CN110243792A - A kind of fluorescence chemical sensor and its detection method and application based on quantum dot and tetrahedron DNA structure - Google Patents
A kind of fluorescence chemical sensor and its detection method and application based on quantum dot and tetrahedron DNA structure Download PDFInfo
- Publication number
- CN110243792A CN110243792A CN201910502257.XA CN201910502257A CN110243792A CN 110243792 A CN110243792 A CN 110243792A CN 201910502257 A CN201910502257 A CN 201910502257A CN 110243792 A CN110243792 A CN 110243792A
- Authority
- CN
- China
- Prior art keywords
- dna
- texas red
- stranded
- tetrahedron
- chemical sensor
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells
Abstract
The present invention provides a kind of fluorescence chemical sensor based on quantum dot and tetrahedron DNA structure and its detection method and application, the fluorescence chemical sensor includes a 3 D stereo DNA structure, tetrahedron DNA is constructed by four oligonucleotides, four oligonucleotides are marked with biotin and the fluorescent dye flower mountain valley with clumps of trees and bamboo 3 (Cy3), texas Red and the flower mountain valley with clumps of trees and bamboo 5 respectively.It is subsequently assembled on the quantum dot of Streptavidin modification, obtains a QD-Cy3-Texas Red-Cy5 tetrahedron DNA up to fluorescence chemical sensor.The fluorescence chemical sensor has space length between specific dyestuff, and the energy transfer between median receptor and terminal receptor has high controllability, the fluorescence chemical sensor can generate the multistep FRET between QD and three color dyestuffs, the detection of a variety of enzymes can be realized as multiple interactive paths FRET of donor using QD and screen inhibitor, the value with good practical application.
Description
Technical field
The invention belongs to biological detections and technical field of molecular biology, and in particular to one kind is based on quantum dot and four sides
The fluorescence chemical sensor and its detection method of body DNA structure and application.
Background technique
Disclosing the information of the background technology part, it is only intended to increase understanding of the overall background of the invention, without
It is existing well known to persons skilled in the art so to be considered as recognizing or imply that information composition has become in any form
Technology.
People design complicated three dimensional DNA paper folding nanostructure, thus in bio-sensing, optics, and drug delivery and gene
Adjusting aspect has extensive use.However, it is found by the inventors that being mostly based on the molecular detection system of three dimensional DNA nanostructure
It is merely able to a type of molecule, and often relates to complicated logical operation.
Usually using fluorescent organic dyes in fluorescence resonance energy transfer (FRET) system, however, fluorescent organic dyes are easy
In photobleaching, fluorescence lifetime is shorter, and spectra overlap is serious and has high background fluorescence in the biological sample.In order to overcome fluorescence
A large amount of Illuminant nanometer materials are widely applied in some limitations of organic dyestuff, people, such as: quantum dot (QD), bar code particle
The materials such as the nano SiO 2 particle of son, polymer nano granules and organic dyestuff package.Inventors have found that semiconductor amount
Son point has long fluorescence lifetime, from the wide absorption spectrum of ultraviolet-near-infrared, narrow and adjustable emission spectrum, and resistance toization
The features such as corrosion, photism is good.And under single light source excitation, single quantum dot, which can couple one or more fluorescence, to be had
Engine dyeing material is as FRET receptor.Therefore, QD is realized as the ideal donor of multiple FRET configuration.
Summary of the invention
In view of the above shortcomings of the prior art, the present invention provides a kind of fluorescence based on quantum dot and tetrahedron DNA structure
Chemical sensor and its detection method and application, the fluorescence chemical sensor include a novel 3 D stereo DNA structure,
Tetrahedron DNA is constructed by four oligonucleotides, and four oligonucleotides are marked with biotin (biotin) and fluorescent dye flower respectively
The mountain valley with clumps of trees and bamboo 3 (Cy3), texas Red (Texas Red) and the flower mountain valley with clumps of trees and bamboo 5 (Cy5).It is subsequently assembled to the quantum dot of Streptavidin modification
(QD) on, a QD-Cy3-Texas Red-Cy5 tetrahedron DNA i.e. fluorescence chemical sensor is obtained.Fluorescence chemical sensing
Device has space length between specific dyestuff, and the energy transfer between median receptor and terminal receptor has height can
Control property, which can generate the multistep between QD and three color dyestuffs (Cy3, Texas Red, and Cy5)
FRET can realize the detection of a variety of enzymes as multiple interactive paths FRET of donor using QD and screen inhibitor have good
The value of good practical application.
The present invention is achieved through the following technical solutions:
The first aspect of the invention provides a kind of based on the fluorescence chemical of quantum dot and tetrahedron DNA structure sensing
Device, the fluorescence chemical sensor include at least quantum dot and the tetrahedron DNA structure for being assembled in the quantum dot surface, institute
It states tetrahedron DNA to be constructed by four oligonucleotides, four oligonucleotides are marked with biotin (biotin) and different glimmering respectively
Photoinitiator dye.
The fluorescent dye includes but is not limited to the flower mountain valley with clumps of trees and bamboo 3 (Cy3), texas Red (Texas Red) and the flower mountain valley with clumps of trees and bamboo 5
(Cy5)。
At least there is a kind of restriction enzyme site of enzyme to be measured on one oligonucleotide chain.
The second aspect of the invention is provided based on above-mentioned fluorescence chemical sensor while the method that detects a variety of enzymes, institute
The method of stating includes:
S1, the above-mentioned tetrahedron DNA structure of building；
S2, above-mentioned tetrahedron DNA structure is added in testing sample solution, carries out incubation reaction, quantum is then added
Point forms 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA nanostructure；
S3, Single Molecule Detection fluorescence signal, a variety of enzyme contents to be measured of quantitative analysis.
The third aspect of the invention provides above-mentioned fluorescence chemical sensor and/or detection method in enzyme quantitative detection
And/or the application in inhibitor sifting.
The invention has the advantages that:
(1) the present invention is based on the fluorescence chemical sensors in the entire reaction process detected to enzyme to be measured not
Need complicated instrument or red tape；Meanwhile extremely simple, low in cost, simple operation is operated, while not being related to patrolling
Collect operation, effectively save cost.
(2) fluorescence chemical sensor of the present invention is using combination tetrahedron DNA and 525QD, and it is logical to be combined multiple FRET
Road further can carry out the sieve of enzyme inhibitor so as to three kinds of highly sensitive quantitative detection even a variety of enzymes simultaneously simultaneously
Choosing, therefore the prospect with good practical application.
