KR20160099268A - Pcr microdevice for self-actuated pumping with uniform rate - Google Patents

Pcr microdevice for self-actuated pumping with uniform rate Download PDF

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Publication number
KR20160099268A
KR20160099268A KR1020150021510A KR20150021510A KR20160099268A KR 20160099268 A KR20160099268 A KR 20160099268A KR 1020150021510 A KR1020150021510 A KR 1020150021510A KR 20150021510 A KR20150021510 A KR 20150021510A KR 20160099268 A KR20160099268 A KR 20160099268A
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South Korea
Prior art keywords
pcr
substrate
fluid
conduit
outlet conduit
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Application number
KR1020150021510A
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Korean (ko)
Inventor
이내윤
큐더론 트린
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가천대학교 산학협력단
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Priority to KR1020150021510A priority Critical patent/KR20160099268A/en
Publication of KR20160099268A publication Critical patent/KR20160099268A/en

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L7/00—Heating or cooling apparatus; Heat insulating devices
    • B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/02—Adapting objects or devices to another
    • B01L2200/026—Fluid interfacing between devices or objects, e.g. connectors, inlet details
    • B01L2200/027—Fluid interfacing between devices or objects, e.g. connectors, inlet details for microfluidic devices

Abstract

Provided is a polymerase chain reaction (PCR) microdevice for a semiautomatic fluid transfer with a uniform flow rate. The PCR microdevice according to an embodiment of the present invention comprises: an inlet conduit through which a fluid is introduced; an outlet conduit through which the fluid is discharged; and a substrate having one end connected with the inlet conduit and the other end connected with the outlet conduit, wherein a PCR reaction channel is formed on the substrate and a fluid passes through the PCR reaction channel to generate a PCR reaction having multiple cycles. The outlet conduit is made of a gas-permeable substance, and the substrate is made of a gas-impermeable substance. According to the present invention, a fluid can be delivered to an outlet hole at a uniform flow rate by maintaining a difference of fluid pressure at front and rear sides within a constant range.

Description

PCR MICRODEVICE FOR SELF-ACTUATED PUMPING WITH UNIFORM RATE FOR SEMICONDUCTOR FLUID TRANSPORT

The present invention relates to a PCR microdevice for performing PCR (polymerase chain reaction), and more particularly, to a PCR microdevice for semi-automatic fluid transportation having a uniform flow rate.

Due to the importance of fluid transport, micropumps have been recognized as one of the most important components in the micro total analysis system (TAS). This is the reason why related studies are actively conducted. Particularly, a lot of studies have been made to simplify fluid injection and transportation. Recently, research and development of a self-actuated micropump has been active.

The semi-automatic micropump has various advantages over the micropump using an external power source, but some limitations have been pointed out in the case of the semi-automatic micropump disclosed to date. First, it does not have a uniform fluid flow rate when the fluid transport length is long (e.g., a few meters). Second, it does not reliably transport fluids under conditions such as high temperature conditions (e.g., 95 DEG C). Therefore, a method for overcoming the above limitations is required.

Patent Document 1: Korean Patent No. 10-1392724 (published on September 30, 2013)

The present invention aims to provide a PCR microdevice which has a uniform fluid flow rate even when the fluid transportation length is several meters and is stably driven even under high temperature conditions.

According to an aspect of the present invention, there is provided an apparatus comprising: an inlet conduit through which fluid is introduced; An outlet conduit through which the fluid flows; And a first substrate on which a PCR reaction channel is formed, one end of which is connected to the inlet conduit and the other end of which is connected to the outlet conduit to cause a PCR reaction having a plurality of cycles while the fluid passes therethrough, The first substrate may be made of a gas impermeable material, and a PCR micro device may be provided.

At this time, the first substrate may be formed of at least one of PMMA (polymethyl methacrylate), PET (polyethylene terephthalate), PBT (polybutylene terephthalate), PC (polycarbonate), PS (polystyrene) And may be formed of PE (polyethylene), PTFE (polytetrafluoroethylene), or PU (polyurethane).

