Study On Strand Displacement Kinetics Of Nucleic Acid Bulge Structure

Deoxyribonucleic acid(DNA)is a kind of gene carrier that stores and transmits genetic information in organisms.It plays an important role in the growth and genetic regulating process of organisms.Based on sequence specificity,unique programmability and diversity,DNA has been explored for the rational design of many nanoscale structures and functions in the field of DNA nanotechnology.It exhibits outstanding potential for applications in biosensing,logic operations,disease diagnosis and therapy.The rational design of these nanoscale structures and functions is based on nucleic acid hybridization and strand displacement.In this paper,we focus on the kinetics of nucleic acid hybridization and the effects of bulge structure on rates of hybridization and strand displacement.The detailed works are as follows:1.First,the hybridization kinetics of double strands with and without the bulge structure were preliminarily investigated.The effects of temperatures,salt concentrations on the kinetics of duplex formation with bulge structure and the formation of fully matched duplex were explored;and different ratios were examined for single stranded substrates.The trend of reaction rates under different conditions and the effect of different bulge lengths on hybridization rate were obtained by comparing and analyzing the kinetic curves.2.Based on kinetic results of double strand hybridization,the effect of bulge structure on the kinetics of double strand displacement reaction was explored.First,the optimal ratio of the two single strands to form a duplex and reaction extents were initially determined by gel electrophoresis.Then,the fluorescence kinetics characterization was performed to study the effects of temperatures and salt concentrations.The influence of bulge structure with the same length at different positions on strand displacement rates was also explored.3.Inspired by the results of strand displacement reaction for double strand DNA with bulge,similar bulge regions were applied to the Y-shaped structure formed by three strands.Firstly,gel electrophoresis was used to find out the optimal reaction ratio.Secondly,we explored the displacement rates of Y-structure with different lengths of bulges,and the effect of equal length of bulges at different positions of Y-structure.Then,the kinetics of reverse displacement of fully matched Y structure with bulge single strand were also studied.Finally,the effect of toehold length on reaction rates were also considered.In summary,based on double strand hybridization reaction and toehold mediated strand displacement reaction,the effects of bulge structure on strand displacement reaction were studied with gel electrophoresis and fluorescence spectrometry.This study provides a new idea for the construction of multifunctional DNA logic circuit or the design of complex nanostructures,and provides crucial information for DNA bulge related conformation,which is beneficial to disease diagnosis.