CN111483096B

CN111483096B - Rhizosphere channel micro-fluidic chip, colloidal block thereof and mold for manufacturing colloidal block

Info

Publication number: CN111483096B
Application number: CN201911257089.9A
Authority: CN
Inventors: 祝贞科; 葛体达; 吴金水; 王季斐; 魏晓梦; 魏亮; 袁红朝; 王久荣
Original assignee: Institute of Subtropical Agriculture of CAS
Current assignee: Institute of Subtropical Agriculture of CAS
Priority date: 2019-12-10
Filing date: 2019-12-10
Publication date: 2022-04-08
Anticipated expiration: 2039-12-10
Also published as: CN111483096A

Abstract

The invention discloses a colloid block, which comprises a body, wherein a channel groove is arranged on the body, the invention also provides a mold for manufacturing the colloid block, the mold is used for manufacturing the colloid block, in addition, the invention also provides a rhizosphere channel micro-fluidic chip containing the colloid block, the micro-fluidic chip comprises a glass substrate and the colloid block which are connected, the channel groove is arranged towards the glass substrate, and a rhizosphere channel is formed in a space surrounded by the channel groove and the glass substrate. When the rhizosphere channel microfluidic chip is used for researching the interaction process of microorganism-plant micro-interfaces, a polyethylene tube is inserted into the rhizosphere channel, plants are planted, a microorganism solution is introduced into the rhizosphere channel, after specific culture, a fluorescence microscope is carried out to observe the microorganism activity of a sample, and a confocal microscope qualitatively or quantitatively detects the active enzyme of the sample on the chip.

Description

Rhizosphere channel micro-fluidic chip, colloidal block thereof and mold for manufacturing colloidal block

Technical Field

The invention relates to the technical field of rhizosphere microorganism research and peripheral supporting facilities thereof, in particular to a rhizosphere channel microfluidic chip, a colloidal block thereof and a mold for manufacturing the colloidal block.

Background

The soil environment is an aggregate of various micro-interfaces such as soil components, plant roots, soil organisms, soil solutions and the like. The rhizosphere microorganisms are various in types, large in quantity and rich in diversity, and influence on an ecosystem is different. Most rhizosphere microorganisms need to establish a parasitic, symbiotic (symbiotic) relationship with plants in order to take up host plant photosynthetic products. The rhizosphere microorganisms and plants have competition for nutrients and space, are interdependent and promote growth mutually. At present, many scholars research rhizosphere microorganisms and plants, but many research methods are to sample roots and soil and then bring the sampled roots and soil back to a laboratory for research. Although the treatment method discloses the general situation of rhizosphere microorganisms-plants, in the soil environment microorganism-plant rhizosphere interface, dark areas and hot areas with the action of microorganisms are formed by the contact distance between key elements and microorganisms, and the hot areas are ignored by the current research institute of homogenization and homogenization.

The micro-fluidic chip technology is widely applied to the research of micro-scale environment and biological interaction as a micro-processing and micro-operation means. Currently, microfluidics technology has been applied to a variety of research fields, including chemical synthesis, DNA analysis, proteomics, single cell analysis, tissue engineering, high throughput screening, environmental analysis and medical diagnosis, however, the application of microfluidics technology in soil micro-ecology is still in the first stage, and the research on the micro-process of related rhizosphere microorganism-plant interaction is rarely reported.

Therefore, how to construct a microfluidic chip to study the rhizosphere microorganism-plant micro-interface interaction process becomes a problem to be solved urgently by those skilled in the art.

Disclosure of Invention

The invention aims to provide a rhizosphere channel microfluidic chip, a colloid block thereof and a mold for manufacturing the colloid block, so as to solve the problems in the prior art, and enable researchers to use the microfluidic chip to realize in-situ observation of the plant growth process so as to research the rhizosphere microorganism-plant micro-interface interaction process.

In order to achieve the purpose, the invention provides the following scheme: the invention provides a colloid block which comprises a body, wherein channel grooves are formed in the body and comprise cylindrical grooves and prism grooves, the prism grooves are communicated with the cylindrical grooves, the prism grooves are uniformly distributed in the circumferential direction around the axis of the cylindrical grooves, terminal grooves are further formed in one ends, far away from the cylindrical grooves, of the prism grooves, and the terminal grooves extend to the edge of the body.

Preferably, the body is a regular pentagonal prism structure, the cylindrical grooves and the colloid blocks are coaxially arranged, and the number of the prismatic grooves is five.

Preferably, the diameter of the cylindrical groove is 2.8mm, the prismatic groove is hexagonal, the longest diagonal of the cross section of the prismatic groove is 7.9mm, the distance between two planes parallel to the longest diagonal of the prismatic groove is 1.8mm, and the obtuse included angle between two adjacent inner side walls of the prismatic groove is 130 °.