The PCR micro device may further include a second substrate covering the upper portion of the first substrate, wherein the second substrate is at least one of PMMA (polymethyl methacrylate), PET (polyethylene terephthalate), PBT (polybutylene terephthalate ), PC (polycarbonate), PS (polystyrene), PP (polypropylene), PE (polyethylene), PTFE (polytetrafluoroethylene) or PU (polyurethane).

Also, the outlet conduit may be a silicone tube.

According to another aspect of the present invention, the PCR microdevice; And a phage syringe connected to the inlet conduit of the PCR microdevice for introducing the fluid and compressing air trapped therein to raise the pressure of the rear end of the fluid above the atmospheric pressure, have.

The apparatus may further include a heater disposed under the first substrate of the PCR micro device.

The PCR microdevice according to embodiments of the present invention can be manufactured by making the PCR reaction channel and the phage syringe as a gas impermeable material and manufacturing the outlet conduit as a gas permeable material, The fluid can be delivered to the outlet at the flow rate.

1 is a view for explaining a PCR micro device according to an embodiment of the present invention.
FIG. 2 is a simplified view of the PCR microdevice of FIG. 1. FIG.
FIG. 3 is a graph showing the pressure gradient at the front and rear ends of the sample fluid of FIG. 2 and the residence time of the sample fluid.
4 is a view for explaining a process of manufacturing a PCR micro device according to an embodiment of the present invention.
Figure 5 is an image of a semi-automated fluid flow test.
6 is a graph showing the results of the semi-automatic fluid flow test.
7 is an image for explaining PCR execution and results.

Hereinafter, the present invention will be described in detail with reference to the accompanying drawings.

1 is a view for explaining a PCR microdevice 100 according to an embodiment of the present invention. Referring to FIG. 1, the PCR microdevice 100 may include an inlet conduit 110, an outlet conduit 120, and a substrate 140 having a PCR reaction channel 130 formed thereon. Also, in the PCR microdevice 100, the inlet conduit 110 may be connected to the phalanx syringe 200 and may be disposed on the heater, although not shown in FIG.

The PCR microdevice 100 functions to amplify PCR (polymerase chain reaction) amplification of the sample fluid introduced into the PCR microdevice 100. The sample fluid may refer to a reagent in which test samples (e.g., DNA template, PCR reagent, etc.) are mixed. The form of the sample fluid may be, for example, a sample-plug.

The inlet conduit 110 serves to inject the sample fluid into the PCR reaction channel 130 and the outlet conduit 120 functions to drain the sample fluid from the PCR reaction channel 130. That is, the inlet conduit 110 is connected to one end of the PCR reaction channel 130, and the outlet conduit 120 is connected to the other end of the PCR reaction channel 130.

The inlet conduit 110 and the outlet conduit 120 can be made tubular. At this time, the size of the outer diameter or the inner diameter is not specified. The outlet conduit 120 is also made of a gas-permeable material. The inlet conduit 120 may be made of a gas permeable material similar to the outlet conduit 120 and may be made of a gas-impermeable material, unlike the outlet conduit 120.

Here, the gas permeable material means a material (material) that can be permeated to the outside by the gas trapped therein. For example, the outlet conduit 120 may be formed of a silicon tube having gas permeability.

The PCR reaction channel 130 may be formed on the upper surface of the substrate 140 where the PCR reaction occurs. The PCR reaction channel 130 may function as a flow path through which the sample fluid flows and the sample fluid may be PCR amplified along the PCR reaction channel 130.

The width, depth, etc. of the PCR reaction channel 130 are not specified. For example, the width and depth of the PCR reaction channel 130 can each have a size of tens to hundreds of micrometers.

