Magnetic Immobilization Of RNA And A Novel Model Of Its Drug Delivery System

Nowadays, people attach more and more attention to gene therapy for diseases, which mainly shows that oligonucleotides are taken as a clinical drug. In the early stage, people researched on antisense oligonucleotides but siRNAs-based genetic medicine has made great progress in recent years. The siRNAs can specifically and quickly treat many diseases, so it has great potency of application. However, how to quickly and effectively deliver these micromolecular substances to specific cells and tissues is crucial for clinical application. Targeting-delivery can enhance efficiency and specificity of oligonucleotide drugs, which can be effectively delivered to designated parts to work.In recent years, magnetic polymer microsphere is emerged as a kind of new-type functional material, which is widely used in many fields such as biomedicine, bioengineering and cytology. Chitosan has been widely used in medicine because of its good biocompatibility and effective biodegradation. Besides, catabolite has no side effect on our body and immunogenicity and precipitation in our body. Most important, chitosan has therapeutic effect on regulating blood fat, blood pressure, and blood sugar level and curing tumors. With its biocompatibility, stability, magnetism, rich active group on surface, chitosan, having no side effect, can be applied into these fields like targeted drug, immunoassays and cellular segregation as a carrier combined with multi-functional molecules.The paper will explore nanometer-scale magnetic microsphere immobilization siRNA and its application in new-type drug delivery system. The research can be divided into three parts:1) Adoption of coprecipitation preparation of nanometer-scale Fe3O4magnetofluid and chemical crosslinking. Dispersing magnetofluid into chitosan fluid and then add adequate glutaraldehyde as cross-linking agent to produce Fe3O4as kernel and chitosan’s nanometer-scale magnetic chitosan composite microsphere as outer sphere.2) Extraction of RNA of beer yeast by phenol method. Taking magnetic chitosan microsphere as a carrier and glutaraldehyde as a cross-linking agent, RNA is magneticly immobilized. By comparing the content of inorganic phosphorus and the total phosphorus in reaction system, immobilized condition of RNA is optimized. In the research of RNA immobilization, when magnetic chitosan microsphere as a carrier and glutaraldehyde as a cross-linking agent, the optimum condition of RNA immobilization is as follows:the optimum response time of magnetic immobilization of yeast RNA is4h, the optimum reaction temperature is40℃, the optimum reaction pH is5.5, the optimum additive amount of RNA is20mL (2mg/mL), and the optimum additive amount of glutaraldehyde is100μL (50%), the optimum rotational speed of oscillator is60r/min.3) Exploration of interference of immobilized siRNA by the experiment of animal tissue culture. Use the optimal condition of immobilized RNA to immobilize siRNA by cross-linking method. Rat liver is divided into two groups for tissue culture:group1, adding siRNA of magnetic immobilization; group2, control group, not adding siRNA of magnetic immobilization. Analyzing interference of siRNA by testing the enzymatic activity of glutamic-pyruvic transaminase. The results are enzymatic activity in group1takes up53%of fresh liver,68%in group2. Group1is significantly lower than in group2. The experimental result shows magnetic immobilized siRNA has good paramagnetism and interference effect on rat liver GPT gene expression.