Detailed description of the invention
The Figure of description for constituting a part of the invention is used to provide further understanding of the present invention, of the invention
Illustrative embodiments and their description are used to explain the present invention, and are not constituted improper limitations of the present invention.
Fig. 1 is the mechanism figure that fluorescence chemical sensor detects a variety of different restriction endonucleases in the embodiment of the present invention 1；
Fig. 2 (A) is the biotin-Cy3-Texas Red-Cy5 tetrahedron DNA polypropylene for not carrying out SYBR Gold dyeing
Acrylamide gel electrophoretogram；Fig. 2 (B) is the biotin-Cy3-Texas Red-Cy5 tetrahedron DNA for carrying out SYBR Gold dyeing
Polyacrylamide gel electrophoresis figure；Fig. 2 (C) is 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA agarose gel electrophoresis
Figure；Wherein band 1 is biotin-Cy3-Texas Red-Cy5 tetrahedron DNA electrophoretic image figure, and band 2 is Streptavidin package
525QD electrophoretic image figure, band 3 be 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA electrophoretic image figure, band 4, band 5, band
6 be electrophoretic image figure when adding HaeIII, EcoRV and PvuII respectively；Band 7 is electrophoretic image figure in the presence of three kinds of enzymes are equal.Figure
3 (A) are 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA fluorescence spectra；Insertion portion is in acceptor emission spectral regions
Between fluorescence spectrum；Fig. 3 (B) forms FRET efficiency between tetrahedron DNA for not isoacceptor and compares figure.
Fig. 4 (A) is 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA single molecular imaging figure；Fig. 4 (B) is to exist
Tetrahedron DNA single molecular imaging figure when HaeIII；Fig. 4 (C) is that there are mono- points of tetrahedron DNA when HaeIII, EcoRV and PvuII
Sub- image；Fig. 4 (D) is HaeIII enzyme cleavage reaction fluorescence spectra；Fig. 4 (E) is that the HaeIII of various concentration corresponds to Cy5
The variation of fluorescence intensity decreasing value, illustration are the line between the fluorescence intensity decreasing value of Cy5 and the logarithmic form of HaeIII concentration
Sexual intercourse figure；Fig. 4 (F) is EcoRV enzyme cleavage reaction fluorescence spectra；Fig. 4 (G) is that the EcoRV of various concentration corresponds to Texas
The variation diagram of the fluorescence intensity decreasing value of Red, illustration are the fluorescence intensity decreasing value of Texas Red and the logarithm of EcoRV concentration
Linear relationship chart between form；Fig. 4 (H) is PvuII enzyme cleavage reaction fluorescence spectra；Fig. 4 (I) is various concentration
PvuII corresponds to the variation of FRET efficiency decreasing value, and illustration is between FRET efficiency decreasing value and the logarithmic form of PvuII concentration
Linear relationship.
Fig. 5 (A) is that different specificity experiments groups correspond to FRET efficiency chart；Wherein, (a) normal HaeIII+EcoRV+
PvuII reaction group；(b) heat inactivation HaeIII+EcoRV+PvuII reaction group；Fig. 5 (B) is that HaeIII methylase reacts fluorescence
Spectrogram, HaeIII concentration are 0.5 fixed every microlitre of unit；Fig. 5 (C) is the HaeIII transmethylase pair of various concentration
The variation diagram of the fluorescence intensity raising value of Cy5 is answered, the fluorescence intensity raising value and HaeIII transmethylase that illustration is Cy5 are dense
Linear relationship chart between the logarithmic form of degree；Fig. 5 (D) is HaeIII under different EDTA concentration, EcoRV and PvuII activity is closed
System's figure.
Specific embodiment
It is noted that following detailed description is all illustrative, it is intended to provide further instruction to the application.Unless
Otherwise indicated, all technical and scientific terms used herein has and the application person of an ordinary skill in the technical field
Normally understood identical meanings.
It should be noted that term used herein above is merely to describe specific embodiment, and be not intended to restricted root
According to the illustrative embodiments of the application.As used herein, unless the context clearly indicates otherwise, otherwise singular shape
Formula be also intended to include plural form, additionally, it should be understood that, when in the present specification use term "comprising" and/or
When " comprising ", existing characteristics, step, operation, device, component and/or their combination are indicated.
As previously mentioned, people design complicated three dimensional DNA paper folding nanostructure, thus in bio-sensing, optics, drug
Have in terms of transmitting and Gene regulation and is widely applied.However, it is found by the inventors that being mostly based on point of three dimensional DNA nanostructure
Sub- detection system is merely able to a type of molecule, and often relates to complicated logical operation.
In view of this, of the invention develops a kind of novel 3 D stereo DNA structure, tetrahedron DNA is by four few cores
Thuja acid building, four oligonucleotides are marked with biotin (biotin) and the fluorescent dye flower mountain valley with clumps of trees and bamboo 3 (Cy3), texas Red respectively
(Texas Red) and the flower mountain valley with clumps of trees and bamboo 5 (Cy5).It is subsequently assembled on the quantum dot (QD) of Streptavidin modification, obtains a QD-
Cy3-Texas Red-Cy5 tetrahedron DNA, that is, fluorescence chemical sensor.The fluorescence chemical sensor have specific dyestuff it
Between space length, and the energy transfer between median receptor and terminal receptor has high controllability, which can produce
Multistep FRET between raw QD and three color dyestuffs (Cy3, Texas Red, and Cy5), can be using QD as the multiple of donor
It realizes the detection of a variety of enzymes and screens inhibitor in the interaction path FRET.
In the specific embodiment of the present invention, provide a kind of Fluoresceinated based on quantum dot and tetrahedron DNA structure
Sensor is learned, the fluorescence chemical sensor includes at least quantum dot and the tetrahedron DNA knot for being assembled in the quantum dot surface
Structure, the tetrahedron DNA are constructed by four oligonucleotides, and four oligonucleotides are marked with biotin (biotin) and not respectively
Same fluorescent dye.
In still another embodiment of the invention, the fluorescent dye includes but is not limited to the flower mountain valley with clumps of trees and bamboo 3 (Cy3), De Kesa
This red (Texas Red) and the flower mountain valley with clumps of trees and bamboo 5 (Cy5).
