Dna Condensed Matter And Cell Patterning In Gene Transfection In Basic Research,

With the development of gene therapy and tissue engineering, effective gene transfection has become a hot topic. There are mainly two steps in gene transfection: preparing the complexes of DNA/gene delivery vectors and gene expression in mammalian cells. Preparation of the complexes of DNA/gene delivery vectors referes to the interaction between DNA and gene delivery vectors, which will cause great changes into the structure of DNA and is referred to as DNA condensation. Gene expression in mammalian cells will be influenced by the cell status. Consequently, gene transfection studies on patterned cells are of great meaning for their gene therapeutic applications. In this study, we investigated the influence of DNA condensation and cell patterning on gene transfection for the purpose of improving gene transfection efficiency, as follows:Fisrt, we investigated the influence of DNA condensation on gene transfection efficiency. In the first part, we investigated how the structure of DNA influenced the gene transfection efficiency. We treated the plasmid DNA with high temperature or alkali and investigated their DNA condensation efficiency as well as their gene transfection efficiency. We found that the heat or alkali treatment greatly changed the structure of DNA. The change significantly increased their DNA condensation efficiency. Further research showed that heat denatured DNA had higher gene transfection efficiency than the native DNA. Besides, the alkali treated DNA were shown to be more sensitive to PEI and could perform gene transfection experiment in the presence of less PEI. In the second part, we investigated the influence of gene delivery vectors on gene transfection efficiency. We synthesized a supermolecular chemicals containing Ru, PPRs with CB[6], PPRs of similar structures but different lengths and a few-positively-charged polymer poly(PEGMA)-4N. We investigated their DNA condensation efficiency and gene transfection efficiency. We also explained the mechanism about DNA condensation. We found that the positive charges, the damage to DNA and the cell toxicity had an influence on the gene transfection efficiency. We found that CB[6] and the length of PPR could effectively control the interaction between DNA and PPR. We detailed the mechanism of DNA condensation in electric field, pointing out that the destruction of DNA/PPRs complexes in electric field was related to the stability between DNA and PPR. We detailed the mechanism of DNA condensation induced by cationic polymers, pointing out that the DNA condensation was caused by both static attraction and depletion effect, the latter of which played a more important part in polymers with few positive charges.Second, we investigated the gene transfection on the patterned cells. We developed new methods to pattern cells on both hydrophilic mica surface and hydrophobic PDMS surface and investigated the gene transfection efficiency on the patterned cells. Data showed that by micropatterning collagen, cell patterns could be obtained on hydrophilic mica surface. The patterned cells were of good gene transfection efficiency. Data also showed that self-assembly of hydrophobin HFBI could effectively convert the hydrophobic PDMS surface into a hydrophilic one, which should facilitate protein immobilization and cell adhesion. Due to the amphiphilic property of HFBI, cell patterns could be obtained on PDMS surface. Data also showed that these cells were of good gene treasfection efficiency.This research investigated the influence of DNA condensation and cell pattern on gene transfection efficiency. The data not only contained new methods for improving transfection efficiency, new methods to pattern cells and new methods for controlling DNA condensation, but also explained the mechanism about DNA condensation. Our research will contribute to gene therapy and tissue engineering fields.