The form of the PCR reaction channel 130 is also not specified. However, for the sake of convenience, the PCR reaction channel 130 has a serpentine form. Here, the serpentine form means that the channel is formed in a serpentine form like a so-called "serpentine shape". As shown in FIG. 1, the PCR reaction channel 130 has a plurality of flow paths arranged in parallel in a transverse direction at predetermined intervals, and one flow path end is connected to another adjacent flow path end. Accordingly, the fluid flowing along the PCR reaction channel 130 can flow along the advancing direction of the fluid up and down in the vertical direction.

In general, DNA amplification using PCR requires temperatures suitable for the denaturation, annealing and extension stages (ie, there are three temperature zones, one for each step). If the size of the target amplicon is less than 300 bp, the binding / extension step may be performed at the same temperature.

Accordingly, the PCR reaction channel 130 may include at least two different temperature zones on the same plane. For example, the PCR reaction channel 130 may be subjected to a thermal denaturation step in the upper part (inlet degree observation) based on the same plane, and to a binding / expansion step in the lower part (outlet degree observation). At this time, the division of the upper part and the lower part depends on the temperature applied by the heater (heat source) disposed at the lower part of the substrate 140, and is not limited to a specific part. That is, the portion where the heat denaturation step or the bonding / stretching step is performed may be changed depending on the position of the heater or the temperature adjustment.

Meanwhile, the substrate 140 can be formed in a plate shape and is made of a gas-impermeable material. Here, the gas impermeable material means a material (material) in which the gas trapped therein can not be transmitted to the outside. The gas permeability may vary depending on the temperature, the polymer composition, the gas to be permeated, etc. In the present specification, the gas impermeability is such that the permeability coefficient of the gas is approximately 20 × 10 -13 cm 3 · cm -2 s -1 Pa -1 ≪ / RTI > Examples of such gas impermeable materials include polymethyl methacrylate (PMMA), polyethylene terephthalate (PET), polybutylene terephthalate (PBT), polycarbonate (PC), PS (polystyrene) (Polyethylene), PTFE (polytetrafluoroethylene), or PU (polyurethane). Any gas-impermeable material may be used as the material of the substrate 140 . For convenience of description, the description will be made mainly in the case where the substrate 140 is made of PMMA.

The PCR micro device 100 may further include another substrate 150 covering an upper portion of the substrate 140 on which the PCR reaction channel 130 is formed. Hereinafter, the substrate 140 on which the PCR reaction channel 130 is formed will be referred to as a first substrate, and another substrate 150 covering an upper portion of the first substrate 140 will be referred to as a second substrate. The PCR reaction channel 130 is formed on the upper surface of the first substrate 140 at a recessed angle and the second substrate 150 covers the upper portion of the first substrate 140 to form the closed PCR reaction channel 130. [ Can be completed.

The second substrate 150 may be formed in the same plate shape as the first substrate 140 and may be made of a gas-impermeable material. For example, the second substrate 150 may be formed of a material such as PMMA (polymethyl methacrylate), PET (polyethylene terephthalate), PBT (polybutylene terephthalate), PC (polycarbonate), PS (polystyrene) (Polyethylene), PTFE (polytetrafluoroethylene), PU (polyurethane), or the like.

The PCR microdevice 100 may be a PCR microdevice system with a phage type syringe 200 and a heater (not shown).

The phage syringe 200 is for injecting fluid into the inlet conduit 110 of the PCR microdevice 100 and may be a hand-held syringe. The end of the phalanx syringe 200 may be connected to the inlet conduit 110. 1, the end of the outlet conduit 120 of the PCR microdevice 100 is closed (in FIG. 1, a closure of the end of the outlet conduit 120 is shown with a clip), a phage syringe 200 can compress the air trapped in the channel by the piston movement of the piston.

The phalanx syringe 200 is made of a gas-impermeable material. For example, the phalanx syringe 200 may be formed of a gas-impermeable plastic, and such a plastic syringe is available by purchasing a commercially available one.

The heater functions as a heat source and controls the temperature applied to the PCR reaction channel 130 formed on the first substrate 140 of the PCR micro device 100. Such a heater may be disposed under the first substrate 140.