Preferably, the channel groove further comprises a communication groove, the prism groove is communicated with the cylinder groove by the communication groove, and the number of the communication grooves is consistent with that of the prism grooves and corresponds to that of the prism grooves one to one.

Preferably, the length of each of the communication groove and the terminal groove is 1.6mm, and the width of each of the communication groove and the terminal groove is 1.2 mm.

The invention also provides a rhizosphere channel microfluidic chip which utilizes the colloid block and comprises a glass substrate and the colloid block which are connected, wherein the channel groove is arranged towards the glass substrate, and a rhizosphere channel is formed in a space formed by the channel groove and the glass substrate.

Preferably, the glass substrate is bonded and connected with the colloid block.

The invention also provides a die for manufacturing the colloidal block, which comprises a base, a silicon wafer and a fixed frame, wherein the base is arranged at the bottom of the silicon wafer, the fixed frame is arranged at the top of the silicon wafer, the fixed frame can fix the relative positions of the silicon wafer and the base, the fixed frame is of a hollow structure, a cavity formed by the base, the fixed frame and the silicon wafer in a surrounding mode forms a forming cavity, a convex block is arranged at the top of the silicon wafer, and the shape of the convex block is matched with that of the channel groove.

Preferably, the base is provided with a positioning groove, the silicon chip is arranged in the positioning groove, the base is further provided with a connecting column, the fixing frame is provided with a connecting hole matched with the connecting column, and the connecting column penetrates through the connecting hole and is fixed through a fastening nut.

Preferably, the number of the convex blocks is multiple, and the convex blocks are uniformly distributed at equal intervals.

Compared with the prior art, the invention has the following technical effects: the colloid block comprises a body, wherein the body is provided with channel grooves, the channel grooves comprise cylindrical grooves and prismatic grooves, the prismatic grooves are communicated with the cylindrical grooves, the prismatic grooves are uniformly distributed in the circumferential direction around the axis of the cylindrical grooves, one ends of the prismatic grooves, far away from the cylindrical grooves, are also provided with terminal grooves, and the terminal grooves extend to the edge of the body. The invention also provides a die for manufacturing the colloidal block, which comprises a base, a silicon wafer and a fixed frame, wherein the base is arranged at the bottom of the silicon wafer, the fixed frame is arranged at the top of the silicon wafer, the fixed frame can fix the relative position of the silicon wafer and the base, the fixed frame is of a hollow structure, a cavity surrounded by the base, the fixed frame and the silicon wafer forms a molding cavity, a bump is arranged at the top of the silicon wafer, and the shape of the bump is matched with that of the channel groove. The invention also provides a rhizosphere channel microfluidic chip containing the colloid block, which comprises a glass substrate and the colloid block which are connected, wherein the channel groove is arranged towards the glass substrate, and a rhizosphere channel is formed in a space surrounded by the channel groove and the glass substrate. When the rhizosphere channel microfluidic chip is used for researching the interaction process of microorganism-plant micro-interfaces, a polyethylene tube is inserted into the rhizosphere channel, plants are planted, a microorganism solution is introduced into the rhizosphere channel, after specific culture, a fluorescence microscope is carried out to observe the microorganism activity of a sample, and a confocal microscope qualitatively or quantitatively detects the active enzyme of the sample on the chip.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings needed to be used in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings without inventive exercise.

FIG. 1 is a schematic diagram of the construction of the colloidal block of the present invention;

FIG. 2 is a schematic structural view of the mold for producing the colloidal block of the present invention;

FIG. 3 is a schematic structural diagram of a rhizosphere channel microfluidic chip according to the present invention;

wherein, 1 is the body, 2 is the cylinder recess, 3 is prismatic recess, 4 is the terminal recess, 5 is the intercommunication recess, 6 is the base, 7 is the silicon chip, 8 is the lug, 9 is the mount, 10 is the spliced pole, 11 is the glass substrate, alpha is the obtuse angle contained angle between two adjacent inside walls of prismatic recess.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

In order to make the aforementioned objects, features and advantages of the present invention comprehensible, embodiments accompanied with figures are described in further detail below.

Referring to fig. 1-3, fig. 1 is a schematic structural diagram of a colloidal block of the present invention, fig. 2 is a schematic structural diagram of a mold for manufacturing a colloidal block of the present invention, and fig. 3 is a schematic structural diagram of a rhizosphere channel microfluidic chip of the present invention.

