We described a new strategy to combine hybridization chain reaction(HCR) process and triplex DNA through Ag+/cysteine,thus to pursue the controllable process of HCR. H1, a specially-designed triplex DNA, with homopyri...We described a new strategy to combine hybridization chain reaction(HCR) process and triplex DNA through Ag+/cysteine,thus to pursue the controllable process of HCR. H1, a specially-designed triplex DNA, with homopyrimidines which can bind to dsDNA through Hoogsteen bonding, forming a cap-like triplex DNA. The presence of Ag^+ can play the role as a H1 locker to stop initiator from triggering HCR process, forming Ag+-stabilized triplex DNA molecular switch(denoted as Ag^+-STDMS). However,the strong binding ability of cysteine towards Ag^+ forming stable Ag^+-cysteine can release the locking tail of H1 and realize the retriggering of HCR. This study presented a promising tool to regulate the self-assembly processes of DNA-based nanostructures in neutral environment. Under the optimum conditions, fluorescence intensity(peaking at 582 nm) of HCR is proportional to the concentration of Ag+ in the 0.2–4.0 μM range. Within the presence of Ag^+, the fluorescence intensity is also proportional to the concentration of cysteine in the 0.2–3.0 μM range. The method can successfully manipulate the assembly and disassembly of DNA in HCR.展开更多
THE observation of the formation of triplex DNA opened a possibility to control gene expression. Since then vast applications on biology and novel approaches to therapeutics have been accumulated considerably. Novel a...THE observation of the formation of triplex DNA opened a possibility to control gene expression. Since then vast applications on biology and novel approaches to therapeutics have been accumulated considerably. Novel approaches that have been developed in the past decades have the capability to supplement or to replace the classical drug paradigm of small molecules to the development of pharmaceuticals. The distinct field of oligonucleotide therapeutic can be possible when an additive oligonucleotide is targeted to a specified double helical DNA. The use of complementary DNA sequences to arrest translation of mRNAs also opens new possibilities for an artificial control of gene expression. The interest in the stability of the formation of a triplex DNA has been driven largely by the ability to synthesize oligonucleotides.展开更多
Conformation stabilities of intramolecular triple-helical DNAs (intra-tnplex DNAs: 5-d(TC)6-d(T)m-d(CT)6-d(C)n-d(AG)6-3’,where m and n are chosen to be 4 and 3) have been examined by molecular mechanics.The four intr...Conformation stabilities of intramolecular triple-helical DNAs (intra-tnplex DNAs: 5-d(TC)6-d(T)m-d(CT)6-d(C)n-d(AG)6-3’,where m and n are chosen to be 4 and 3) have been examined by molecular mechanics.The four intra-triplexes (H-DNAs) are compared with the mtermolecular triplexes d(TC)6 * d ( AG)6 d(CT)6.The results revealed that loops do not significantly influence the mtratriplex conformations,loop conformations,however,depend partly on its length.Loop also makes strand Ⅱ of every intra-triplex DNA longer than that of the inter-triplex DNA.Most of residue sugar conformations of triple-helical DNAs are S-type,there aiso exist,however,N-type conformation and the conformations between S-type and N-type Possible models of the five triplex DNAs are presented.展开更多
The preparations,the formation conditions and stabilities of duplex dA<sub>10</sub>·dT<sub>10</sub>and triplaxdA<sub>10</sub>·2dT<sub>10</sub>,and the interact...The preparations,the formation conditions and stabilities of duplex dA<sub>10</sub>·dT<sub>10</sub>and triplaxdA<sub>10</sub>·2dT<sub>10</sub>,and the interactions of ethidium bromide with them in an appropriate buffer are reported.Flu-orescence spectra show that ethidium can be used as a useful fluorescence probe to detect triplex formation,and its fluorescence is significantly increased by either the duplex or triplex,but less in the case of thetriplex.Thermal denaturation profiles demonstrate that the stability of the triplex is enhanced by ethidium.Fluorescence energy transfer studies suggest the existence of similar energy transfer from the triplex or du-plex to the bound ethidium but the presence of the triplex results in substantially smaller energy transfer thanthat of the duplex does.Furthermore,fluorescence quenching using the anionic quencher[Fe（CN）<sub>6</sub>]<sup>4-</sup>can-not decrease the fluorescence intensities of triplex/ethidium complex.These results demonstrate that ethidi-um has significantly binding affinity with the triplex,and interacts with it via intercalative mechanism,thusincreases its stability.展开更多
