Detection of miRNAs presents a particular challenge because of their limited size, high sequence homo- logy and greatly various expression level. In this work, an ultrasensitive, label-free and isothermal miRNA detect...Detection of miRNAs presents a particular challenge because of their limited size, high sequence homo- logy and greatly various expression level. In this work, an ultrasensitive, label-free and isothermal miRNA detection was developed based on asymmertric hairpin probe, exonuclease I（Exo I） and SYBR Green I. The method employed asymmetric hairpin probe to perform cycled polymerization and Exo I to reduce background signal. In the presence of the target miRNA, the target triggers probe-mediated cycled polymerization reactions to generate lots of dsDNA products. The dsDNA product effectively prevents itself from being degraded by Exo I and permitted the insertion of more fluorescence dye into it to enlarge the fluorescence signal. In the absence of the target miRNA, there was no probe-mediated polymerization reaction, and the probe was digested by Exo I added, which minimized the intercala- tion of fluorescence dye to reduce the background signal. The combination of the probe-mediated cycled polymeriza- tion with the Exo 1-assisted background reduction allows us to achieve a detection limit of 5× 10^-18 mol/L. Owing to its ultrasensitivity, excellent specificity, convenience and low-cost, this method might hold out great promise in miRNA detection.展开更多
Simian virus 40（SV40） is a polyomavirus and can induce a series of different tumors. The recognition of SV40 genome is crucial to tumor diagnosis and gene therapy. Herein, a sensitive and selective colorimetric meth...Simian virus 40（SV40） is a polyomavirus and can induce a series of different tumors. The recognition of SV40 genome is crucial to tumor diagnosis and gene therapy. Herein, a sensitive and selective colorimetric method for sequence-specific recognition of homopyrimidine-homopurine duplex DNA（dsDNA） of SV40（4424-4440, gp6） was established with a hairpin probe based upon the formation of triplex DNA. Hairpin probe 5＇-CCC TAC CCA TTT TTT CTT CTC TTT CCT GGG TAG GGC GGG TTG GG-3＇（HP） containing G-rich sequence and 17-bp triplex-forming sequence was used as the signal probe, which was stem-loop structure alone and exhibited low catalytic activity. Upon its binding to the target duplex of SV40, hairpin probe transferred from stem-loop structttre to parallel triplex DNA, accompanied by the recovery of catalytic activity of DNAzyme and a sharp increase of absorbance. Under optimum conditions, the absorbance was increased proportionally to the concentration of dsDNA over the range from 500 pmol/L to 40.0 nmol/L with a detection limit of 433 pmol/L. Moreover, satisfied results were obtained when the assay was used to recognize the mismatched sequences.展开更多
The hairpin probe using microwave resonance in plasma is applicable to high pressure 1.33 ×10^3-1.01×10^5 Pa）） as developed recently. In this work, an analytic model of the hairpin resonator probe surround...The hairpin probe using microwave resonance in plasma is applicable to high pressure 1.33 ×10^3-1.01×10^5 Pa）） as developed recently. In this work, an analytic model of the hairpin resonator probe surrounded by a thin dielectric layer and a sheath layer is proposed. The correction factor due to these surroundings is analytically found and confirmed by electromagnetic field finite difference time domain simulation, thus enabling the accurate measurement of electron density in a high-pressure non-equilibrium uniform discharge.展开更多
The double-coupled microwave resonance probe(DMRP)based on the hairpin probe is proposed for diagnosing atmospheric plasma jet(ne<1017 m-3).In this work,the resonance characteristics of DMRP are investigated by num...The double-coupled microwave resonance probe(DMRP)based on the hairpin probe is proposed for diagnosing atmospheric plasma jet(ne<1017 m-3).In this work,the resonance characteristics of DMRP are investigated by numerical simulation.It shows that two resonance peaks on the reflectance spectrum can be observed,and influenced significantly by some parameters,such as the probe separation,the distance to the handheld radio frequency atmospheric pressure glow discharge plasma jet(RF-APGDPJ)and the plasma electron density less than 1017 m-3.Based on two resonance modes of DMRP,the electron densities in the afterglow of RF-APGDPJ at the different rf powers and helium flow rates are diagnosed experimentally by matching the change of FWHM(Df1-Df1,airand Df2-Df2,air)measured by vector network analyzer with the simulated relation between the FWHM changes and the plasma density.展开更多
设计合成了一种长臂发夹型核酸探针,结合核酸外切酶Ⅲ水解反应建立了一种免标记荧光信号放大高灵敏检测DNA的新方法。当不存在靶DNA时, SYBR Green Ⅰ荧光染料能够嵌入发夹型探针的茎部而发出很强的荧光,而当存在靶DNA并与发夹型探针杂...设计合成了一种长臂发夹型核酸探针,结合核酸外切酶Ⅲ水解反应建立了一种免标记荧光信号放大高灵敏检测DNA的新方法。当不存在靶DNA时, SYBR Green Ⅰ荧光染料能够嵌入发夹型探针的茎部而发出很强的荧光,而当存在靶DNA并与发夹型探针杂交后,核酸外切酶Ⅲ从杂交产物的3’端开始水解发夹型探针,释放出靶DNA,并触发下一个酶水解反应,同时SYBR Green Ⅰ染料也随发夹型探针水解而释放,导致荧光信号降低,从而实现了对DNA的免标记荧光信号放大高灵敏检测。该方法的检出限低至320 fmol/L,比传统双标的分子信标的方法降低了4~5个数量级,且该方法还具有免标记、简单、快速的特点。展开更多