The invention provides a colloid block which comprises a body 1, wherein channel grooves are formed in the body 1 and comprise cylindrical grooves 2 and prism grooves 3, the prism grooves 3 are communicated with the cylindrical grooves 2, a plurality of prism grooves 3 are uniformly distributed in the circumferential direction around the axis of the cylindrical grooves 2, terminal grooves 4 are further formed in one ends, far away from the cylindrical grooves 2, of the prism grooves 3, and the terminal grooves 4 extend to the edge of the body 1. The colloidal block is made of polydimethylsiloxane, a body 1 of the colloidal block is provided with a channel groove, wherein the channel groove comprises a cylindrical groove 2 and a prismatic groove 3, and a terminal groove 4 extends to the edge of the body 1.

Specifically, body 1 is regular pentagonal prism column structure, and cylindrical groove 2 sets up with the colloid piece is coaxial, and prismatic groove 3's quantity is five, and five prismatic grooves 3 are the equipartition in cylindrical groove 2's periphery and are linked together with cylindrical groove 2 respectively in the circular matrix form.

In this embodiment, the diameter of the cylindrical groove 2 is 2.8mm, the prismatic groove 3 is hexagonal prism-shaped, the longest diagonal of the cross section of the prismatic groove 3 is 7.9mm, the distance between two planes parallel to the longest diagonal of the prismatic groove 3 is 1.8mm, and the obtuse included angle between two adjacent inner side walls of the prismatic groove 3 is 130 °.

Wherein, the passageway recess still includes intercommunication recess 5, and prismatic recess 3 utilizes intercommunication recess 5 to be linked together with cylindrical groove 2, and the quantity of intercommunication recess 5, terminal recess 4 all is unanimous and the one-to-one with prismatic recess 3, and five terminal recesses 4 are perpendicular with the sideline of the body 1 of pentaprism form respectively.

In the present embodiment, the communication groove 5 and the terminal groove 4 each have a length of 1.6mm and a width of 1.2 mm.

The invention also provides a rhizosphere channel microfluidic chip which utilizes the colloid block and comprises a glass substrate 11 and the colloid block which are connected, the channel groove is arranged towards the glass substrate 11, and a rhizosphere channel is formed in a space surrounded by the channel groove and the glass substrate 11.

When in use, the colloid block is subjected to surface hydrophilic modification treatment by adopting oxygen plasma, the colloid block made of polydimethylsiloxane is connected with the glass substrate 11, the channel groove and the glass substrate 11 enclose a rhizosphere channel, in addition, polyethylene pipes are respectively inserted into the cylindrical groove 2 and the prismatic groove 3, one polyethylene pipe is used for planting plants, the other polyethylene pipe is connected with a peristaltic pump to introduce a microorganism solution (different microorganisms of the same plant) into the rhizosphere channel, that is, if plants are planted in the polyethylene pipe at the cylindrical grooves 2, a microbial solution is introduced at the prismatic grooves 3, if microorganism solution is introduced into the polyethylene pipe at the cylindrical groove 2, plants are planted in the polyethylene pipe at the prismatic groove 3, after specific culture, and (3) observing the microbial activity of the sample by using a fluorescence microscope, and qualitatively or quantitatively detecting the active enzyme of the sample on the chip by using a confocal microscope. In this embodiment, the polyethylene tube has an internal diameter of 1mm or 2 mm. The rhizosphere channel microfluidic chip can be used for detecting the process of the geochemical interface of the rice soil organism, can realize the parallel detection of a plurality of samples, reduces the using amount of reagents and the samples, can carry out qualitative or quantitative detection on the element content and the enzyme activity on the analyte with short time scale and limited volume, has simple and convenient operation, is suitable for simulating the soil environment and observing the dynamic change of the geochemical interface of the soil organism.

Wherein, the glass substrate 11 is bonded with the colloid block and is tightly connected.

The invention also provides a die for manufacturing the colloidal block, which comprises a base 6, a silicon wafer 7 and a fixing frame 9, wherein the base 6 is arranged at the bottom of the silicon wafer 7, the fixing frame 9 is arranged at the top of the silicon wafer 7, the fixing frame 9 can fix the relative positions of the silicon wafer 7 and the base 6, the fixing frame 9 is of a hollow structure, a groove-shaped forming cavity is formed by a cavity surrounded by the base 6, the fixing frame 9 and the silicon wafer 7, a convex block 8 is arranged at the top of the silicon wafer 7, and the shape of the convex block 8 is matched with that of a channel groove. The projection 8 on the silicon chip 7 is made by adopting a photoetching technology, the processing precision is improved, the projection 8 can be used for forming a channel groove of a colloid block, when the colloid block is manufactured, the polydimethylsiloxane prepolymer and the curing agent are mixed according to the mass ratio of 13:1, the mixture is fully stirred, bubbles in the mixed solution are removed, the mixture is poured into a forming cavity formed by the base 6, the fixing frame 9 and the silicon chip 7, and the mixture is cured for 4 hours at the temperature of 55 ℃, so that the colloid block can be obtained, and the colloid block has good softness and is not easy to damage.

