Construction And Preliminary Application Of New DNA Molecular Logic Gates

Nucleic acid (DNA/RNA) has a precise molecular recognition capabilities and powerful signal storage capacity. The use of the biological characteristics of nucleic acid (DNA/RNA) molecules to build molecular-level logic gates and logic operations to achieve the emerging field of molecular biology and computer science in recent years, cross-generation, Especially construction and application of new DNA molecular logic gates, attracted wide attention of researchers at home and abroad.Input and output signals of logic gates target DNA molecule is at the molecular level to achieve a complex of two or more of computing systems, the output signal of the object is obtained by calculating the corresponding logic gate signal to the signal applied to the "1" and "0" binary logic operation, enabling digital operation.Thrombin is a very important role in the coagulation cascade human anticoagulant, procoagulant the bioactive protein. In recent years, clinical, laboratory, pathology and pharmacology studies have shown. Coagulation system in the majority of cancer patients who are active, thrombin promotes the growth of tumor cells. Spontaneity invasion and metastasis, but also stimulate tumor angiogenesis. Thus, rapid and accurate detection of thrombin has important practical significance.MicroRNA (miRNA) are a class of endogenous gene encoding a length of about 20-25 nucleotides in non-coding single-stranded RNA molecules, after their participation in the transcriptional regulation of gene expression in plants and animals. Therefore, to build a new type of DNA molecule logic gate of miRNA diagnosis is very important.Logic gate is the most basic application in integrated circuits. Integrated circuits, transistors are the constituent elements of simple logic gates. DNA molecules are DNA molecules for the logic gate of the logic gate operation target using a DNA molecule as the input signal to the logic gate, in order to achieve DNA level logical operation. DNA logic gate has an important role in molecular computers and computing technology on the molecular level.Based on this, the thesis based on molecular engineering and nanotechnology to build a new type of DNA molecule logic gate and miRNA detection of thrombin, mainly carried out on the following three aspects:1. Developed based on aptamers-DNA molecular logic gate system colloidal gold technology to build proteins. We thrombin protein detection target to build a new type of DNA molecular logic gates. In this system, the aptamer and thrombin colloidal gold technology for protein analysis through the use of functionalized gold nanoparticles, and colloidal gold clever design and aptamer target substance combined with the choice of thrombin proteins as logic gates input, the use of thrombin aptamer specifically binding thereto, the release of colloidal gold-binding site on the monitoring probe, causing agglomeration of colloidal gold, to implement logic gate output signal.2. Development of the substitution reaction based chain-fluorescence resonance energy transfer construct microRNA molecule of DNA logic gate system. We microRNA-195 and microRNA-21 for the detection of target DNA molecules to build a new type of logic gate. microRNA-195 is a miRNA-15a,15b,16,195,497 family important one, which plays an important role in the development of tumors and the malignant behavior regulation. microRNA-21-encoding gene located in the 17q23.2, namely vacuolar membrane protein gene (vacuole membrane protein-1, VMP1) coding region, the tenth intron VMP1 gene. Study found that miRNA-21 play in the development of tumors in a very important role in the expression of different tumor types were noticeable increase in the proliferation of tumor cells, invasion, angiogenesis and the biological behavior of drug resistance and other related. We have constructed a diagnostic test based on miRNA-mediated FRET foothold strand displacement and two logic gates (YES AND gates and doors) system. Logic gates can perform simple calculations, and in response to the disease indicators show the correct symbol. Using foothold mediated strand displacement, in the proposed logic gates absence of enzyme under full output signal. Furthermore, in principle, any miRNA, including the level of specific genes or partial DNA sequence for any number of diseases, can be used as an input signal, we are built-in logic to detect and diagnose system. This shows that our logic gate having a potential value on the application of biological samples.3. We developed an exonuclease-based-DNA polymerase molecule logic gate system Construction of miRNA. We miRNA let-7d to build a new type of DNA molecular logic gates to detect targets. miRNAlet-7d miRNA let-7 family is one, the expression levels of let-7 family in lung cancer, ovarian cancer, breast cancer cells was significantly reduced. We based exonuclease I constructed a YES door. Exonuclease I have a great advantage, the first to join this exonuclease I, when there is no presence of the target substance detection systems for nucleic acid probes and primers are degraded, so that the fluorescent dye (SYBRGreen I, SG) the doping amount of the reaction mixture less, i.e., when the target substance is no input, the output signal of the fluorescence of fluorescent dyes particularly low, almost no output, that is, no input is no output, greatly reduces the background signal, improving the detection sensitivity. Second, adjusting the design of nucleic acid probes, the target binding domain are conducive to the length of the extension product of the polymerization reaction and amplifying the embedded fluorescent dyes in response to the signal SG, so that I exonuclease capable of degrading single-stranded very few, so when the target substance input, the output signal of the fluorescent dye is particularly evident, the detection limit can reach 50 zM.