This work investigates internal plasma process parameters using a hairpin resonance probe and optical emission spectroscopy. The dependence of electron density and atomic fluorine on the percentage of oxygen in an SF6...This work investigates internal plasma process parameters using a hairpin resonance probe and optical emission spectroscopy. The dependence of electron density and atomic fluorine on the percentage of oxygen in an SF6/O2 discharge was measured using these methods. An RIE Oxford Instruments 80 plus chamber was used for the experiments. Two different process powers （100 W and 300 W） at a constant pressure （100 mTorr） were used, and it was found that the optical emission intensity of the 703.7 nm and 685.6 nm lines of atomic fluorine increased rapidly as oxygen was added to the SF6 discharge, reached their maximum at an O2 fraction of 20% and then decreased with further addition of oxygen. The plasma electron density was also strongly influenced by the addition of O2.展开更多
Changes of the electron dynamics in hydrogen （H2） radio-frequency （RF） inductively coupled plasmas are investigated using a hairpin probe and an intensified charged coupled device （ICCD）. The electron density, p...Changes of the electron dynamics in hydrogen （H2） radio-frequency （RF） inductively coupled plasmas are investigated using a hairpin probe and an intensified charged coupled device （ICCD）. The electron density, plasma emission intensity, and input current （voltage） are measured during the E to H mode transitions at different pressures. It is found that the electron density, plasma emission intensity, and input current jump up discontinuously, and the input voltage jumps down at the E to H mode transition points. And the threshold power of the E to H mode transition decreases with the increase of the pressure. Moreover, space and phase resolved optical emission spectroscopic measurements reveal that, in the E mode, the RF dynamics is characterized by one dominant excitation per RF cycle, while in the H mode, there are two excitation maxima within one cycle.展开更多
基金Supported by the National Natural Science Foundation of China（No.21275043） and the National Basic Research Program of China（No .2009CB421601）.
文摘Detection of miRNAs presents a particular challenge because of their limited size, high sequence homo- logy and greatly various expression level. In this work, an ultrasensitive, label-free and isothermal miRNA detection was developed based on asymmertric hairpin probe, exonuclease I（Exo I） and SYBR Green I. The method employed asymmetric hairpin probe to perform cycled polymerization and Exo I to reduce background signal. In the presence of the target miRNA, the target triggers probe-mediated cycled polymerization reactions to generate lots of dsDNA products. The dsDNA product effectively prevents itself from being degraded by Exo I and permitted the insertion of more fluorescence dye into it to enlarge the fluorescence signal. In the absence of the target miRNA, there was no probe-mediated polymerization reaction, and the probe was digested by Exo I added, which minimized the intercala- tion of fluorescence dye to reduce the background signal. The combination of the probe-mediated cycled polymeriza- tion with the Exo 1-assisted background reduction allows us to achieve a detection limit of 5× 10^-18 mol/L. Owing to its ultrasensitivity, excellent specificity, convenience and low-cost, this method might hold out great promise in miRNA detection.
基金Supported by the National Natural Science Foundation of China（No.21375153）, the Fundamental Research Funds for the Central Universities of China（No. 131gzd05） and the Open Project of Beijing National Laboratory for Molecular Sciences, China.
文摘Simian virus 40（SV40） is a polyomavirus and can induce a series of different tumors. The recognition of SV40 genome is crucial to tumor diagnosis and gene therapy. Herein, a sensitive and selective colorimetric method for sequence-specific recognition of homopyrimidine-homopurine duplex DNA（dsDNA） of SV40（4424-4440, gp6） was established with a hairpin probe based upon the formation of triplex DNA. Hairpin probe 5＇-CCC TAC CCA TTT TTT CTT CTC TTT CCT GGG TAG GGC GGG TTG GG-3＇（HP） containing G-rich sequence and 17-bp triplex-forming sequence was used as the signal probe, which was stem-loop structure alone and exhibited low catalytic activity. Upon its binding to the target duplex of SV40, hairpin probe transferred from stem-loop structttre to parallel triplex DNA, accompanied by the recovery of catalytic activity of DNAzyme and a sharp increase of absorbance. Under optimum conditions, the absorbance was increased proportionally to the concentration of dsDNA over the range from 500 pmol/L to 40.0 nmol/L with a detection limit of 433 pmol/L. Moreover, satisfied results were obtained when the assay was used to recognize the mismatched sequences.
