CN114354696B - DNA biosensor driven by friction nano generator and application thereof - Google Patents
DNA biosensor driven by friction nano generator and application thereof Download PDFInfo
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Abstract
The invention belongs to the technical field of biological detection, and particularly relates to a DNA biosensor driven by a friction nano generator and application thereof. A friction nano-generator driven DNA biosensor comprising: the friction nano generator is used for converting external mechanical energy into electric energy and outputting the electric energy; the resistance type biosensor is electrically connected with the friction nano generator and captures target DNA under the drive of electric energy output by the friction nano generator; and a carbon nanotube for binding the target DNA and causing the resistive biosensor to generate a sensing signal; a surface-modified DNA capture probe of the resistive biosensor; and the surfaces of the carbon nanotubes are modified with DNA signal probes. The DNA sensor has a simple structure, is driven by a friction nano generator, does not need an external power supply, and improves the portability of the sensor; the DNA probe technology is adopted, so that the concentration detection of the pM-grade DNA can be realized, and the sensitivity and the specificity are very high.
Description
Technical Field
The invention belongs to the technical field of biological detection, and particularly relates to a DNA biosensor driven by a friction nano generator and application thereof.
Background
The current "gold standard" for detecting and identifying bacteria is the use of culture-based methods. While conventional culture-based techniques are reliable and accurate and relatively low cost, the method is time consuming and laborious, may take days or even weeks to obtain results, and is limited to culturable bacteria. In addition, there are some important bacteria that are difficult or impossible to culture and are not detected by the culture method. Molecular techniques are particularly useful for detecting a variety of substances or low template amounts. Detection of specific DNA sequences is becoming increasingly important in bacterial detection. The analysis of bacterial DNA is a common bacterial analysis means, which is of great importance in molecular diagnosis of bacteria, rapid classification of bacterial species and phenotyping. Many detection methods have been proposed, including fluorescence, electrochemiluminescence, enzymatic, surface plasmon resonance, polymerase Chain Reaction (PCR) detection and 16S ribosomal RNA (16S rRNA) fragment-related diagnostic methods. However, these methods have the disadvantages of severe experimental conditions, complex operation, environmental pollution, unstable activity and sometimes false positive when applied to complex samples, which makes them an important bottleneck for molecular diagnosis. There is therefore a need for a faster, more convenient and easier to handle method of detecting bacterial DNA fragments.
Electrochemical DNA sensors have become an interesting choice in the existing methods because of their simplicity, portability and low cost. But its application is limited because it requires additional tags and power sources. Since 2012, friction nano-generators have been proposed based on coupling of triboelectrification and electrostatic induction effects as efficient means and methods of converting various mechanical energy (e.g., human walking, wind energy, water wave energy, etc.) into electrical energy. Meanwhile, a friction nano generator (TENG) is also used to prepare various self-driven sensors. The self-driven DNA sensor does not need a power supply and has high sensitivity, so that the development of a novel self-driven DNA sensor based on TENG is a promising thought.
Disclosure of Invention
The invention aims at solving the problems in the existing biological detection technology and provides a DNA biosensor driven by a friction nano generator, wherein the DNA biosensor uses the friction nano generator as a driving power supply to detect DNA of a biological sample and has the advantages of high sensitivity, rapid detection, no need of an external power supply and the like.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows: a friction nano-generator driven DNA biosensor comprising:
the friction nano generator is used for converting external mechanical energy into electric energy and outputting the electric energy;
the resistance type biosensor is electrically connected with the friction nano generator and captures target DNA under the drive of electric energy output by the friction nano generator; the method comprises the steps of,
a carbon nanotube for binding the target DNA and causing the resistive biosensor to generate a sensing signal;
a surface-modified DNA capture probe of the resistive biosensor;
and the surface of the carbon nano tube is modified with a DNA signal probe.
Further, the friction nano generator comprises two conductive layers, wherein the friction layer is arranged on the surface of one conductive layer; the friction layer is used as a negative electrode of the power generation layer, the other conductive layer is used as a positive electrode of the power generation layer, and the positive electrode of the power generation layer and the negative electrode of the power generation layer generate electricity through friction under the action of external force.
Further, the friction layer is a perfluoroethylene propylene copolymer film having a thickness of 50 μm.
Further, the conductive layer is an aluminum film, and the thickness of the conductive layer is 50 micrometers.
Further, a supporting substrate is arranged on one surface of the conductive layer opposite to the friction surface of the conductive layer; an elastic element is connected between the two support substrates.
Further, the resistance type sensor is made of etched ITO conductive glass, a gap with the width of 50 microns is etched in the middle of the ITO conductive glass, and the etching depth is equal to the thickness of the ITO film layer; the etched ITO conductive glass surface is decorated with silane.
