The Research On Some Chemical And Physical Test Methods In The Chemical And Fiber Production Process

The construction of efficient,sensitive and stable biosensors and nanomedicines is very important to biosensing,disease diagnosis and therapy.Nucleic acids can not only store and transmit genetic information,but also has excellent recognition and self-assembly functions,so it can be used to design various types of biosensors(such as fluorescent probes,electrochemical probes,etc.)and nanomedicines.However,the traditional nucleic acid probes are mostly linear probes,which are easily degraded by nucleases,resulting in poor stability and low cellular uptake efficiency,resulting in compromised performance in complex physiological environments is greatly compromised.In addition,the development of nucleic acid drugs is limited by low cellular uptake efficiency and poor intracellular stability,resulting in low drug delivery efficiency.The development of framework nucleic acids(FNAs)is expected to solve the abovementioned problems.FNAs refers to a three-dimensional shell or outer frame skeleton composed of DNA.Compared with traditional linear nucleic acids,FNAs are internalized into cells via caveolae mediated endocytosis,and have good resistance to nuclease cleavage,and maintain sensing or therapeutic performance in complex physiological environments.Therefore,this research makes full use of the advantages of the FNAs to construct more stable,efficient,and highly biocompatible fluorescent biosensor.At the same time,the introduction of photoacoustic dye to improve the penetration depth and imaging performance in biological tissues,and constructs CRISPR-Cas9 carrier based on FNAs to realize gene silencing in cells.The specific research content includes the following aspects:(1)In chapter 2,in order to solve the problem that traditional linear nucleic acid probes are susceptible to nuclease degradation,we designed a self-assembly DNA nanocage structure,and encapsulated functional nucleic acid probes(aptamer probes targeting ATP)in the internal cavity of the DNA nanocage structure to construct a functional nucleic acid probe loaded in the DNA nanocage(Cage-probe).A series of experiments have proved that,compared with traditional linear aptamer probes,Cage-probe has excellent resistance to nuclease cleavage and can be taken up by cells via caveolae mediated endocytosis without the transfection process.The strategy of loading functional nucleic acid probes with DNA nanocages constructed in this chapter provides a new idea for constructing biosensors with high stability and high biocompatibility.(2)In chapter 3,in order to overcome light scattering in biological tissues,the photoacoustic nucleic acid probe is encapsulated in the self-assembly DNA nanocage structure construct a photoacoustic nucleic acid probe loaded in the DNA nanocage(Cage-PA-probe).Compared with traditional fluorescent biosensors,Cage-PA-probe exhibited stronger tissue penetration depth and successfully achieved the imaging of targets in vivo(taking mi RNA-21 as an example).The Cage-PA-probe provide a new method for constructing highly stable in vivo imaging probes.(3)In chapter 4,in order to achieve cell delivery of the CRISPR-Cas9 system,a strategy of using DNA tetrahedron to carry the CRISPR-Cas9 system for intracellular gene silencing was developed.In this work,firstly through the construction of plasmids,high-purity Cas9 protein with normal cleavage activity was obtained.Then,confocal laser microscopy and flow cytometry were used to successfully prove the feasibility of the strategy of using DNA tetrahedrons to carry the CRISPR-Cas9 system.The delivery strategy which based on DNA tetrahedron proposed in this chapter is expected to provide new ideas for the application of the CRISPR-Cas9 system in the field of gene therapy.