In still another embodiment of the invention, at least there is a kind of enzyme to be measured on a kind of oligonucleotide chain
Restriction enzyme site；
In still another embodiment of the invention, the quantum dot is coated with Streptavidin, thus with biology is marked with
The oligonucleotides of element connects and then makes the assembling of tetrahedron DNA structure over the qds；
In still another embodiment of the invention, the quantum dot is 525QD；
In still another embodiment of the invention, when restriction endonuclease to be measured is HaeIII, EcoRV and PvuII；
In still another embodiment of the invention, the base sequence of four oligonucleotides is as follows:
(1) DNA of biotin labeling is single-stranded: 5 '-CTA TGT GGC CAA TCA AAC GAG AGC AAG TGT ATG
AGT AAG ATC GCG ACC AT–biotin-3'(SEQ ID NO.1)；
(2) DNA of Cy3 label is single-stranded: 5 '-CAT GGG ATA TCT ACG GAC ATA CAC TTG CTC TCG
AAG ACT TCA GCT GGT TA-Cy3-3'(SEQ ID NO.2)；
(3) DNA of Texas Red label is single-stranded: 5 '-Texas Red-CCG TAG ATA TCC CAT GAG TTG
AGC CTG GAC AGG AAT GGT CGC GAT CTT AC-3'(SEQ ID NO.3)；
(4) DNA of Cy5 label is single-stranded: 5 '-Cy5-TTG ATT GGC CAC ATA GAC CTG TCC AGG CTC
AAC ATA ACC AGC TGA AGT CT-3’(SEQ ID NO.4)。
In still another embodiment of the invention, provides based on above-mentioned fluorescence chemical sensor while detecting a variety of enzymes
Method, which comprises
S1, the above-mentioned tetrahedron DNA structure of building；
S2, above-mentioned tetrahedron DNA structure is added in testing sample solution, carries out incubation reaction, quantum is then added
Point forms 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA nanostructure；
S3, Single Molecule Detection fluorescence signal, a variety of enzyme contents to be measured of quantitative analysis.
In still another embodiment of the invention, in the step S1,
Construct tetrahedron DNA structure method particularly includes: prepare a variety of single stranded DNAs, including different length biotin labeling
DNA is single-stranded；The DNA of Cy3 label is single-stranded；The DNA of Texas Red label is single-stranded；The DNA of Cy5 label is single-stranded；And with it is above-mentioned
DNA single stranded sequence is identical, but the DNA of unmarked dyestuff single-stranded (sp chain).Required DNA is added to 1 × CutSmart buffer
In, mixture is added to 92~96 DEG C (preferably 95 DEG C) 5~15 minutes (preferably 10 minutes)；Be subsequently placed on ice 0.5~
1.5 hours (preferably 1 hour)；
In still another embodiment of the invention, the tetrahedron DNA sequence dna for being self-assembled into 16 base-pair side lengths is as follows:
(1) DNA of biotin labeling is single-stranded: 5 '-CTA TGT GGC CAA TCA AAC GAG AGC AAG TGT ATG
AGT AAG ATC GCG ACC AT–biotin-3'(SEQ ID NO.1)；
(2) DNA of Cy3 label is single-stranded: 5 '-CAT GGG ATA TCT ACG GAC ATA CAC TTG CTC TCG
AAG ACT TCA GCT GGT TA-Cy3-3'(SEQ ID NO.2)；
(3) DNA of Texas Red label is single-stranded: 5 '-Texas Red-CCG TAG ATA TCC CAT GAG TTG
AGC CTG GAC AGG AAT GGT CGC GAT CTT AC-3'(SEQ ID NO.3)；
(4) DNA of Cy5 label is single-stranded: 5 '-Cy5-TTG ATT GGC CAC ATA GAC CTG TCC AGG CTC
AAC ATA ACC AGC TGA AGT CT-3’(SEQ ID NO.4)。
In still another embodiment of the invention, in the step S2,
Incubation reaction condition specifically: 30~40 DEG C (preferably 37 DEG C) reactions, 0.5~1.5 hour (preferably 1 hour)；
In still another embodiment of the invention, in the step S3, it is based on full interior angle reflected fluorescent light technology (TIRF)
Single molecular imaging system detected.Specifically, excitation light source is 405 nano lasers, excitation intensity is 30 milliwatts.
525QD uses 500-550 nanometers of optical filter, and Cy3/Texas Red is received using 573-617 nanometers of optical filter and 565-605
The optical filter of rice, Cy5 use 672-712 nanometers of optical filter.
In still another embodiment of the invention, it is fixed in enzyme to provide above-mentioned fluorescence chemical sensor and/or detection method
Application in amount detection and/or inhibitor sifting.
Explanation is further explained to the present invention by the following examples, but is not construed as limiting the invention.It should be understood that
These examples are only for illustrating the present invention and are not intended to limit the scope of the present invention.In addition, unspecified in embodiment
Molecular biology method is the method for this field routine, and concrete operations can be referring to molecular biosciences guide or product description.
Embodiment 1
1. testing principle of the present invention and method and step:
The experimental principle of the technology is marked with biotin, Cy3, four DNA widows of Texas Red and Cy5 as shown in Figure 1:
Nucleotide is self-assembly of biotin-Cy3-Texas Red-Cy5 tetrahedron DNA.Then, biotin- made of self assembly
Cy3-Texas Red-Cy5 tetrahedron DNA is connected to Streptavidin package by special biotin and Streptavidin
The surface QD forms 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA.When 392 nanometers of excitation wavelengths excite tetrahedron DNA,
The fluorescence signal of tetra- kinds of 525QD, Cy3, Texas Red, Cy5 fluorescent materials can be observed simultaneously.The system includes QD-
The FRET of Cy3, QD-Texas Red, QD-Cy5, Cy3-Texas Red, Cy3-Cy5, Texas Red-Cy5.Wherein, QD-
The FRET of Cy3 is beneficial to the fluorescent emission signals of Cy3；The FRET of QD-Texas Red and Cy3-Texas Red are beneficial to
The fluorescent emission signals of Texas Red；The FRET of QD-Cy5, Cy3-Cy5 and Texas Red-Cy5 are beneficial to the fluorescence of Cy5
Emit signal.
