The Construction Of DNA Tweezers Mediated Proximity Analysis Switch And Its Sensing Application

DNA is a carrier of biological genetic information,DNA nanomaterials have excellent controllability,programmability and predictability due to the strict base complementary pairing principles,and are widely used in the construction of biosensors.The immunopolymerase chain reaction transformed the immunoassay of protein into the detection of DNA.With the aid of the amplification ability of DNA nanomaterials,the sensitivity of protein detection was greatly improved,but it required complicated fixation and tedious washing process.In order to solve this problem,Fredrisson et al.first proposed the DNA-mediated proximity assay,which avoids washing or separation steps,and enabling the homogeneous and sensitive detection of protein.This strategy brings two or more DNA sequences into proximity via the affinity interactions identify and binding target protein to produce the detectable DNA sequences.However,because the target recognition method is a dual recognition mode,there are certain limitations,such as poor versatility,low detection accuracy and sensitivity,and irreversible proximity binding reactions,effective regulation of neighboring binding cannot be achieved.As a kind of DNA molecular machines,DNA tweezers have two conformations of "open" and "close".Under the stimulation of specific nucleic acid molecules,p H changes,small molecules,etc.,they can dynamically switch between the two conformations,and then realize the capture,hold and release of specific stimuli.Through effective regulation of DNA tweezer,it is possible to construct a universal proximity analysis switch.There are four parts in this paper:Chapter one is the introduction section,and it mainly introduces DNA nanomaterials,DNA-mediated proximity analysis,the basic situation of DNA tweezers.In addition,its contains the scientific problems and main research content.In chapter two,a proximity analysis switch with DNA tweezers as a scaffold was constructed.The toehold domain(TD)and branch migration domain(BMD)were respectively designed on ends of the DNA tweezer arms,and the dynamic conversion of the tweezers was used to achieve effective regulation of proximity analysis.Combining it with hybridization chain reaction(HCR),the ultrasensitive detection of miRNA was achieved,and the detection limit was 9.4 f M.By changing the aptamer sequence,the sensitivity and specificity of thrombin was also achieved,which improves the versatility of proximity analysis.Based on the conformational transformation of DNA tweezers,Boolean logic platform was constructed to realize the logical operation of the system.This strategy has great application potential in the field of biological detection and biomedical research.In chapter three,a DNA tweezers-actuated regenerated DNA nanoreactor was constructed.The reactor used DNA tweezers as a scaffold,which can be reversibly controlled during target detection,and a regenerated DNAzyme was achievedfor the first time.This method realized the sensitive and selective detection of miRNA,and the detection limit was1.23 p M.In addition,the sensitive and selective detection of small molecule adenosine was also achieved,which can be used to distinguish the content of adenosine in urine samples of normal people and cancer patients,indicating that this strategy can be used for sensitive analysis of multiple targets,and had good flexibility and versatility,provides a repeatable and reproducible platform for the sensitive and selective detection of cancer-related biomolecules in clinical samples.Chapter four is a conclusion and expectation section,which summarizes and prospects the research results of the paper.