear force.

[0047] It should also be noted that the advantages of tangential flow inlet and outlet compared to traditional vertical flow inlet and outlet are as follows: In traditional vertical flow, the inlet is at the top and the outlet is at the bottom, perpendicular to the filter membrane; while in tangential flow, the outlet and inlet are horizontal, parallel to the filter membrane, which can utilize shear force to reduce particle deposition.

[0048] In some embodiments, the openings of the upper and lower seats are flush and fit together, and the outer and inner diameters of the upper and lower seats are the same, so that the upper and lower seats are matched.

[0049] Optionally, the side wall of the upper body provided in this application embodiment may also integrate a pump interface; the peristaltic pump interface is externally connected to the pump, so as to integrate the power supply and the pump into a single device.

[0050] As one implementation of this application, the power supply can be an external portable power supply, and the pump can be a peristaltic pump. Accordingly, the peristaltic pump interface integrated on the side wall of the upper body can be connected to an external portable power supply and a peristaltic pump, so as to integrate the power supply, portable power supply and peristaltic pump into an integrated device.

[0051] It should be noted that the peristaltic pump pumps environmental water into the environmental DNA extraction device through the tangential inlet, allowing a large amount of environmental water to pass through the detachable module, where the environmental DNA is adsorbed onto the detachable module. Since the inlet is tangential, power is required to allow the environmental water to enter the environmental DNA extraction device. Furthermore, due to the fine mesh of the detachable membrane module, water flow is difficult to pass through the membrane without external power.

[0052] Optional, combined Figure 1 See Figure 3 The environmental DNA extraction device also includes a ring clamp fixing component 7; the ring clamp fixing component 7 is disposed in the gap between the upper body and the lower body, and is used to fix the detachable membrane component between the upper body and the lower body.

[0053] It should be noted that a ring-shaped clamp fixing component is installed at the gap between the upper and lower seats, which can firmly fix the detachable membrane assembly between the upper and lower seats.

[0054] Optional, combined Figure 3 See Figure 4 This application provides a detachable membrane module, a ceramic filter membrane 3-1 and an adsorption module 3-2; the ceramic filter membrane is connected to the lower end of the upper seat as the upper end of the detachable membrane module; the adsorption module is connected to the upper end of the lower seat as the lower end of the detachable membrane module; wherein, the adsorption module contains salt-activated broken glass fiber particles.

[0055] It should be noted that broken glass fiber particles can effectively increase the specific surface area of ​​the adsorption module, thereby increasing DNA adsorption within a limited space. High-concentration salt solutions can further promote DNA adsorption.

[0056] In some embodiments, the removable membrane assembly can be a standardized, replaceable component, and the size of the removable membrane assembly matches the size of the upper and lower bodies, thereby improving the assembly and disassembly efficiency of the environmental DNA extraction device.

[0057] It should be noted that the ceramic filter membrane has multiple pores, and the pore size of the ceramic filter membrane can be within a first preset range, which can be 10-100nm, for filtering and removing suspended solids, colloids and other large particulate matter from environmental water.

[0058] In some embodiments, ambient water passes through a tangential inlet and undergoes initial filtration via a ceramic membrane, trapping large particulate impurities on its surface. Subsequent ambient water then flows in tangentially, utilizing shear force to remove these impurities and prevent clogging of the adsorption assembly. The filtered filtrate permeates the ceramic membrane into the adsorption assembly, where it adsorbs the incoming filtrate, retaining environmental DNA for subsequent DNA extraction.

[0059] Optional, combined Figure 4 See Figure 5 The adsorption component includes a polyester fiber mesh wrapping layer 3-2-1 and a salt-activated glass fiber adsorption layer 3-2-2; the polyester fiber mesh wrapping layer is used to wrap the salt-activated glass fiber adsorption layer, wherein the salt-activated glass fiber adsorption layer is composed of salt-activated broken glass fiber particles; the polyester fiber mesh wrapping layer is connected to the upper end of the lower seat body as the lower end of the detachable membrane component.