The 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA of the Technology design, each side are the double-strand of 16 base-pairs
DNA, the catalytic substrate of each Bian Douyou enzyme.There is the catalytic site of PvuII nickase at 7 base-pairs of distance Cy3,
There is the catalytic site of EcoRV nickase at 8 base-pairs of distance Texas Red, has at 8 base-pairs of distance Cy5
The catalytic site of HaeIII nickase.In the presence of only a kind of restriction endonuclease (by taking HaeIII as an example), it is anti-that HaeIII causes digestion
The DNA fragmentation that Cy5 should be caused to mark is separated with tetrahedron, subsequently, as QD-Cy5, Cy3-Cy5 and Texas Red-Cy5
The signal of the disappearance of FRET access, Cy5 is reduced or is disappeared.When there are two types of restriction endonucleases (by taking HaeIII and EcoRV as an example) to exist
When, the DNA fragmentation that HaeIII causes endonuclease reaction and Cy5 is caused to mark is separated with tetrahedron, and EcoRV, which causes endonuclease reaction, to be caused
The DNA fragmentation of Texas Red label is separated with tetrahedron.Therefore, because QD-Cy5, Cy3-Cy5 and Texas Red-Cy5
The signal of the disappearance of FRET access, Cy5 is reduced or is disappeared；Due to the FRET of QD-Texas Red and Cy3-Texas Red
The signal of the disappearance of access, Texas Red is reduced or is disappeared.When three kinds of restriction endonucleases (HaeIII, EcoRV and PvuII) exist
When, the DNA fragmentation for by the endonuclease reaction that HaeIII, EcoRV and PvuII are caused Cy3 being marked, Texas Red label
The DNA fragmentation of DNA fragmentation and Cy5 label is separated from tetrahedron DNA, due to QD-Cy3, QD-Texas Red, QD-Cy5Cy3-
The disappearance of FRET access between Texas Red, Cy3-Cy5 and Texas Red-Cy5, can not observe Cy3 signal again,
Texas Red signal and Cy5 signal.Indicate that HaeIII, Texas Red indicate that EcoRV, Cy3 indicate PvuII by Cy5, it should
Technology can be realized while detect 3 kinds of enzymes and screens a variety of enzyme inhibitors.
Building tetrahedron DNA: prepare a variety of single stranded DNAs respectively, the DNA including different length biotin labeling is single-stranded；
The DNA of Cy3 label is single-stranded；The DNA of Texas Red label is single-stranded；The DNA of Cy5 label is single-stranded；With above-mentioned DNA single stranded sequence phase
Together, but the DNA of unmarked dyestuff single-stranded (sp chain).Required DNA (1 every liter of micromole) is added to 1 × CutSmart buffer
In, mixture is added to 95 degrees Celsius 10 minutes.It is subsequently placed with 1 hour on ice.
It is self-assembled into the tetrahedron DNA sequence dna of 16 base-pair side lengths:
(1) DNA of biotin labeling is single-stranded: 5 '-CTA TGT GGC CAA TCA AAC GAG AGC AAG TGT ATG
AGT AAG ATC GCG ACC AT–biotin-3'(SEQ ID NO.1)；
(2) DNA of Cy3 label is single-stranded: 5 '-CAT GGG ATA TCT ACG GAC ATA CAC TTG CTC TCG
AAG ACT TCA GCT GGT TA-Cy3-3'(SEQ ID NO.2)；
(3) DNA of Texas Red label is single-stranded: 5 '-Texas Red-CCG TAG ATA TCC CAT GAG TTG
AGC CTG GAC AGG AAT GGT CGC GAT CTT AC-3'(SEQ ID NO.3)；
(4) DNA of Cy5 label is single-stranded: 5 '-Cy5-TTG ATT GGC CAC ATA GAC CTG TCC AGG CTC
AAC ATA ACC AGC TGAAGT CT-3’(SEQ ID NO.4)。
Endonuclease reaction: 0.12 every liter of micromole tetrahedron DNA is added in 1 × CutSmart buffer, is then added not
With nickase HaeIII, EcoRV and the HaeIII of concentration.Mixture system reacts 1 hour at 37 degrees Celsius.Then, reactant
The 525QD of the Streptavidin package of 5 every liter of nanomoles is added in system.To form 525QD-Cy3-Texas Red-Cy5 tetra-
Face body DNA nanostructure.
Methylase reaction: 0.12 every liter of micromole tetrahedron DNA is added in 1 × CutSmart buffer, is then added
It is single to be eventually adding 0.5 for the HaeIII transmethylase for entering every liter of S-adenosylmethionine of 60 micromole (SAM) and various concentration
The every microlitre of HaeIII in position.Mixture system reacts 1 hour at 37 degrees Celsius.Then, 5 every liter of nanomoles are added in reaction system
Streptavidin package 525QD after carry out spectral measurement.
Biofluid endonuclease reaction: 0.12 every liter of micromole tetrahedron DNA is added in 1 × CutSmart buffer, with
The nickase of various concentration and the fetal calf serum of 10% volume fraction are added afterwards, mixture system is small in 37 degrees Celsius of reactions 1
When.
Inhibitors experiment: prepare 525QD-Cy3-Texas Red-Cy5,525QD-Cy3-Texas Red-sp, 525QD-
Cy3-sp-sp is respectively used to HaeIII, EcoRV and PvuII inhibitors experiment.0.12 every liter of micromole tetrahedron DNA is added
Into 1 × CutSmart buffer, nickase corresponding to various concentration inhibitor and different type tetrahedron DNA is added, cuts
Cutting enzyme concentration is 0.5 every microlitre of unit.Mixture system reacts 1 hour at 37 degrees Celsius.
Gel electrophoresis: using 8% polyacrylamide gel electrophoresis to unused SYBR Gold DNA indicator dyeing and
It is imaged using the biotin-Cy3-Texas Red-Cy5tetrahedral DNA that SYBR Gold DNA indicator dyes,
Reaction system is tbe buffer liquid (44.5 mMs of every liter of Tris- boric acid, 1 mM of every liter of EDTA), and electrophoresis time is 45 points
Clock.525QD-Cy3-Texas Red-Cy5 tetrahedron DNA is imaged using 1% agarose gel electrophoresis, reaction system is
TAE buffer (40 mMs of every liter of Tris- acetic acid, 2 mMs of every liter of EDTA), electrophoresis time are 30 minutes.Finally by solidifying
Glue imaging system carries out gel imaging.525QD-Cy3-Texas Red-Cy5 tetrahedron DNA uses multiple excitation light sources, excitation
When light source is blue light (460-490 nanometers of excitation wavelength), 525QD, 525QD is excited to use 518-546 nm filter.Excitation
When light source is green light (520-545 nanometers of excitation wavelength), Cy3 perhaps Texas Red fluorophor Cy3 or Texas is excited
Red fluorophor uses 577-613 nm filter.When excitation light source is feux rouges (625-650 nanometers of excitation wavelength), excitation
Cy5 fluorophor, 525QD use 675-725 nm filter.
Fluorescence measurement: fluorescence signal collects fluorescence signal, excitation wavelength 392nm using luminoscope measurement.It collects respectively
The emission peak intensity of 530 nanometers, 565 nanometers, 613 nanometers and 670 nanometers, respectively corresponds 525QD, Cy3, Texas Red and
The fluorescence intensity of Cy5.