The heater may use a single heater or a plurality of heaters, and in the latter case, the plurality of heaters may be driven at different temperatures. That is, when there are two heaters, one heater is driven to a temperature for performing the thermal denaturation step, and the other heater can be driven to a temperature for performing the bonding / stretching step.

Hereinafter, the principle of the present invention will be described. It is noted that the same reference numerals are used for the same components as those described in FIG. 1 for convenience of explanation.

FIGS. 2 and 3 are diagrams for explaining the operation principle of the PCR microdevice of FIG. Fig. 2 is a simplified view of the PCR microdevice of Fig. 1, and Fig. 3 is a graph showing the pressure gradient at the front and rear ends of the sample fluid s in Fig. 2 and the residence time of the sample fluid s.

Referring to FIG. 2, the end of the phage syringe 200 is connected to the conduit f. Here, 'conduit (f)' is used to collectively refer to the inlet conduit 110, the PCR reaction channel 130, and the outlet conduit 120 described above.

As described above, the phalanx syringe 200 is formed of a gas-impermeable material (e.g., plastic). And the inlet conduit 110 in the conduit f and the PCR reaction channel 130 are formed of a gas impermeable material such as PMMA and the outlet conduit 120 is formed of a gas permeable material such as a silicone tube. That is, the gas can be transmitted to the outside at the end of the conduit f opposite to the phagocytic syringe 200 with respect to the traveling direction of the fluid.

As shown in Fig. 2, it is assumed that the outlet side end of the conduit f is closed (e.g., closed by a clip) and the sample fluid s is located in the conduit f. In this case, Pp shown in Fig. 2 is the air pressure inside the rear end of the sample fluid (s), and Pa is the air pressure inside the front end of the sample fluid (s). The rear end means the back side of the sample fluid s with reference to the flow direction of the sample fluid s, and the front end means the front side of the sample fluid s. At this time, the pressure gradient Pg between the front and rear ends of the sample fluid s can be defined as the difference between Pp and Pa.

The air trapped in the closed conduit f can be compressed by the piston motion of the phalanx syringe 200, in which case the pressure inside the rear end becomes higher than the atmospheric pressure. Before the sample fluid s is introduced into the conduit f, the air pressures in the conduit f are all the same at the front and rear ends (i.e., Pp = Pa). However, once the sample fluid s is introduced into the conduit f, there is no air exchange between the front and rear ends of the sample fluid s, so that the front and rear ends are physically separated. At this time, since the front end portion includes the outlet conduit 120, it has a gas permeable property, and the rear end portion does not include the outlet conduit 120, and thus has a gas impermeable property. Of course, the inlet conduit 110 may be made of a gas permeable material, but in this case, the gas permeable nature of the inlet conduit 110 is negligible considering the specific gravity of the inlet conduit 110 at the rear end. It can be assumed that the air in the conduit f only diffuses outwardly through the outlet conduit 120 only.

When air diffusion from the front end of the sample fluid (s) to the outside occurs, a pressure drop occurs at the front end. On the other hand, there is no pressure drop at the rear end (negligible even if it occurs). Therefore, the sample fluid s can always continue to advance semi-automatically along the conduit f as a high pressure is always maintained within the rear end compared to the front end of the sample fluid s.

Also, since the pressure in the rear end is kept constant, the pressure gradient between the front and rear ends can also be maintained at a certain level (see FIG. The fact that the pressure gradient between the front and rear ends is maintained at a constant level means that the sample fluid s can advance in the conduit f at a constant speed. This is because the back end pressure of the sample fluid (s) is maintained at a constant level while the sample fluid (s) moves along the conduit (f). Therefore, even if the length of the conduit f is long, the sample fluid s can move at a constant speed, so that the residence time of the sample fluid s can be kept constant (see Fig. 3B). On the other hand, the flow velocity of the sample fluid s is controlled by adjusting the length of the conduit f, the size (outer diameter, inner diameter, height, width, etc.) of the outlet conduit 120, the inner pressure of the front end portion (degree of gas permeability) Can be adjusted accordingly.