基金supported by the National Natural Science Foundation of China (Grant No. 10835004 )the 2nd Knowledge Cluster Research Project of Japan:Tokai Region Nanotechnology Manufacturing Cluster (Innovation of Environment Friendly Highly Functional Materials and Devices)
文摘The hairpin probe using microwave resonance in plasma is applicable to high pressure 1.33 ×10^3-1.01×10^5 Pa）） as developed recently. In this work, an analytic model of the hairpin resonator probe surrounded by a thin dielectric layer and a sheath layer is proposed. The correction factor due to these surroundings is analytically found and confirmed by electromagnetic field finite difference time domain simulation, thus enabling the accurate measurement of electron density in a high-pressure non-equilibrium uniform discharge.
基金supported by National Natural Science Foundation of China(No.11075033)。
文摘The double-coupled microwave resonance probe(DMRP)based on the hairpin probe is proposed for diagnosing atmospheric plasma jet(ne<1017 m-3).In this work,the resonance characteristics of DMRP are investigated by numerical simulation.It shows that two resonance peaks on the reflectance spectrum can be observed,and influenced significantly by some parameters,such as the probe separation,the distance to the handheld radio frequency atmospheric pressure glow discharge plasma jet(RF-APGDPJ)and the plasma electron density less than 1017 m-3.Based on two resonance modes of DMRP,the electron densities in the afterglow of RF-APGDPJ at the different rf powers and helium flow rates are diagnosed experimentally by matching the change of FWHM(Df1-Df1,airand Df2-Df2,air)measured by vector network analyzer with the simulated relation between the FWHM changes and the plasma density.
文摘设计合成了一种长臂发夹型核酸探针,结合核酸外切酶Ⅲ水解反应建立了一种免标记荧光信号放大高灵敏检测DNA的新方法。当不存在靶DNA时, SYBR Green Ⅰ荧光染料能够嵌入发夹型探针的茎部而发出很强的荧光,而当存在靶DNA并与发夹型探针杂交后,核酸外切酶Ⅲ从杂交产物的3’端开始水解发夹型探针,释放出靶DNA,并触发下一个酶水解反应,同时SYBR Green Ⅰ染料也随发夹型探针水解而释放,导致荧光信号降低,从而实现了对DNA的免标记荧光信号放大高灵敏检测。该方法的检出限低至320 fmol/L,比传统双标的分子信标的方法降低了4~5个数量级,且该方法还具有免标记、简单、快速的特点。
基金supported by the EC Framework 7 IMPROVE research project (IR -2008-0013)the Science Foundation Ireland PRECISION project (08-SRC-I1411)
文摘This work investigates internal plasma process parameters using a hairpin resonance probe and optical emission spectroscopy. The dependence of electron density and atomic fluorine on the percentage of oxygen in an SF6/O2 discharge was measured using these methods. An RIE Oxford Instruments 80 plus chamber was used for the experiments. Two different process powers （100 W and 300 W） at a constant pressure （100 mTorr） were used, and it was found that the optical emission intensity of the 703.7 nm and 685.6 nm lines of atomic fluorine increased rapidly as oxygen was added to the SF6 discharge, reached their maximum at an O2 fraction of 20% and then decreased with further addition of oxygen. The plasma electron density was also strongly influenced by the addition of O2.
基金Project supported by the National Natural Science Foundation of China(Grant Nos.11075029,11175034,and 11205025)the Fundamental Research Funds for Central Universities,China(Grant No.DUT12RC(3)14)
文摘Changes of the electron dynamics in hydrogen （H2） radio-frequency （RF） inductively coupled plasmas are investigated using a hairpin probe and an intensified charged coupled device （ICCD）. The electron density, plasma emission intensity, and input current （voltage） are measured during the E to H mode transitions at different pressures. It is found that the electron density, plasma emission intensity, and input current jump up discontinuously, and the input voltage jumps down at the E to H mode transition points. And the threshold power of the E to H mode transition decreases with the increase of the pressure. Moreover, space and phase resolved optical emission spectroscopic measurements reveal that, in the E mode, the RF dynamics is characterized by one dominant excitation per RF cycle, while in the H mode, there are two excitation maxima within one cycle.