Single Molecule Detection: unimolecule is carried out using the utilizing total internal reflection fluorescence microscope equipped with oil mirror and inverted microscope
Imaging.Excitation light source is 405 nano lasers, and excitation intensity is 30 milliwatts.525QD uses 500-550 nanometers of optical filter,
Cy3/Texas Red uses 573-617 nanometers of optical filter and 565-605 nanometers of optical filter, and Cy5 uses 672-712 nanometers
Optical filter.Then, data analysis is carried out to the region for collecting 70 × 70 pixels.Test result analysis
1. gel electrophoresis analysis
The present embodiment uses the tetrahedron DNA of non denatured polyacrylamide gel (PAGE) electrophoretic analysis self assembly.Such as
Shown in Fig. 2, in Fig. 2A and Fig. 2 B, (Fig. 2A is with 1,2,3 for single stranded DNA；Fig. 2 B band 1) it is fastest in gel electrophoresis, two
(Fig. 2A is with 4-13 for chain or three chain Self-assembled DNAs；Fig. 2 B band 2,3) speed are then slower than single stranded DNA, four DNA self assemblies
(Fig. 2A is with 14；Fig. 2 B band 4) speed is most slow.And every band shows biotin-Cy3-Texas Red-Cy5 tetra- all without miscellaneous band
Face body DNA success self assembly.
The present embodiment analyzes 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA using agarose gel electrophoresis.Scheming
In 2C, 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA (Fig. 2 C band 3) and the four sides biotin-Cy3-Texas Red-Cy5
Body DNA (Fig. 2 C band 1) position is different and without miscellaneous band, shows that biotin-Cy3-Texas Red-Cy5 tetrahedron DNA is successfully assembled
To the surface 525QD.
HaeIII (Fig. 2 C band 4) is added then to biotin-Cy3-Texas Red-Cy5 tetrahedron DNA, EcoRV (figure
2C band 5), PvuII (Fig. 2 C, band 6) and HaeIII+EcoRV+PvuII (Fig. 2 C band 7), it may be observed that carry Cy5, Texas
The small-sized segment and 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA of Red, Cy3 and Cy5+Texas Red+Cy3 subtract
It is few.Show that three kinds of enzymes have caused the cutting of tetrahedron DNA.
2. spectrofluorimetry
The present embodiment then uses the tetrahedron DNA of spectrofluorimetry self assembly, as shown in figure 3, in Fig. 3 A, it is single
Dye acceptor (Cy3) is assembled to the reduction that QD fluorescence intensity is caused on the surface QD.Double dye acceptor (Cy3 and Texas Red) assemblings
Cause the reduction of QD intensity to the surface QD, and reduces amplitude and be greater than single dye acceptor (Cy3).Three dye acceptors are assembled to QD table
The reduction of QD intensity is caused in face, and reduces amplitude and be greater than single dye acceptor (Cy3) and double dye acceptor (Cy3 and Texas
Red).Fig. 3 B shows to change with the FRET efficiency of QD- tetrahedron DNA with the change of acceptor type and acceptor quantity, and
And there are three types of reach maximum when acceptor dye on the surface 525QD.This result is also demonstrated that biotin-Cy3-Texas Red-
The formation of Cy5 tetrahedron DNA and the tetrahedron are successfully assembled to the surface 525QD.
3. Single Molecule Detection is analyzed
It is as shown in Figure 4: as 405 nanometer lasers excitation 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA, to obtain simultaneously
Can to 525 (525QD) and the fluorescence imaging of 595 (Cy3 perhaps Texas Red) nanometers while also obtain 585 (Cy3 or
Texas Red) and 692 (Cy5) nanometers fluorescence imaging.In the absence of enzyme, it may be observed that the fluorescence of apparent 525QD
Signal (Fig. 5 A, a) with Cy3/Texas Red fluorescence signal (Fig. 5 A, b and d) and Cy5 signal (Fig. 5 A, e).This result is same
Sample shows biotin-Cy3-Texas Red-Cy5 tetrahedron DNA success self assembly.
In the presence of HaeIII, it may be observed that (Fig. 5 B a) believes with Cy3/Texas Red fluorescence the fluorescence signal of 525QD
Number (Fig. 5 B, b and d), however can not observe Cy5 signal (Fig. 5 B, e).When HaeIII+EcoRV+PvuII is existed simultaneously,
It can only observe that (Fig. 5 C a), and can not observe Cy3 or Texas Red (Fig. 5 C, b and d) or Cy5 for the signal of 525QD
Fluorescence signal (Fig. 5 C, e).This result again shows that enzyme has caused the cutting of tetrahedron DNA.
4. endonuclease minimal detectable concentration
The signal that the cleavage reaction that HaeIII, EcoRV and PvuII cause generates can be as the function as enzyme concentration.
There is the reduction for causing Cy5 fluorescence signal intensity in HaeIII, and with the increase of QD fluorescence intensity signals (figure
4D).Meanwhile HaeIII concentration is higher, the reduction of Cy5 signal strength is bigger (Fig. 4 E).The reduction of Cy5 fluorescence intensity with
HaeIII has good linear relationship (Fig. 4 E illustration), related letter between the logarithm of every microlitre of concentration of 0.025 to 0.5 unit
Number is Δ I=107.96+51.85log10C(R2=0.990), the concentration that wherein C is HaeIII, Δ I are that the reduction of Cy5 is strong
Degree.Add the standard deviation of three times to obtain detection by calculating average value and is limited to 0.0112 every microlitre of unit.
Fig. 4 F indicates that there are glimmering accordingly when the EcoRV of fixed concentration HaeIII (0.5 every microlitre of unit) and various concentration
Light spectrum.There is no when EcoRV, it may be observed that apparent Texas Red signal (Fig. 4 F) shows to occur QD in Texas Red
Between FRET.In contrast, there is the reduction for causing Texas Red fluorescence signal intensity and with QD fluorescence intensity in EcoRV
The increase (Fig. 4 F) of signal, meanwhile, EcoRV concentration is higher, and the reduction of Texas Red signal strength is bigger (Fig. 4 G).Texas
The reduction of Red fluorescence intensity and EcoRV have good linear relationship between the logarithm of every microlitre of concentration of 0.025 to 0.5 unit
(Fig. 4 G illustration), correlation function are Δ I=78.13+42.43log10C(R2=0.998), wherein C is EcoRV concentration, and Δ I is
The reduction intensity of Texas Red, detection are limited to 0.0233 every microlitre of unit.