In other words, the diffusion of the trapped air in the conduit f to the outside takes place only at the outlet conduit 120, which acts as a driving force to semi-automatically advance the sample fluid s. Also, since the trailing edge of the sample fluid s has gas impermeable properties, the initial pressure formed by the phage syringe 200 can be maintained at a constant level during movement of the sample fluid s. Therefore, even if the length of the conduit f becomes long, the moving speed of the sample fluid s is not reduced, and the residence time can be kept constant.

As described above, the PCR microdevice according to the embodiments of the present invention can produce a PCR reaction channel and a phage type syringe as a gas impermeable substance and produce an outlet conduit as a gas permeable substance, So that the fluid can be delivered to the outlet at a uniform flow rate.

Hereinafter, specific examples and experimental examples of the PCR microdevice according to the present invention will be described. It is to be understood that the present invention is not limited by the following examples and experimental examples.

1. Manufacture of PCR microdevices

4 is a view for explaining a process of manufacturing a PCR micro device according to an embodiment of the present invention. Referring to FIG. 4, a supernatant microchannel (PCR reaction channel) was formed on a PMMA substrate (40 × 40 × 2 mm) using a computer numerical control (CNC) milling machine. The microchannel was formed to have a width of 200 mu m and a depth of 50 mu m (see Figs. 4A and 4B).

Next, another PMMA substrate having the same size as that of the PMMA substrate is prepared, and a hole through which the inlet conduit and the outlet conduit are inserted is formed through the drilling machine. The holes are formed at positions corresponding to both ends of the microchannel when the two PMMA substrates are stacked. Both PMMA substrates are then thermally bonded at about 105 DEG C (see FIG. 4C).

Finally, a silicone tube having an inner diameter of 0.2 mm and an outer diameter of 2 mm is used as an inlet conduit, and a silicone tube having an inner diameter of 1 mm and an outer diameter of 0.2 mm is inserted into the hole as an outlet conduit, and is hardened by using a PDMS prepolymer ).

The PCR microdevice thus prepared has 25 thermal cycles and the total length of the microchannels is 1.25 m, corresponding to a length of 5 cm per cycle.

2. Semi-automatic fluid flow test

FIG. 5 is an image of a semi-automatic fluid flow test for confirming the performance of the PCR micro device manufactured in the above 1. FIG. Referring to FIG. 5, first, red ink was used to confirm the position of the sample fluid, and the sample fluid containing the red ink was sucked into the pulp type syringe (see upper left corner of FIG. 5A).

The exit conduit was clipped and closed before the phage type syringe (20 mL) was connected to the PCR microdevice. The piston of the phage syringe was then pulled to the proper position and connected to the inlet conduit of the PCR microdevice (see FIG. 5b).

Next, the piston of the gripping type syringe is pushed to a suitable position to compress the air inside the gripping type syringe, and the gripping body of the gripping type syringe and the piston are strapped (see Fig. 5C). Thereby causing the sample fluid to advance along the conduit. In Fig. 5D, the advancement of the sample fluid can be confirmed.

More specifically, describing the test conditions, three tests were performed for each item for reproducibility, and the initial internal pressure was set at approximately 2 atm for all tests. This could be accomplished by pushing the piston of the phalanx syringe from 20 to 10 with respect to the scale shown on the body.

6 is a graph showing the results of the semi-automatic fluid flow test. 6, the x-axis represents the cycle and the y-axis represents the residence time of the sample fluid. Hereinafter, the results of this test will be described with reference to Fig.