Nucleic acid amplification test is a reliable method for primary human immunodeficiency virus(HIV) infection diagnosis.Herein, a novel fluorescent method for sequence-specific recognition of DNA fragment of HIV-1 was ...Nucleic acid amplification test is a reliable method for primary human immunodeficiency virus(HIV) infection diagnosis.Herein, a novel fluorescent method for sequence-specific recognition of DNA fragment of HIV-1 was established based upon nicking-assisted strand displacement amplification(SDA) and triplex DNA. In the presence of target dsDNA, nicking-assisted SDA process generated a lot of ssDNA, which hybridized with molecular beacon to produce signal. The fluorescence intensity was proportional to the concentration of target dsDNA within the range from 5 to 1000 pmol/L, with a detection limit of 1.4 pmol/L. Moreover, it successfully distinguished target dsDNA from the nucleic acid extractive of human blood. Thus this method has the merit of high sensitivity, and it is suitable for sequence-specific recognition of target dsDNA in complex matrices, which made it a potential application in diagnosis of acquired immunodeficiency syndrome(AIDS) in the future.展开更多
Strand displacement reaction is a crucial component in the assembly of diverse DNA-based nanodevices,with the toehold-mediated strand displacement reaction representing the prevailing strategy.However,the single-stran...Strand displacement reaction is a crucial component in the assembly of diverse DNA-based nanodevices,with the toehold-mediated strand displacement reaction representing the prevailing strategy.However,the single-stranded Watson-Crick sticky region that serves as the trigger for strand displacement can also cause leakage reactions by introducing crosstalk in complex DNA circuits.Here,we proposed the toeless and reversible DNA strand displacement reaction based on the Hoogsteen-bond triplex,which is compatible with most of the existing DNA circuits.We demonstrated that our proposed reaction can occur at pH 5 and can be reversed at pH 9.We also observed an approximately linear relationship between the degree of reaction and pH within the range of pH 5-6,providing the potential for precise regulation of the reaction.Meanwhile,by altering the sequence orientation,we have demonstrated that our proposed reaction can be initiated or regulated through the same toeless mechanism without the requirement for protonation in low pH conditions.Based on the proposed reaction principle,we further constructed a variety of DNA nanodevices,including two types of DNA logic gates that rely on pH 5/pH 9 changes for initiating and reversing:the AND gate and the OR gate.We also successfully constructed a DNA Walker based on our proposed reaction modes,which can move along a given track after the introduction of a programmable DNA sequence and complete a cycle after 4 steps.Our findings suggest that this innovative approach will have broad utility in the development of DNA circuits,molecular sensors,and other complex biological systems.展开更多
基金supported by the National Natural Science Foundation of China (21375153)the Fundamental Research Funds for the Central Universities (171gpy79)
文摘We described a new strategy to combine hybridization chain reaction(HCR) process and triplex DNA through Ag+/cysteine,thus to pursue the controllable process of HCR. H1, a specially-designed triplex DNA, with homopyrimidines which can bind to dsDNA through Hoogsteen bonding, forming a cap-like triplex DNA. The presence of Ag^+ can play the role as a H1 locker to stop initiator from triggering HCR process, forming Ag+-stabilized triplex DNA molecular switch(denoted as Ag^+-STDMS). However,the strong binding ability of cysteine towards Ag^+ forming stable Ag^+-cysteine can release the locking tail of H1 and realize the retriggering of HCR. This study presented a promising tool to regulate the self-assembly processes of DNA-based nanostructures in neutral environment. Under the optimum conditions, fluorescence intensity(peaking at 582 nm) of HCR is proportional to the concentration of Ag+ in the 0.2–4.0 μM range. Within the presence of Ag^+, the fluorescence intensity is also proportional to the concentration of cysteine in the 0.2–3.0 μM range. The method can successfully manipulate the assembly and disassembly of DNA in HCR.
文摘THE observation of the formation of triplex DNA opened a possibility to control gene expression. Since then vast applications on biology and novel approaches to therapeutics have been accumulated considerably. Novel approaches that have been developed in the past decades have the capability to supplement or to replace the classical drug paradigm of small molecules to the development of pharmaceuticals. The distinct field of oligonucleotide therapeutic can be possible when an additive oligonucleotide is targeted to a specified double helical DNA. The use of complementary DNA sequences to arrest translation of mRNAs also opens new possibilities for an artificial control of gene expression. The interest in the stability of the formation of a triplex DNA has been driven largely by the ability to synthesize oligonucleotides.