[0060] In some embodiments, the polyester fiber web wrapping layer is a sealed interlayer formed by two layers of polyester web.

[0061] It should be noted that the polyester fiber mesh wrapping layer is a sealed interlayer formed by two layers of polyester mesh, with the edges bonded to prevent particle leakage, and the thickness of the interlayer is 3-5 times that of a single layer of mesh.

[0062] Optionally, the area of ​​the salt-activated glass fiber adsorption layer is smaller than the area of ​​the polyester fiber mesh wrapping layer, and the ratio of the area of ​​the salt-activated glass fiber adsorption layer to the area of ​​the polyester fiber mesh wrapping layer is within a second preset range.

[0063] It should be noted that the second preset range can be 0.7-0.8, which means that the area of ​​the salt-activated glass fiber adsorption layer is 70%-80% of that of the polyester fiber mesh wrapping layer, and the coating thickness is 3-5 times the thickness of a single layer of polyester fiber mesh.

[0064] In the embodiments of this application, the salt-activated glass fiber adsorption layer can be broken glass fiber particles activated by a 1.0 M sterile NaCl solution. Through salt induction, its ability to bind to DNA can be significantly enhanced, so as to efficiently adsorb environmental DNA filtrate from filtered environmental water.

[0065] In some embodiments, after the ambient water flows through the ceramic filter membrane via the tangential inlet, large particulate impurities in the ambient water can be removed. The filtered filtrate penetrates the ceramic filter membrane and enters the salt-activated glass fiber adsorption layer wrapped by the polyester fiber mesh membrane. Under the induction of 1.0 M sterile NaCl solution, the environmental DNA in the filtrate is efficiently adsorbed, so that the environmental DNA is adsorbed onto the salt-activated broken glass fiber particles in the salt-activated glass fiber adsorption layer. The salt-activated broken glass fiber particles with adsorbed environmental DNA are transferred to centrifuge tubes so that a low concentration of salt solution can be added to elute and release the DNA or to extract the environmental DNA from the environmental DNA filtrate using DNA extraction reagents.

[0066] Based on the environmental DNA extraction device provided in the above embodiments of this application, correspondingly, the embodiments of this application provide an environmental DNA extraction method, such as... Figure 6 As shown, the method includes the following steps:

[0067] S601: Ambient water is tangentially flowed into the removable membrane module through the tangential inlet of the upper body.

[0068] In this embodiment, a peristaltic pump interface is integrated on the side wall of the upper body, allowing for the connection of a portable power supply and a peristaltic pump, thus integrating the power supply, portable power supply, and peristaltic pump into a single device. The peristaltic pump pumps environmental water into the environmental DNA extraction device through the tangential inlet, allowing a large amount of environmental water to pass through the removable membrane assembly.

[0069] S602: Filters and adsorbs environmental water through a detachable membrane module to adsorb environmental DNA in the environmental water onto the detachable module, and discharges the remaining filtrate through the filtrate outlet of the lower body.

[0070] In this embodiment, the detachable membrane assembly includes a ceramic filter membrane and an adsorption assembly. The adsorption assembly includes a polyester fiber mesh wrapping layer and a salt-activated glass fiber adsorption layer, which is composed of salt-activated broken glass fiber particles. After the ambient water flows through the ceramic filter membrane via the tangential inlet, large particulate impurities in the ambient water can be removed. The filtered filtrate penetrates the ceramic filter membrane and enters the salt-activated glass fiber adsorption layer wrapped by the polyester fiber mesh wrapping layer. Under the induction of 1.0M sterile NaCl solution, the environmental DNA in the filtrate is efficiently adsorbed, so that the environmental DNA is adsorbed onto the salt-activated broken glass fiber particles in the salt-activated glass fiber adsorption layer. The salt-activated broken glass fiber particles adsorbed with environmental DNA are transferred to a centrifuge tube so that a low-concentration salt solution can be added to elute and release the DNA or to extract the environmental DNA from the environmental DNA filtrate using a DNA extraction reagent.