(1) First test: Flow test according to the change of outlet conduit length

In the first test, the flow test was carried out by varying the length of the outlet conduit. The results are shown in Fig. 6A. Referring to FIG. 6A, the lengths of the outlet conduits were set at 1 cm, 2 cm, and 3 cm, respectively. As a result of the test, the retention time of the sample fluid becomes shorter as the length of the outlet conduit becomes longer, which means that the flow rate of the sample fluid increases. Specifically, when the lengths of the outlet conduits were 3 cm, 2 cm, and 1 cm, respectively, the total residence time of the sample fluid was 21 minutes, 31 minutes, and 50 minutes, respectively, and the average residence time was approximately 49 seconds, 72 seconds, 119 Seconds. This means that the longer the outlet conduit is, the higher the gas permeability is, and the higher the flow velocity is interpreted. From this test, it was confirmed that the flow rate of the sample fluid can be controlled by adjusting the length of the outlet conduit.

(2) Second test: Flow test according to depth change of microchannel

In the second test, the flow test was performed while varying the depth of the microchannel (PCR reaction channel). The results are shown in Fig. 6B. Referring to FIG. 6B, the depths of the microchannels were set to 20 μm, 50 μm, and 100 μm, respectively. The width of the microchannel was 200 mu m and the length of the outlet conduit was fixed at 3 cm. As a result of the test, the shorter the depth of the microchannel, the shorter the retention time of the sample fluid, which means that the flow rate of the sample fluid increases. Specifically, when the depths of the microchannels were 20 μm, 50 μm, and 100 μm, the total residence time of the sample fluid was 12 minutes, 21 minutes, and 27 minutes, and the average residence time was approximately 28 seconds, 49 seconds , And 63 seconds. This is interpreted as a result of lowering the pressure gradient as the depth of the microchannel is deeper and the flow velocity is lowered.

(3) Third test: Flow test according to microchannel length change

In the third test, the flow test was performed while changing the length of the microchannel (PCR reaction channel). Referring to FIG. 6C, the lengths of the microchannels were set to 1.25 m (25 cycles), 2.25 m (45 cycles), and 2.75 m (55 cycles), respectively. The width of the microchannel was 200 μm, the depth was 80 μm, and the length of the outlet conduit was fixed at 3 cm. As a result of the test, it was found that the change of the length of the microchannel did not greatly affect the flow rate of the sample fluid. Specifically, when the microchannel lengths were 25, 45, and 55 cycles, the total residence time of the sample fluid was 21 minutes, 37 minutes, and 49 minutes. However, the average residence time was 49 seconds, 48 seconds, and 53 seconds There was no difference. This is interpreted to be due to the same level of pressure at the back end of the sample fluid, regardless of the length of the microchannel.

From the first to third tests as described above, it has been found that the length of the outlet conduit has the greatest influence on the flow velocity (because it is the main factor in forming the pressure gradient), and the depth of the microchannel also affects the flow velocity . Conversely, it was confirmed that the flow rate can be controlled by controlling the length of the outlet conduit and the depth of the microchannel.

3. PCR and Results

PCR was performed using the prepared PCR microdevices. Figure 7 is an image for explaining PCR execution and results. Referring to FIG. 7, a phage type syringe was connected to a PCR micro device according to an embodiment of the present invention and placed on two copper blocks (see FIG. 7A). The surface temperature measurement of the PCR microdevices was performed using an infrared (IR) camera (FLIR Thermovision A320) (see FIG. 7B).

E. coli and human genomic DNA (Roche) containing pGEM-3Zf (+) plasmid vector were used as DNA template for 2-temperature PCR. The heat denaturation temperature was adjusted to 95 ± 0.5 ° C and the binding / elongation temperature was adjusted to 68 ± 0.2 ° C for pGEM-3Zf (+) plasmid vector and 63 ± 0.1 ° C for human genomic DNA. The primer sequences for amplifying the 230 bp gene fragment in the pGEM-3Zf (+) plasmid vector are as follows: 5'-CCG GCG AAC GTG GCG AGA AAG GAA GGG AAG AAA GC- TGC AGC ACA TCC CCC TTT CGC CAG C-3 '(retrograde). The primer sequences for amplification of human genomic DNA at the D1S80 position are: 5'-GAA ACT GGC CTC CAA ACA CTG CCC GCC G-3 '(transitional) and 5'- GTC TTG TTG GAG ATG CAC GTG CCC CTT GC-3 '(retrograde).

