| It is well known that DNA is the chromosomes of the main chemical composition and it is the undertaker genetic information. The researchers found that specific sequence for nucleotide bases decides biology gene genetic characteristics and the protein synthesis templates oforganism, and the specific sequence for nucleotide bases is biodiversity in nature and maintain life all sorts of function normal operation of the basic structure unit. The reasons for a lots of human genetic diseases are related to the variation of nucleotide molecular base sequences, such as Patient fibrosis, Benzene acetone urine disease, Sickle cell anemia, hemophilia, tumor and so on. So it is necessary to develop rapid and sensitive methods for the detection of specific DNA sequences.Eletrogenerated chemiluminescence is a technique that combines chemiluminescence and electrochemistry. The chemiluminesence reaction is initiated by an electrochemical reaction atelectrode surface. Such an elelctro-initiation reaction introduces a large number of additional advantagges, such as high sensitivity and selectivity, rapid and convenient operation and relatively simple instrumentation system. Due to these inherent advantages, eletrogenerated chemiluminescence method has attracted much attention from all analytical fields, especially from biochemical analysis. The aim of the present work is to develop a series of methods highly sensitive, simple and rapid for the DNA hybridization detection basing on electrogenerated chemiluminescence. Combining the technology of DNA hybridization and immobilization techniques, we have developed a series of eletrogenerated chemiluminescence methods for the determination of sepecial DNA sequence. In this work, we have utilized the influence on eletrogenerated chemiluminescence signal which were initiated to the difference interaction frombetween single strand DNA or duplex DNA morphology and porous structure of modified electrode to develop two kinds of methods simple, lable free, rapid, as well as high selective. This thesis includes review and research section. In the review, the basic pinciples, characteistics and systems of ECL, the principle of DNA detection are introduced. The principles and characteristics of various kinds of DNA detection are summarized. Finally the purpose and content of the thesis are presented.The research section contains two subunits.Subunit1, the study of DNA ECL biosensor basing polyaniline porous structure modified electrode.ECL detection methods with high sensitivity had been used in DNA sequence recognition for livingmatter combining of fixed DNA technology, DNA hybridization. Polyaniline porous structure was electrodynamically deposited onto the graphite electode. Oligonucleotides with amino-group at the5' end were covalently linked onto amidogroups of polyaniline porous structure in the presence of the soluble coupling reagent glutaraldehyde. Label-free oligonucleotide ECL sensor that used a change in the interaction between DNA and the surface of graphite electrode modified polyaniline to signal the formation of a DNA duplex with luminol as detection probe was investigated, which was simple, rapid, and could detect single-base mismatch DNA sequence. Under the optimized test conditions, the detection limit of ECL sensors was1.1×10-10mol/L to the complementary target DNA, and it furthered expand the application of the modified electrode in the field of biological analysis.Subunit2, the study of DNA ECL biosensor basing Sol-Gel Silica-Chitosan-Ru(bpy)32+composite silicon membrane porous structure modified electrodeIn this work, we firstly modified Silica-Chitosan-Ru(bpy)32+composite silicon membrane rough and poriferous on the surface of graphite electrode making use of classical electro-deposition Sol-Gel, and then immobilized linear DNA label-free with electrostatic adsorption. Label-free oligonucleotide ECL sensor that used a change in the interaction between DNA and the surface of graphite electrode modified Silica-Chitosan composite films to signal the formation of a DNA duplex with Ru(bpy)32+as detection probe was investigated. The results showed that it was able to recognize complementay sequence to form dsDNA. The linear range for complementary sequence was1.0×10-11～l.0×10-9mol/L. The detection limit was2.7×10-12mol/L for complementary target DNA, which was much lower than of subunit1. The Sol-Gel films porous provided a novel means for the ssDNA immobilization and sequence-specific DNA detection. |
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