[0071] In summary, firstly, the water inlet on the upper body of the environmental DNA extraction device provided in this application adopts a tangential flow structure design. Compared with traditional vertical filtration, the tangential force of the water flow can effectively flush away impurities in the environmental water attached to the membrane surface, thereby avoiding the accumulation and clogging of large particles, slowing down the rate of membrane fouling, and extending the service life of the membrane.

[0072] Secondly, the detachable membrane component in the environmental DNA extraction device provided in this application uses a salt-activated glass fiber adsorption layer containing broken glass fiber particles, which can effectively increase the specific surface area of ​​the DNA adsorption layer. At the same time, the glass fiber activated by 1.0 M sterile NaCl solution significantly improves the DNA adsorption rate, thereby achieving efficient enrichment and extraction of DNA in environmental water.

[0073] Then, the upper body of the environmental DNA extraction device provided in this application is integrated with a peristaltic pump interface, which supports the integration of an external portable power supply and a peristaltic pump into an integrated device, thereby realizing rapid sampling and DNA enrichment extraction in the field water environment.

[0074] Furthermore, the detachable membrane component in the environmental DNA extraction device provided in this application is a standardized and replaceable component, which is not only easy to install and disassemble, but also allows for integrated structure and mass production, facilitating the replacement and use of the membrane component during field sampling.

[0075] Furthermore, the materials selected for each structure in the environmental DNA extraction device provided in this application are inexpensive and readily available, and all possess good chemical stability and biocompatibility. They can withstand the chemical erosion of various environmental water samples and will not adsorb or degrade the environmental DNA, thus maximizing the integrity and extraction purity of the environmental DNA.

[0076] Finally, during the installation and fixing process of the environmental DNA extraction device provided in this application, it should be ensured that the device and the filter assembly are tightly connected, and that the filter membrane is free from wrinkles and displacement to prevent water sample leakage and thus affect the filtration effect.

[0077] To better understand the environmental DNA extraction device provided in the embodiments of this application, the following description uses examples:

[0078] Example 1: DNA was extracted from tap water using the environmental DNA extraction device provided in this application embodiment, such as... Figure 7 As shown, 10 is a peristaltic pump, 11 is an environmental DNA extraction device, 12 is a tap water collection tank, and 13 is a filtrate collection tank. The specific process is as follows:

[0079] First, 0.1 g of glass fiber filter paper was mixed with 10.0 ml of 1.0 M sodium chloride buffer solution. Zirconia microspheres with a diameter of 5.0 mm were added and the mixture was thoroughly broken up in a tissue homogenizer. The mixture was shaken at a frequency of 25 Hz for 1 min to obtain broken glass fiber particles in the salt-activated glass fiber adsorption layer. The salt-activated glass fiber adsorption layer containing glass fiber particles was uniformly dispersed in a polyester fiber mesh wrapping layer and prepared together with a ceramic filter membrane to form a DNA enrichment membrane module (detachable membrane module).

[0080] Secondly, a ring clamp is used to fix the detachable membrane assembly in the gap between the upper and lower bodies of the environmental DNA extraction device. The inlet pipe and outlet pipe are connected to the tangential inlet and outlet pipes respectively. A peristaltic pump is connected between the inlet pipe and the tap water sample collection bucket.

[0081] Finally, under the action of the peristaltic pump, tap water is filtered and DNA enriched through the detachable membrane module; after 5 minutes, the peristaltic pump is turned off to stop filtration, and the broken glass fiber particles in the water environment DNA enrichment device are transferred to a centrifuge tube, where the broken glass fiber particles at this time are adsorbed with environmental DNA from the tap water; 100 μl of DNA extraction reagent is added to the offline tube, heated at 95°C for 10 minutes, and centrifuged at 3000 rpm for 3 minutes. The supernatant is the solution containing environmental DNA, which can be used as a DNA template solution for PCR reaction for DNA amplification and subsequent DNA detection.