The PCR reagents included a green buffer (5x), a mixture of 0.2 mM dNTPs, 1 mg / ml BSA, 1 μM forward and reverse primers and 0.075 U / μl Taq polymerase. The commercially available human genomic DNA (200 ng / [mu] L) was diluted and added to PCR reagents to make the final template concentration and similarly the pGEM-3Zf (+) plasmid vector was diluted and added to the PCR reagent to the final template concentration (5ng per reaction).

The PCR results are shown in FIGS. 7C and 7D. FIG. 7C shows the amplification result of the 230 bp gene fragment obtained from the pGEM-3Zf (+) plasmid vector, and FIG. 7D shows the amplification result of the D1S80 position (369-801 bp) obtained from the human genomic DNA. 25 cycles were used for amplification. For comparison, the same PCR reaction solution was introduced using a thermal cycler (using a 100 bp DNA class marker).

Referring to FIG. 7C, the average intensity of the target appplicons obtained using the PCR microdevice according to an embodiment of the present invention was approximately 97.2% of the average intensity obtained using the thermal cycler. Referring to FIG. 7D, the average intensity of the target amblikers obtained using the PCR microdevice according to an embodiment of the present invention was approximately 71.7% of the average intensity obtained using the thermal cycler. That is, in both cases, it can be confirmed that the target bands are successfully amplified, which is a result of showing the reliability of the PCR microdevice according to the present invention.

The embodiments of the present invention have been described above. It will be apparent to those skilled in the art that various modifications and variations can be made in the present invention without departing from the spirit or scope of the inventive concept as defined by the appended claims. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the scope of the present invention.

100: PCR microdevice
110: inlet conduit
120: outlet conduit
130: PCR reaction channel
140: first substrate
150: second substrate
200: Finger Syringe

Claims (6)

An inlet conduit through which the fluid flows;
An outlet conduit through which the fluid flows; And
A first substrate on which a PCR reaction channel is formed to cause a PCR reaction having a plurality of cycles while one end is connected to the inlet conduit and the other end is connected to the outlet conduit,
Wherein the outlet conduit is made of a gas-permeable material, and wherein the first substrate is made of a gas-impermeable material. The method according to claim 1,
The first substrate may be formed of at least one selected from the group consisting of PMMA (polymethylmethacrylate), PET (polyethylene terephthalate), PBT (polybutylene terephthalate), PC (polycarbonate), PS (polystyrene) ), PTFE (polytetrafluoroethylene), or PU (polyurethane). The method of claim 2,
And a second substrate covering the upper portion of the first substrate,
The second substrate may be formed of a material selected from the group consisting of PMMA (polymethyl methacrylate), PET (polyethylene terephthalate), PBT (polybutylene terephthalate), PC (polycarbonate), PS (polystyrene) ), PTFE (polytetrafluoroethylene), or PU (polyurethane). The method of claim 2,
Wherein the outlet conduit is a silicon tube. A PCR microdevice according to any one of claims 1 to 4; And
And a phage syringe connected to the inlet conduit of the PCR microdevice for introducing the fluid and for compressing the trapped air to raise the pressure of the rear end of the fluid above atmospheric pressure. The method of claim 5,
And a heater disposed below the first substrate of the PCR micro device.
KR1020150021510A 2015-02-12 2015-02-12 Pcr microdevice for self-actuated pumping with uniform rate KR20160099268A (en)

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Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR101392724B1 (en) 2012-03-20 2014-05-09 가천대학교 산학협력단 Microfluid device for continuoous-flow pcr

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR101392724B1 (en) 2012-03-20 2014-05-09 가천대학교 산학협력단 Microfluid device for continuoous-flow pcr

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