Fig. 4 H indicates that (0.5 unit is per micro- there are fixed concentration HaeIII (0.5 every microlitre of unit) and fixed concentration EcoRV
Rise) and various concentration PvuII when corresponding fluorescence spectrum.There is no when PvuII, it may be observed that apparent Cy3 signal
(Fig. 4 H), shows that FRET occurs between QD and Cy3.In contrast, there is the reduction of initiation Cy3 fluorescence signal intensity simultaneously in PvuII
With the increase (Fig. 4 H) of QD fluorescence intensity signals, meanwhile, FRET efficiency increases significantly (figure with the increase of PvuII intensity
4I).FRET efficiency and PvuII have good linear relationship (Fig. 4 I between the logarithm of every microlitre of concentration of 0.025 to 0.5 unit
Illustration), correlation function is Δ E=7.69+3.92log10 C(R2=0.997), wherein C is PvuII concentration, and Δ E is that FRET is strong
Spend reduction amount.Detection is limited to 0.0233 every microlitre of unit.
5. tetrahedron specificity is analyzed
In order to further verify specificity of the 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA to restriction endonuclease, this reality
Applying example uses enzyme XhoI and KpnI as negative control group.XhoI can be identified and be cut 5 '-CTCGAG-3 ' sequence, and KpnI can
It identifies and cuts 5 '-GGTACC-3 ' sequence.The rwo enzyme can not identify and cut 525QD-Cy3-Texas RedCy5 tetra-
Face body DNA.Consistent as expected, XhoI and KpnI control group does not have apparent FRET signal to reduce (Fig. 5 A).Meanwhile in heating
There is also apparent signals to reduce (Fig. 5 A b) for enzyme cutting HaeIII+EcoRV+PvuII experimental group, the reason is that heating can make
HaeIII and EcoRV inactivation, but PvuII but has certain resistance to high temperature.With above-mentioned phenomenon on the contrary, being added active
HaeIII+ EcoRV+PvuII when, it may be observed that apparent FRET reduces (Fig. 5 A a).This result is strong to be shown
Specificity of the 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA to restriction endonuclease.
6. methyl transferase activity is analyzed
It is living that 525QD-Cy3-Texas Red-Cy5 tetrahedron DNA equally can further detect HaeIII transmethylase
Property.HaeIII transmethylase can be catalyzed the methylation of 5 '-GGCC-3 ' sequence and generate 5 '-GGmCC-3 ' sequence.And HaeIII
It can not identify and cut 5 '-GGmCC-3 ' sequence.When HaeIII transmethylase is not present, 5 '-GGCC-3 ' sequence can not
Methylation cuts to be identified by HaeIII, therefore can not observe Cy5 signal (Fig. 5 B)
When there are HaeIII transmethylase, 525QD fluorescence intensity is reduced, and Cy5 fluorescence intensity increases (Fig. 5 B).
HaeIII transmethylase concentration is higher, and Cy5 fluorescence signal is stronger (Fig. 5 C), Cy5 fluorescence intensity and HaeIII transmethylase
Have between the logarithm of concentration good linear relationship (Fig. 5 C illustration), correlation function is Δ I=70.47+31.45log10C(R2
=0.993), wherein C is the concentration of HaeIII transmethylase, and Δ I is the intensity of Cy5.Detection is limited to 0.01 unit per micro-
It rises.This is the result shows that the tetrahedron equally can detecte other kinds of enzymatic activity.
7. inhibitors experiment
525QD-Cy3-Texas Red-Cy5 tetrahedron DNA can equally be further used for screening enzyme inhibitor.This reality
Applying example uses dehydration disodium EDTA (EDTA) as model inhibitor.By measuring 525QD-Cy3-Texas
The intensity of Cy5 in Red-Cy5 tetrahedron DNA, can verify inhibition of the EDTA to HaeIII.By measuring 525QD-Cy3-
The intensity of Texas Red in Texas Red-sp can verify inhibition of the EDTA to EcoRV.By measuring 525QD-Cy3-
The intensity of Cy3 in sp-sp can verify inhibition of the EDTA to PvuII.HaeIII, EcoRV and PvuII activity intensity can be made
IC is used for function (Fig. 5 D) the present embodiment of EDTA concentration50The value of (half inhibitor) is as the finger for measuring inhibitory effect
Mark.IC of the EDTA inhibitor to different enzymes50Show following trend: PvuII (26.02 mMs every liter) < HaeIII (33.23 millis
Mole every liter) < EcoRV (49.65 mMs every liter).
It should be noted that above example is only used to illustrate the technical scheme of the present invention rather than is limited.Although ginseng
It is described the invention in detail according to given example, but those skilled in the art can be as needed to this
The technical solution of invention is modified or replaced equivalently, without departing from the spirit and scope of the technical solution of the present invention.
SEQUENCE LISTING
<110>Shandong Normal University
<120>a kind of fluorescence chemical sensor and its detection method and application based on quantum dot and tetrahedron DNA structure
<130>
<160> 4
<170> PatentIn version 3.3
<210> 1
<211> 50
<212> DNA
<213>artificial synthesized
<400> 1
ctatgtggcc aatcaaacga gagcaagtgt atgagtaaga tcgcgaccat 50
<210> 2
<211> 50
<212> DNA
<213>artificial synthesized
<400> 2
catgggatat ctacggacat acacttgctc tcgaagactt cagctggtta 50
<210> 3
<211> 50
<212> DNA
<213>artificial synthesized
<400> 3
ccgtagatat cccatgagtt gagcctggac aggaatggtc gcgatcttac 50
<210> 4
<211> 50
<212> DNA
<213>artificial synthesized
<400> 4
ttgattggcc acatagacct gtccaggctc aacataacca gctgaagtct 50
Claims (10)
1. a kind of fluorescence chemical sensor based on quantum dot and tetrahedron DNA structure, which is characterized in that the fluorescence chemical passes
Sensor includes at least quantum dot and the tetrahedron DNA structure for being assembled in the quantum dot surface, and the tetrahedron DNA is by four widows
Nucleotide construction, four oligonucleotides are marked with biotin and different fluorescent dyes respectively；
The fluorescent dye includes but is not limited to the flower mountain valley with clumps of trees and bamboo 3, texas Red and the flower mountain valley with clumps of trees and bamboo 5；
At least there is a kind of restriction enzyme site of enzyme to be measured on one oligonucleotide chain.
2. fluorescence chemical sensor as described in claim 1, which is characterized in that the quantum dot is coated with Streptavidin, from
And it is connect with the oligonucleotides for being marked with biotin and then makes the assembling of tetrahedron DNA structure over the qds.