Further, the sensing system also includes a variable resistor connected in series between the friction nano-generator and the resistive biosensor.
Further, the two ends of the variable resistor are connected with an alarm device in parallel.
The invention also provides application of the DNA biosensor driven by the friction nano generator, and the sensor can be applied to detecting bacterial DNA.
The invention also provides a DNA detection method driven by the friction nano generator, which comprises the following steps:
(1) Soaking the resistance type biosensor in a DNA solution to be detected, and incubating for 45min at 37 ℃;
(2) Taking out the sensor, soaking the sensor in a carbon nano tube solution modified with a DNA signal probe, and incubating for 45min at 37 ℃;
(3) Taking out the sensor, lightly flushing the unbound carbon nanotubes with deionized water, and naturally air-drying;
(4) The friction nano generator is used for driving, and voltage values at two ends of the resistance type biosensor are tested;
(5) And calculating the concentration of the target DNA in the solution to be detected by utilizing a pre-fitted voltage-DNA concentration curve equation.
The invention adopts the capture probe and the signal probe which can be specifically and complementarily combined with the target DNA to form a sandwich structure with the target DNA, thereby combining the carbon nano tube modified with the signal probe onto the resistance type biosensor to cause resistance change, further changing the voltage at two ends of the resistance, and inverting the concentration of the target DNA according to the voltage.
The DNA biosensor driven by the friction nano generator has the following beneficial effects:
1. in the invention, the sensor is driven by the friction nano generator, so that the mechanical energy can be converted into electric energy, an external power supply is not needed, and the portability of the sensor is improved;
2. the DNA sensor of the invention adopts the DNA probe technology, can realize the concentration detection of the pM-grade DNA, and has high sensitivity;
3. the base sequences of different bacterial DNA are different, so that the variety of bacteria can be simply identified through voltage signals;
4. the detection method is simple and quick, and results can be obtained only in a few hours, so that compared with the traditional detection method, the efficiency is greatly improved;
5. the DNA sensor has the advantages of simple structure, working at normal temperature and normal pressure and no need of external power supply, is favorable for industrialized popularization, and has potential application value in developing a self-driven DNA sensing network facing wireless sensing.
Drawings
FIG. 1 is a schematic diagram of a friction nano-generator driven DNA biosensor according to an embodiment of the present invention;
FIG. 2 is a schematic diagram of a friction nano-generator according to an embodiment of the present invention;
FIG. 3 is an equivalent circuit diagram of a friction nano-generator driven DNA biosensor according to an embodiment of the present invention;
FIG. 4 (a) is the output voltage of a friction nano-generator;
FIG. 4 (b) is the stability of the friction nano-generator;
FIG. 5 is a representation of the ITO conductive glass after modification; wherein a is an XPS diagram of ITO conductive glass modified silane, b is an XPS diagram of ITO conductive glass modified silane connected with a capture probe, c is an XPS diagram of a carbon nanotube, d is an XPS diagram of a signal probe on the carbon nanotube, and e is an XPS diagram of silane and the capture probe on the ITO conductive glass modified silane;
FIG. 6 is a feasibility analysis of the friction nano-generator driven DNA biosensor;
FIG. 7a is a graph showing the output voltage as a function of DNA concentration;
FIG. 7b is a graph of a linear fit of the error bars after three replicates of DNA at different concentrations;
FIG. 7c is an SEM image of the ITO conductive glass surface at different concentrations of DNA and in blank sets.
Fig. 8 is a 3D view of a portable device of the friction nano-generator driven DNA biosensor.
Detailed Description
In order that the invention may be readily understood, a more particular description thereof will be rendered by reference to specific embodiments that are illustrated in the appended drawings. Preferred embodiments of the present invention are shown in the drawings. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete.
Example 1 this example provides a friction nano-generator driven DNA biosensor, the structure of which is shown in fig. 1. As shown in fig. 1, the sensor includes a friction nano-generator 100, a resistive biosensor 200, a variable resistor 300, and an LED lamp 400. The friction nano-generator 100 is used to convert external mechanical energy into electrical energy to output an electrical signal to an external circuit. The resistive biosensor 200 is electrically connected with the friction nano-generator 100 such that the friction nano-generator 100 supplies power to the resistive biosensor 200 for detecting a sensing signal. The variable resistor 300 is connected in series with the resistive biosensor 200 to divide a portion of the voltage. The LED lamps 400 are connected in parallel across the variable resistor 300 for emitting a light alarm signal when the sensing signal is greater than a certain signal threshold.