In order to avoid causing the impact to lead to its dislocation to silicon chip 7 when pouring mixed liquid into the shaping intracavity, set up the constant head tank on base 6, silicon chip 7 sets up in the constant head tank, be convenient for fix a position silicon chip 7 fast, still set up spliced pole 10 on base 6, be provided with on the mount 9 with the connecting hole of spliced pole 10 looks adaptation, spliced pole 10 passes the connecting hole and fixed through fastening nut, connect the fastening, easy dismounting. In the present embodiment, the diameter of the silicon wafer 7 is 55mm, and the height of the bump 8 is determined according to the specific experimental conditions.

In addition, in order to improve the working efficiency of the support colloidal block, the number of the bumps 8 is multiple, the bumps 8 are uniformly distributed at equal intervals, and after the production is finished, a finished product is cut to obtain the colloidal block.

The principle and the implementation mode of the invention are explained by applying a specific example, and the description of the embodiment is only used for helping to understand the method and the core idea of the invention; meanwhile, for a person skilled in the art, according to the idea of the present invention, the specific embodiments and the application range may be changed. In view of the above, the present disclosure should not be construed as limiting the invention.

Claims

1. A colloidal block, comprising: the device comprises a body, wherein channel grooves are formed in the body, the channel grooves comprise cylindrical grooves and prismatic grooves, the prismatic grooves are communicated with the cylindrical grooves, the prismatic grooves are uniformly distributed in the circumferential direction around the axis of the cylindrical grooves, the obtuse included angle between two adjacent inner side walls of the prismatic grooves is 130 degrees, a terminal groove is further formed in one end, away from the cylindrical grooves, of each prismatic groove, and the terminal groove extends to the edge of the body; the body is of a regular pentagonal prism structure, the cylindrical grooves and the colloid blocks are coaxially arranged, and the number of the prismatic grooves is five; during the use, be in respectively cylinder groove department with prism groove insert the polyethylene pipe, the alternative polyethylene pipe plants the plant, in addition polyethylene union coupling peristaltic pump channels into microorganism solution in to the rhizosphere passageway.

2. The gel block of claim 1, wherein: the diameter of cylinder recess is 2.8mm, prismatic groove is hexagonal prism form, prismatic groove's the longest diagonal of cross section is 7.9mm, prismatic groove with the distance between the two planes that the longest diagonal parallels is 1.8mm, the obtuse angle contained angle between the two adjacent inside walls of prismatic groove is 130.

3. A gel block according to claim 2, wherein: the channel groove further comprises a communicating groove, the prismatic groove is communicated with the cylindrical groove by the communicating groove, and the number of the communicating grooves is consistent with that of the prismatic grooves and corresponds to that of the prismatic grooves one to one.

4. The colloidal block of claim 3, wherein: the length of the communication groove and the length of the terminal groove are both 1.6mm, and the width of the communication groove and the terminal groove are 1.2 mm.

5. A rhizosphere channel microfluidic chip utilizing the colloidal block of any one of claims 1-4, wherein: the device comprises a glass substrate and the colloid block which are connected, wherein a channel groove is arranged towards the glass substrate, and a rhizosphere channel is formed in a space surrounded by the channel groove and the glass substrate.

6. The rhizosphere channel microfluidic chip of claim 5, wherein: and the glass substrate is bonded with the colloid block.

7. A mold for making the colloidal block according to any of claims 1-4, wherein: the silicon chip fixing device comprises a base, a silicon chip and a fixing frame, wherein the base is arranged at the bottom of the silicon chip, the fixing frame is arranged at the top of the silicon chip, the fixing frame can fix the relative position of the silicon chip and the base, the fixing frame is of a hollow structure, a cavity enclosed by the base, the fixing frame and the silicon chip forms a forming cavity, a convex block is arranged at the top of the silicon chip, and the shape of the convex block is matched with that of a channel groove.

8. The mold for manufacturing a colloidal block as set forth in claim 7, wherein: the silicon chip fixing device is characterized in that a positioning groove is formed in the base, the silicon chip is arranged in the positioning groove, a connecting column is further arranged on the base, a connecting hole matched with the connecting column is formed in the fixing frame, and the connecting column penetrates through the connecting hole and is fixed through a fastening nut.

9. The mold for manufacturing a colloidal block as set forth in claim 7, wherein: the quantity of lug is a plurality of, and is a plurality of the lug is equidistant equipartition.