[0082] Example 2: Using the environmental DNA extraction device provided in this application, environmental DNA was extracted from natural lake water. Figure 8 As shown, 10 is a peristaltic pump, 11 is an environmental DNA extraction device, 12 is a tap water collection tank, 13 is a filtrate collection tank, 14 is a boat used for DNA extraction from lake water, and 15 is natural lake water. The specific process is as follows:

[0083] First, 0.3 g of glass fiber filter paper was mixed with 30.0 ml of 2.0 M sodium chloride buffer solution. Zirconia microspheres with a diameter of 5.0 mm were added and the mixture was thoroughly broken up in a tissue homogenizer. The mixture was shaken at a frequency of 25 Hz for 3 min to obtain broken glass fiber particles in the salt-activated glass fiber adsorption layer. The salt-activated glass fiber adsorption layer containing glass fiber particles was uniformly dispersed in a polyester fiber mesh wrapper and prepared together with a ceramic filter membrane to form a DNA enrichment membrane module (removable membrane module).

[0084] Secondly, a ring clamp is used to fix the filter membrane assembly in the gap between the upper and lower bodies of the environmental DNA extraction device. The inlet pipe and outlet pipe are connected at the tangential inlet and outlet respectively. A peristaltic pump is connected between the inlet pipe and the natural water sample collection point.

[0085] Finally, under the action of the peristaltic pump, the natural water sample was filtered and DNA enriched through the detachable membrane module; after 8 minutes, the peristaltic pump was turned off to stop filtration, and the broken glass fiber particles in the water environment DNA enrichment device were transferred to centrifuge tubes, where the environmental DNA of the natural water sample was adsorbed on the broken glass fiber particles at this time; 0.2 M sodium chloride solution was added to the centrifuge tubes to elute the environmental DNA adsorbed on the broken glass fiber particles.

[0086] Example 3: This example uses the environmental DNA extraction device provided in the embodiments of this application and a traditional vertical flow device to filter natural lake water and extract environmental DNA. The specific process is as follows:

[0087] The environmental DNA extraction device provided in this application embodiment and the traditional vertical flow device are respectively equipped with the DNA membrane assembly (detachable membrane assembly) provided in this application embodiment. The inlet pipe and outlet pipe are respectively connected to the inlet and outlet of the two devices. A peristaltic pump is connected between the inlet pipe and the natural water sample collection point. The natural water sample is filtered and DNA enriched through the detachable membrane assembly.

[0088] Using peristaltic pumps of the same model from the same manufacturer and with the same flow rate, the initial flow rate Q0 was recorded during the experiment, and the real-time flow rate Q was recorded every 2 minutes. t The membrane fouling rate is: (1-Q) t / Q0)×100%. After 8 min, the peristaltic pump was turned off to stop filtration. The removable membrane module was removed to observe its degree of fouling and calculate its clogging rate. The salt-activated glass fiber adsorption layer of the removable membrane module was placed in 0.2 M NaCl solution for DNA elution. The amount of DNA recovered was detected by real-time PCR. The corresponding experimental results are shown in Table 1.

[0089] As can be seen from the experimental results shown in Table 1, the environmental DNA extraction device provided by the present invention has better membrane module anti-clogging performance and DNA enrichment and extraction effect compared with the traditional vertical flow device.

[0090] Table 1. Comparison of environmental DNA extraction results from the two devices

[0091]

[0092] The various embodiments in this specification are described in a progressive manner. Similar or identical parts between embodiments can be referred to mutually. Each embodiment focuses on describing the differences from other embodiments. In particular, for system or system embodiments, since they are basically similar to method embodiments, the description is relatively simple, and relevant parts can be referred to the descriptions in the method embodiments. The systems and system embodiments described above are merely illustrative. The units described as separate components may or may not be physically separate. The components shown as units may or may not be physical units; that is, they may be located in one place or distributed across multiple network units. Some or all of the modules can be selected to achieve the purpose of this embodiment according to actual needs. Those skilled in the art can understand and implement this without creative effort.