Fig. 2 shows a structural illustration of the friction nano-generator 100. As shown in fig. 2, the friction nano-generator 100 includes two support substrates, an elastic member 160, a first conductive layer 103, a second conductive layer 105, and a first friction layer 104. The two support substrates are a first support substrate 101 and a second support substrate 102, and the support substrate is made of PVC board. The elastic element 106 is connected between two support substrates, and in the embodiment shown in fig. 2, the elastic element 106 is a spring, the number of which is four. The first conductive layer 103 is fixed on the lower surface of the first support substrate 101, and the second conductive layer 105 is fixed on the upper surface of the second support substrate 102. The material of the first conductive layer 103 and the second conductive layer 105 is an aluminum film, and the thickness is 50 μm. A first friction layer 104 is provided on the lower surface of the first conductive layer 103, the first friction layer 104 is a perfluoroethylene propylene copolymer film, the thickness is 50 micrometers, and the size and shape of the first friction layer are the same as those of the two conductive layers. The second conductive layer 105 may serve as an electrode material or a second friction layer, and generates electricity by friction with the first friction layer 104. The first conductive layer 103 and the second conductive layer 105 are respectively glued on the two support substrates by Kapton.
The resistive biosensor 200 is made of etched ITO conductive glass, and as shown in fig. 1, a slit having a width of 50 μm is etched in the middle of the ITO conductive glass, and the etching depth is equal to the thickness of the ITO film layer. The etched ITO conductive glass has faults in the middle of the ITO film layer, so that the resistance of the sensor is infinite.
The etched ITO conductive glass surface needs to be subjected to modification treatment of silane and DNA capture probes in sequence so as to specifically identify and combine target DNA.
The sensing system for bacteria detection driven by the friction nano generator provided by the embodiment has the following working principle: the first supporting substrate 101 is pressed, so that the first friction layer 104 and the second conductive layer 105 (second friction layer) are periodically contacted and rubbed, mechanical energy is converted into electric energy, and current is output to the system circuit through the first conductive layer 103 and the second conductive layer 105, and an equivalent circuit diagram thereof is shown in fig. 3. The output voltage was measured by a programmable multifunctional electrometer (Keithley, model 6514) and it was possible to identify whether it was the DNA of interest by LED light on and off.
In the initial state, the resistance type biosensor 200 is formed by etching ITO conductive glass, and the etched ITO conductive glass is not conductive and is connected in a circuit, so that the resistance is infinite.
After the resistive biosensor 200 is soaked in a solution containing target DNA, the DNA capture probes modified on the surface of the resistive biosensor 200 are clipped complementarily to specifically identify and capture the target DNA, and after the target DNA is captured, the resistive biosensor 200 is soaked in a carbon nanotube solution modified by DNA signal probes, and the target DNA captured by the resistive biosensor 200 is complementarily combined with the DNA signal probes to adsorb the carbon nanotubes, so that the resistance of the resistive biosensor 200 is reduced due to the fact that the carbon nanotubes are conductive. When the target DNA concentration is high, the number of carbon nanotubes bound to the resistive biosensor 200 is also large, and the resistance of the resistive biosensor 200 is smaller, and the voltage across the resistive biosensor 200 is smaller, so that the target DNA concentration can be reflected according to the voltage values across the resistive biosensor 200. When the base sequence of the target DNA is longer, the sensitivity can be lower, and the target DNA of 22bp is detected at present, and the lower limit of detection is 1pM.
Fig. 4a and 4b are respectively the output voltage and the power stability of the friction nano generator. As can be seen from fig. 4, the operating voltage of the friction nano generator in the present invention is about 160V. The characterization of the ITO conductive glass and the characterization of the carbon nano tube are shown in fig. 5, wherein a and b are respectively ITO conductive glass modified silane and a capture probe. c. d is the carbon nanotube after modifying the signal probe separately. Figure 5e demonstrates that modifying silane and capture probe have no effect on output. To further determine the feasibility of the invention, the feasibility analysis of the invention is shown in FIG. 6. It is clear that none of the probes is tested in the absence of any probe and that the voltage drops only when the DNA of interest is present.
The voltage change based on the friction nano-generator self-driven DNA sensor to detect DNA of different concentrations is shown in fig. 7 a. Based on the dependence of output voltage on continuous change of DNA concentration, a mode of detecting concentration is provided, along with the increase of target DNA concentration, the voltage is continuously reduced, the lower detection limit is 1pM, and as the increase of target DNA concentration, the number of adhered carbon nanotubes is also increased, so that the conductivity of ITO conductive glass is increased, and the voltage is reduced. The error bars measured in triplicate experiments are shown in figure 7 b. And a correlation coefficient. Has good linear relation and high sensitivity. FIG. 7 is a drawing showing the adhesion of a carbon nanotube by SEM, and as shown in FIGS. 7c, I and II are SEM images when a signal probe is not modified on a carbon nanotube and when a target DNA is not attached, respectively, as can be seen from the sequential decrease in the concentration of III-IX target DNA of FIG. 7c, the adhered carbon nanotube is indeed decreased with the decrease in the concentration of target DNA. In conclusion, the concentration of the target DNA can be judged by analyzing the output voltage, which is based on the working principle of the friction nano generator DNA sensor.
