Study And Application Of Cationic Comb-type Copolymer Assisted Rapid DNA Self-assembly At Constant Temperature

With the rapid development of DNA nanotechnology,well-defined DNA modules have been designed and manufactured.On this basis,various complex DNA three-dimensional systems can be constructed by hybridization or strand exchange reaction between the sticky ends of DNA modules.However,the assembly processes of DNA modules are restricted by many factors.For example,DNA hybridization and strand exchange reaction are limited by the dynamic hybridization,and the exchange reaction between DNA strands needs competitive replacement,which is completed in a step-by-step manner through slow multiple annealing,thus limiting the self-assembly of DNA assembly efficiency and thermodynamic stability.Increasing the concentration or length of the sticky ends can improve the rate of DNA hybridization and strand exchange reaction to a certain extent.However,it often leads to the formation of cross-linking or entanglement between DNA modules,thus affecting the morphology and structure of DNA assembly.In order to overcome the above shortcomings,the assembly of DNA modules needs new process to realize the accurate and fast process.A series of previous studies of our group have shown that cationic comb-type copolymers can accelerate DNA hybridization and strand exchange reaction at relatively low DNA concentration and less sticky ends,while improving the stability of DNA multilevel structure.From this point of view,we propose the concept of"cationic comb-type copolymers assisted rapid assembly of DNA nanostructures at constant temperature",which is the main content of this thesis.The detailed research work is introduced as follows:1.Cationic comb-type copolymer polylysine grafted dextran PLL-g-Dex was synthesized by reductive amination and its structure was characterized.In the presence of PLL-g-Dex,four single strands at low concentration can assemble rapidly at room temperature to form a double crossover module,and its yield and stability are significantly improved.Without PLL-g-Dex,it is difficult for low concentration DNA to assemble into double crossover structure.The results showed that the cationic comb-type copolymer can accelerate the precise assembly of DNA modules free of annealing treatment.Further,under the action of comb type cationic copolymer,the double crossover modules with sticky ends can quickly form high-ordered network structure.In addition,the successful adding of cationic comb-type copolymer and anionic polymer sodium polyethylene sulfonate PVS can achieve rapid and controllable conversion of double crossover network.2.A label-free fluorescent aptasensor based on G-quadruplex was designed to detect and extract vascular endothelial growth factor(VEGF).The sulfhydryl modified DNA was introduced into chitosan hydrogel by click chemistry reaction,and combined with the DNA chain containing VEGF aptamer and c-myc.In the presence of K~+,the c-myc sequence forms a G-quadruplex.PLL-g-Dex can rapidly assemble the G-chains in the solution into G-nanowires and immobilized on the chitosan hydrogel.The target VEGF can form a complex with aptamers,and induce G-nanowires to release from the hydrogel into the liquid phase.Then,the fluorescence intensity of Th T bound to G-nanowires was significantly enhanced,which can realize the high sensitivity detection of VEGF.When the VEGF aptamer was hybridized with its complementary chain,VEGF was rapidly dissociated from the aptamer to achieve accurate extraction.