基金Project supported by the Chinese Academy of Sciences
文摘Conformation stabilities of intramolecular triple-helical DNAs (intra-tnplex DNAs: 5-d(TC)6-d(T)m-d(CT)6-d(C)n-d(AG)6-3’,where m and n are chosen to be 4 and 3) have been examined by molecular mechanics.The four intra-triplexes (H-DNAs) are compared with the mtermolecular triplexes d(TC)6 * d ( AG)6 d(CT)6.The results revealed that loops do not significantly influence the mtratriplex conformations,loop conformations,however,depend partly on its length.Loop also makes strand Ⅱ of every intra-triplex DNA longer than that of the inter-triplex DNA.Most of residue sugar conformations of triple-helical DNAs are S-type,there aiso exist,however,N-type conformation and the conformations between S-type and N-type Possible models of the five triplex DNAs are presented.
基金the Eighth Five-Year Plan's Key Basic Research Project of Academia Sinica
文摘The preparations,the formation conditions and stabilities of duplex dA<sub>10</sub>·dT<sub>10</sub>and triplaxdA<sub>10</sub>·2dT<sub>10</sub>,and the interactions of ethidium bromide with them in an appropriate buffer are reported.Flu-orescence spectra show that ethidium can be used as a useful fluorescence probe to detect triplex formation,and its fluorescence is significantly increased by either the duplex or triplex,but less in the case of thetriplex.Thermal denaturation profiles demonstrate that the stability of the triplex is enhanced by ethidium.Fluorescence energy transfer studies suggest the existence of similar energy transfer from the triplex or du-plex to the bound ethidium but the presence of the triplex results in substantially smaller energy transfer thanthat of the duplex does.Furthermore,fluorescence quenching using the anionic quencher[Fe（CN）<sub>6</sub>]<sup>4-</sup>can-not decrease the fluorescence intensities of triplex/ethidium complex.These results demonstrate that ethidi-um has significantly binding affinity with the triplex,and interacts with it via intercalative mechanism,thusincreases its stability.
基金supported by the National Natural Science Foundation of China(21375153)
文摘Nucleic acid amplification test is a reliable method for primary human immunodeficiency virus(HIV) infection diagnosis.Herein, a novel fluorescent method for sequence-specific recognition of DNA fragment of HIV-1 was established based upon nicking-assisted strand displacement amplification(SDA) and triplex DNA. In the presence of target dsDNA, nicking-assisted SDA process generated a lot of ssDNA, which hybridized with molecular beacon to produce signal. The fluorescence intensity was proportional to the concentration of target dsDNA within the range from 5 to 1000 pmol/L, with a detection limit of 1.4 pmol/L. Moreover, it successfully distinguished target dsDNA from the nucleic acid extractive of human blood. Thus this method has the merit of high sensitivity, and it is suitable for sequence-specific recognition of target dsDNA in complex matrices, which made it a potential application in diagnosis of acquired immunodeficiency syndrome(AIDS) in the future.
基金financially supported by the National Key Research and Development Program of China(No.2021YFC2701402)the Open Research Fund of State Key Laboratory of Bioelectronics,Southeast University(No.Sklb2021-k06)+1 种基金the Open Foundation of NHC Key Laboratory of Birth Defect for Research and Prevention(Hunan Provincial Maternal and Child Health Care Hospital)(No.KF2020007)the Open Foundation of Translational Medicine National Science and Technology Infrastructure(Shanghai)(No.TMSK-2021-141)。
文摘Strand displacement reaction is a crucial component in the assembly of diverse DNA-based nanodevices,with the toehold-mediated strand displacement reaction representing the prevailing strategy.However,the single-stranded Watson-Crick sticky region that serves as the trigger for strand displacement can also cause leakage reactions by introducing crosstalk in complex DNA circuits.Here,we proposed the toeless and reversible DNA strand displacement reaction based on the Hoogsteen-bond triplex,which is compatible with most of the existing DNA circuits.We demonstrated that our proposed reaction can occur at pH 5 and can be reversed at pH 9.We also observed an approximately linear relationship between the degree of reaction and pH within the range of pH 5-6,providing the potential for precise regulation of the reaction.Meanwhile,by altering the sequence orientation,we have demonstrated that our proposed reaction can be initiated or regulated through the same toeless mechanism without the requirement for protonation in low pH conditions.Based on the proposed reaction principle,we further constructed a variety of DNA nanodevices,including two types of DNA logic gates that rely on pH 5/pH 9 changes for initiating and reversing:the AND gate and the OR gate.We also successfully constructed a DNA Walker based on our proposed reaction modes,which can move along a given track after the introduction of a programmable DNA sequence and complete a cycle after 4 steps.Our findings suggest that this innovative approach will have broad utility in the development of DNA circuits,molecular sensors,and other complex biological systems.