[0093] Those skilled in the art will further recognize that the units and algorithm steps of the various examples described in conjunction with the embodiments disclosed herein can be implemented in electronic hardware, computer software, or a combination of both. To clearly illustrate the interchangeability of hardware and software, the components and steps of the various examples have been generally described in terms of functionality in the foregoing description. Whether these functions are implemented in hardware or software depends on the specific application and design constraints of the technical solution. Those skilled in the art can use different methods to implement the described functions for each specific application, but such implementations should not be considered beyond the scope of this invention.

[0094] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

[0095] The above are merely preferred embodiments of the present invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. An environmental DNA extraction device, characterized in that, The environmental DNA extraction device includes an upper seat, a lower seat, and a detachable membrane assembly; The upper body is a column with a closed upper end and an open lower end. Tangential flow inlet and tangential flow outlet are respectively provided on the two side walls of the upper body. The lower seat has an opening at its upper end, and a filtrate outlet is provided at its lower end. The lower end of the upper seat is connected to the upper end of the detachable membrane assembly, and the upper end of the lower seat is connected to the lower end of the detachable membrane assembly, so that the detachable membrane assembly is disposed between the upper seat and the lower seat.

2. The environmental DNA extraction device according to claim 1, characterized in that, The detachable membrane assembly includes a ceramic filter membrane and an adsorption assembly; The ceramic filter membrane is connected to the lower end of the upper body as the upper end of the detachable membrane assembly; The adsorption component is connected to the upper end of the lower seat as the lower end of the detachable membrane component; wherein the adsorption component contains salt-activated broken glass fiber particles.

3. The environmental DNA extraction device according to claim 2, characterized in that, The adsorption component includes a polyester fiber mesh wrapping layer and a salt-activated glass fiber adsorption layer; The polyester fiber mesh wrapping layer is used to wrap the salt-activated glass fiber adsorption layer; wherein, the salt-activated glass fiber adsorption layer is composed of salt-activated broken glass fiber particles; The polyester fiber mesh wrapping layer serves as the lower end of the detachable membrane assembly and is connected to the upper end of the lower seat.

4. The environmental DNA extraction device according to claim 1, characterized in that, The environmental DNA extraction device also includes a ring clamp fixing assembly; The annular clamp fixing assembly is disposed in the gap between the upper seat and the lower seat, and is used to fix the detachable membrane assembly between the upper seat and the lower seat.

5. The environmental DNA extraction device according to claim 1, characterized in that, The upper seat and the lower seat have their tube openings flush and fit together, and the outer and inner diameters of the upper seat are the same as those of the lower seat.

6. The environmental DNA extraction device according to claim 1, characterized in that, The side wall of the upper body also integrates a pump interface; The peristaltic pump interface is connected to an external power supply and a pump, so as to integrate the power supply and the pump into a single device.

7. The environmental DNA extraction device according to claim 2, characterized in that, The pore size of the ceramic filter membrane is within a first preset range.

8. The environmental DNA extraction device according to claim 3, characterized in that, The polyester fiber mesh wrapping layer is a sealed interlayer formed by two layers of polyester mesh.

9. The environmental DNA extraction device according to claim 3, characterized in that, The area of ​​the salt-activated glass fiber adsorption layer is smaller than the area of ​​the polyester fiber mesh wrapping layer, and the ratio of the area of ​​the salt-activated glass fiber adsorption layer to the area of ​​the polyester fiber mesh wrapping layer is within a second preset range.

10. A method for extracting environmental DNA, characterized in that, The method, applied to the environmental DNA extraction apparatus according to any one of claims 1-9, comprises: Ambient water is tangentially flowed into the removable membrane module through the tangential inlet of the upper body; The environmental water is filtered and adsorbed through the detachable membrane assembly to adsorb environmental DNA from the environmental water onto the detachable module, and the remaining filtrate flows out through the filtrate outlet of the lower body.

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