3. fluorescence chemical sensor as described in claim 1, which is characterized in that the quantum dot is 525QD.
4. fluorescence chemical sensor as described in claim 1, which is characterized in that when restriction endonuclease to be measured be HaeIII, EcoRV and
When PvuII；
The base sequence of four oligonucleotides is as follows:
The DNA of biotin labeling is single-stranded: 5 '-CTA TGT GGC CAA TCA AAC GAG AGC AAG TGT ATG AGT
AAG ATC GCG ACC AT–biotin-3'(SEQ ID NO.1)；
The DNA of Cy3 label is single-stranded: 5 '-CAT GGG ATA TCT ACG GAC ATA CAC TTG CTC TCG AAG ACT
TCA GCT GGT TA-Cy3-3'(SEQ ID NO.2)；
The DNA of Texas Red label is single-stranded: 5 '-Texas Red-CCG TAG ATA TCC CAT GAG TTG AGC CTG
GAC AGG AAT GGT CGC GAT CTT AC-3'(SEQ ID NO.3)；
The DNA of Cy5 label is single-stranded: 5 '-Cy5-TTG ATT GGC CAC ATA GAC CTG TCC AGG CTC AAC ATA
ACC AGC TGA AGT CT-3’(SEQ ID NO.4)。
5. the method that any one of the claim 1-4 fluorescence chemical sensor detects a variety of enzymes simultaneously, which comprises
S1, building tetrahedron DNA structure；
S2, tetrahedron DNA structure is added in testing sample solution, carries out incubation reaction, quantum dot is then added, formed
525QD-Cy3-Texas Red-Cy5 tetrahedron DNA nanostructure, that is, fluorescence chemical sensor；
S3, Single Molecule Detection fluorescence signal, a variety of enzyme contents to be measured of quantitative analysis.
6. the method as claimed in claim 5 for detecting a variety of enzymes simultaneously, which is characterized in that in the step S1,
Construct tetrahedron DNA structure method particularly includes: prepare a variety of single stranded DNAs, the DNA including different length biotin labeling is mono-
Chain；The DNA of Cy3 label is single-stranded；The DNA of Texas Red label is single-stranded；The DNA of Cy5 label is single-stranded；And it is single-stranded with above-mentioned DNA
Sequence is identical, but the DNA of unmarked dyestuff single-stranded (sp chain)；Required DNA is added in 1 × CutSmart buffer, is mixed
Object is added to 92~96 DEG C (preferably 95 DEG C) 5~15 minutes (preferably 10 minutes)；Be subsequently placed on ice 0.5~1.5 hour it is (excellent
It is selected as 1 hour).
7. the method as claimed in claim 6 for detecting a variety of enzymes simultaneously, which is characterized in that
The tetrahedron DNA sequence dna for being self-assembled into 16 base-pair side lengths is as follows:
The DNA of biotin labeling is single-stranded: 5 '-CTA TGT GGC CAA TCA AAC GAG AGC AAG TGT ATG AGT
AAG ATC GCG ACC AT–biotin-3'(SEQ ID NO.1)；
The DNA of Cy3 label is single-stranded: 5 '-CAT GGG ATA TCT ACG GAC ATA CAC TTG CTC TCG AAG ACT
TCA GCT GGT TA-Cy3-3'(SEQ ID NO.2)；
The DNA of Texas Red label is single-stranded: 5 '-Texas Red-CCG TAG ATA TCC CAT GAG TTG AGC CTG
GAC AGG AAT GGT CGC GAT CTT AC-3'(SEQ ID NO.3)；
The DNA of Cy5 label is single-stranded: 5 '-Cy5-TTG ATT GGC CAC ATA GAC CTG TCC AGG CTC AAC ATA
ACC AGC TGA AGT CT-3’(SEQ ID NO.4)。
8. the method as claimed in claim 5 for detecting a variety of enzymes simultaneously, which is characterized in that in the step S2,
Incubation reaction condition specifically: 30~40 DEG C (preferably 37 DEG C) reactions, 0.5~1.5 hour (preferably 1 hour).
9. the method as claimed in claim 5 for detecting a variety of enzymes simultaneously, which is characterized in that in the step S3, in complete
The single molecular imaging system of corner reflection fluorescent technique is detected；
Preferably, excitation light source is 405 nano lasers, and excitation intensity is 30 milliwatts；525QD uses 500-550 nanometers of filter
Mating plate, Cy3/Texas Red use 573-617 nanometers of optical filter and 565-605 nanometers of optical filter, and Cy5 uses 672-712
The optical filter of nanometer.