The DNA biosensor of the embodiment can detect any DNA material, is not limited to bacteria, fungi, viruses, animals, plants and the like, and can realize rapid qualitative and quantitative detection of DNA.
Embodiment 2 this embodiment provides a friction nano-generator driven bacterial DNA detection method comprising the steps of:
(1) Soaking the resistance type biosensor in a DNA solution to be detected, and incubating for 45min at 37 ℃;
(2) Taking out the sensor, soaking the sensor in a carbon nano tube solution modified with a DNA signal probe, and incubating for 45min at 37 ℃;
(3) Taking out the sensor, lightly flushing the unbound carbon nanotubes with deionized water, and naturally air-drying;
(4) The friction nano generator is used for driving, and voltage values at two ends of the resistance type biosensor are tested;
(5) And calculating the concentration of the target DNA in the solution to be detected by utilizing a pre-fitted voltage-DNA concentration curve equation.
Fig. 8 is a 3D diagram of a portable device of the DNA sensor of the friction nano-generator of the present invention, the sensor is placed in the middle of the portable device, and an LED lamp is installed at the upper round hole, so that whether the DNA is the target DNA can be easily distinguished, and ITO conductive glass and a variable resistor are respectively placed at the inner groove.
Claims (10)
1. A friction nano-generator driven DNA biosensor comprising:
the friction nano generator is used for converting external mechanical energy into electric energy and outputting the electric energy;
the resistance type biosensor is electrically connected with the friction nano generator and captures target DNA under the drive of electric energy output by the friction nano generator; the method comprises the steps of,
a carbon nanotube for binding the target DNA and causing the resistive biosensor to generate a sensing signal;
a surface-modified DNA capture probe of the resistive biosensor;
and the surface of the carbon nano tube is modified with a DNA signal probe.
2. The friction nano-generator driven DNA biosensor of claim 1, wherein the friction nano-generator comprises two conductive layers, wherein a friction layer is provided on a surface of one of the conductive layers; the friction layer is used as a negative electrode of the power generation layer, the other conductive layer is used as a positive electrode of the power generation layer, and the positive electrode of the power generation layer and the negative electrode of the power generation layer generate electricity through friction under the action of external force.
3. The friction nano-generator driven DNA biosensor according to claim 2, wherein the friction layer is a perfluoroethylene propylene copolymer film having a thickness of 50 μm.
4. The friction nano-generator driven DNA biosensor according to claim 2, wherein the conductive layer is an aluminum thin film having a thickness of 50 microns.
5. The friction nano-generator driven DNA biosensor according to claim 2, wherein the conductive layer is provided with a support substrate on a side opposite to the friction surface thereof; an elastic element is connected between the two support substrates.
6. The friction nano generator driven DNA biosensor according to claim 1, wherein the resistive sensor is made of etched ITO conductive glass, a slit having a width of 50 μm is etched in the middle of the ITO conductive glass, and the etching depth is equal to the thickness of the ITO film layer; the etched ITO conductive glass surface is decorated with silane.
7. The friction nano-generator driven DNA biosensor of claim 1, further comprising a variable resistor in series between the friction nano-generator and the resistive biosensor.
8. The friction nano generator driven DNA biosensor according to claim 7, wherein both ends of the variable resistor are connected in parallel with an alarm device.
9. Use of a friction nano-generator driven DNA biosensor according to any of claims 1-8, wherein the sensor is applied for detecting bacterial DNA.
10. The DNA detection method driven by the friction nano generator is characterized by comprising the following steps of:
(1) Soaking the resistance type biosensor in a DNA solution to be detected, and incubating for 45min at 37 ℃;
(2) Taking out the sensor, soaking the sensor in a carbon nano tube solution modified with a DNA signal probe, and incubating for 45min at 37 ℃;
(3) Taking out the sensor, lightly flushing the unbound carbon nanotubes with deionized water, and naturally air-drying;
(4) The friction nano generator is used for driving, and voltage values at two ends of the resistance type biosensor are tested;
(5) And calculating the concentration of the target DNA in the solution to be detected by utilizing a pre-fitted voltage-DNA concentration curve equation.
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