10. being detected simultaneously described in any one of the claim 1-4 fluorescence chemical sensor and/or claim any one of 5-9
Application of the method for a variety of enzymes in enzyme quantitative detection and/or inhibitor sifting.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910502257.XA CN110243792A (en) | 2019-06-11 | 2019-06-11 | A kind of fluorescence chemical sensor and its detection method and application based on quantum dot and tetrahedron DNA structure |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910502257.XA CN110243792A (en) | 2019-06-11 | 2019-06-11 | A kind of fluorescence chemical sensor and its detection method and application based on quantum dot and tetrahedron DNA structure |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN110243792A true CN110243792A (en) | 2019-09-17 |
Family
ID=67886546
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201910502257.XA Pending CN110243792A (en) | 2019-06-11 | 2019-06-11 | A kind of fluorescence chemical sensor and its detection method and application based on quantum dot and tetrahedron DNA structure |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN110243792A (en) |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111982867A (en) * | 2020-07-17 | 2020-11-24 | 山东师范大学 | Ratio type fluorescence sensor and preparation method and application thereof |
| CN112326762A (en) * | 2020-10-29 | 2021-02-05 | 常州大学 | Method for detecting DNA tetrahedron by capillary electrophoresis |
| CN113151401A (en) * | 2020-09-07 | 2021-07-23 | 山东师范大学 | Fluorescence sensor based on quantum dots and three-dimensional hybridization structure and preparation and application thereof |
| CN113185973A (en) * | 2021-04-06 | 2021-07-30 | 上海交通大学 | Preparation method of zero-dimensional nano carbon material/origami structure deoxyribonucleic acid composite material |
| CN113652471A (en) * | 2021-07-23 | 2021-11-16 | 中山大学 | DNA biosensor based on FRET fluorescence ratio and detection method and cell classification method thereof |
| CN113866234A (en) * | 2020-06-30 | 2021-12-31 | 上海健康医学院 | Electrochemical biosensor for detecting FR (FR) based on double-DNA (deoxyribonucleic acid) tetrahedral structure |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1735808A (en) * | 2002-11-07 | 2006-02-15 | 鹿特丹伊拉兹马斯大学 | FRET probes and methods for detecting interacting molecules |
| KR20110122320A (en) * | 2010-05-04 | 2011-11-10 | 경원대학교 산학협력단 | Fret-based nano-sensor system and detection method using the same |
| CN105283560A (en) * | 2013-05-24 | 2016-01-27 | 昆塔波尔公司 | Nanopore-based nucleic acid analysis with mixed FRET detection |
| CN106990084A (en) * | 2017-05-27 | 2017-07-28 | 山东师范大学 | A kind of being used for based on single quantum dot detects the nano-sensor of dnmt rna |
| CN108315421A (en) * | 2018-04-04 | 2018-07-24 | 山东师范大学 | Method of the constant-temperature amplification with the combination of quantum dot fluorescence Resonance energy transfer for detecting a variety of MicroRNAs simultaneously |
-
2019
- 2019-06-11 CN CN201910502257.XA patent/CN110243792A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1735808A (en) * | 2002-11-07 | 2006-02-15 | 鹿特丹伊拉兹马斯大学 | FRET probes and methods for detecting interacting molecules |
| KR20110122320A (en) * | 2010-05-04 | 2011-11-10 | 경원대학교 산학협력단 | Fret-based nano-sensor system and detection method using the same |
| CN105283560A (en) * | 2013-05-24 | 2016-01-27 | 昆塔波尔公司 | Nanopore-based nucleic acid analysis with mixed FRET detection |
| CN106990084A (en) * | 2017-05-27 | 2017-07-28 | 山东师范大学 | A kind of being used for based on single quantum dot detects the nano-sensor of dnmt rna |
| CN108315421A (en) * | 2018-04-04 | 2018-07-24 | 山东师范大学 | Method of the constant-temperature amplification with the combination of quantum dot fluorescence Resonance energy transfer for detecting a variety of MicroRNAs simultaneously |
Non-Patent Citations (1)
| Title |
|---|
| 胡娟等: ""Construction of Tetrahedral DNA-Quantum Dot Nanostructure with the Integration of Multistep Förster Resonance Energy Transfer for Multiplex Enzymes Assay"", 《ACSNANO》 * |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113866234A (en) * | 2020-06-30 | 2021-12-31 | 上海健康医学院 | Electrochemical biosensor for detecting FR (FR) based on double-DNA (deoxyribonucleic acid) tetrahedral structure |
| CN113866234B (en) * | 2020-06-30 | 2024-03-12 | 上海健康医学院 | Electrochemical biosensor for detecting FR based on double DNA tetrahedral structure |
| CN111982867A (en) * | 2020-07-17 | 2020-11-24 | 山东师范大学 | Ratio type fluorescence sensor and preparation method and application thereof |
| CN113151401A (en) * | 2020-09-07 | 2021-07-23 | 山东师范大学 | Fluorescence sensor based on quantum dots and three-dimensional hybridization structure and preparation and application thereof |
| CN112326762A (en) * | 2020-10-29 | 2021-02-05 | 常州大学 | Method for detecting DNA tetrahedron by capillary electrophoresis |
| CN112326762B (en) * | 2020-10-29 | 2023-03-21 | 常州大学 | Method for detecting DNA tetrahedron by capillary electrophoresis |
| CN113185973A (en) * | 2021-04-06 | 2021-07-30 | 上海交通大学 | Preparation method of zero-dimensional nano carbon material/origami structure deoxyribonucleic acid composite material |
| CN113185973B (en) * | 2021-04-06 | 2023-04-28 | 上海交通大学 | Preparation method of zero-dimensional nano carbon material/paper folding structure deoxyribonucleic acid composite material |
| CN113652471A (en) * | 2021-07-23 | 2021-11-16 | 中山大学 | DNA biosensor based on FRET fluorescence ratio and detection method and cell classification method thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN110243792A (en) | A kind of fluorescence chemical sensor and its detection method and application based on quantum dot and tetrahedron DNA structure | |
| ES2908919T3 (en) | Fractional primer hybridization chain reaction | |
| US8642260B2 (en) | Single quantum-dot based aptameric nanosensors | |
| US6777184B2 (en) | Detection of nucleic acid hybridization by fluorescence polarization | |
| US11384391B2 (en) | Flourescene energy transfer-based single molecule/ensemble DNA sequencing by synthesis | |
| US6772070B2 (en) | Methods of analyzing polymers using a spatial network of fluorophores and fluorescence resonance energy transfer | |
| EP1402245B1 (en) | Methods for the characterization of single polymers | |
| JP4343682B2 (en) | Methods for the detection and quantification of analytes in complex mixtures | |
| EP1183521B1 (en) | Aptamer-based detection | |
| US6447724B1 (en) | DNA sequencing using multiple fluorescent labels being distinguishable by their decay times | |
| Guo et al. | Hybridization chain reactions on silica coated Qbeads for the colorimetric detection of multiplex microRNAs | |
| US9273345B2 (en) | Methods and compositions for processes of rapid selection and production of nucleic acid aptamers | |
| US20120003648A1 (en) | Signal Multiplexing and Signal Amplification | |
| US7262007B2 (en) | Dye-labeled oligonucleotide for labeling a nucleic acid molecule | |
| CN110398481A (en) | A kind of single quantum dot fluorescence nano-sensor and its preparation method and application based on no enzymatic self assembly | |
| CN105372421B (en) | Avian flu virus detection based on intelligent aqueous gel fluorescence aptamer sensor | |
| US10365213B2 (en) | Reliable fluorescence correction method for two-color measurement fluorescence system | |
| DE69919143T2 (en) | NUCLEIC ACID HYBRIDIZATION TEST PROCEDURE IN SOLUTION | |
| CN111088324B (en) | Single quantum dot nano sensor for detecting methyltransferase and detection method and application thereof | |
| Strohsahl et al. | Towards single-spot multianalyte molecular beacon biosensors | |
| US20070238125A1 (en) | Probe synthesis method for nucleic acid detection | |
| CN106754902B (en) | A kind of fluorescence resonance probe and its application and kit | |
| AU2019214845A1 (en) | Sequential staining for multiplex analyses of tissues and cells | |
| WO2018073872A1 (en) | Method for detecting target nucleic acid molecule | |
| US20100323360A1 (en) | Oligonucleotide arrangements, processes for their employment and their use |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20190917 |
|
| WD01 | Invention patent application deemed withdrawn after publication |