本文選題：RNA納米花 + 癌細胞��； 參考：《曲阜師范大學》2017年碩士論文

【摘要】：本論文在利用熒光、紫外吸收和激光掃描共聚焦顯微鏡等技術下,利用功能化核酸折紙方法和基于雜交鏈式反應的比色放大法分別對細胞進行聯合給藥、靶向治療及miRNA、ATP的檢測。本文主要內容如下:1.基于磁性RNA納米花用于復合給藥和靶向治療的診斷-治療一體化系。本部分開發(fā)了一種基于磁性RNA納米花用于復合給藥和靶向治療的診斷-治療一體化系統。與傳統的核酸結構相比之下,通過將磁性納米粒子(MNP)引入到中可使得RNA NF分離方便。葉酸(FA)修飾的MNP/RNA NF(FA/MNP/RNA NF)作為一種具有優(yōu)良的生物相容性納米靶向載體克服了MNP/RNA NF的非選擇性。然后,抗癌藥物阿霉素(DOX)和光敏劑四甲基吡啶卟啉(TMPyP4)結合RNA NF被用作共同藥物運輸模型,RNA NF首次被用于聯合給藥。因此,成像熒光標記,目標識別元素和藥物分子都聚集在MNP/RNA NF的表面。實驗結果表明,聯合給藥平臺(FA/MNP/RNA NF/D/T)的療效優(yōu)于單一給藥平臺(FA/MNP/RNA NF/D)。此外,FA/MNP/RNA NF被作為探針用于癌細胞的檢測,檢出限為500 cell/m L。綜上所述,對于其他生物分子的細胞內定量,基于FA/MNP/RNA NF的共給藥平臺以及診斷-治療一體化系統是一種很有前途的方法。2.一種通用的基于雜交鏈式反應的比色放大法對核酸和適體特異性配體的檢測。我們提出了一種基于金納米粒子-DNA(GNP-DNA)雜交鏈式反應(HCR)并用于檢測核酸和適體特異性配體的通用放大比色法。這種通用的陣列由捕獲探針和發(fā)夾DNA-GNP組成。首先,捕獲探針特異性地識別目標分子,并導致引發(fā)劑序列的釋放。然后,發(fā)夾DNA修飾的金納米粒子通過引發(fā)劑引發(fā)的HCR進而由分散態(tài)聚集成相互交聯的聚集體。我們采用miRNA的靶序列(miRNA-203)和核酸適體的特異性配體(ATP)作為靶分子來驗證這種方法。在miRNA-203的強競爭力下,引發(fā)序列(DNA 2)從捕獲探針(MNP/DNA1/2共軛物)釋放,發(fā)夾DNA(H 1和H 2)可以與DNA 2互補從而形成GNP-H1/GNP-H2聚集體,所用miRNA-203濃度在1.0×10-11-3.0×10-10M范圍內時,記錄GNP-H1/GNP-H2聚集體溶液在620 nm和520 nm處的吸光度比值(A620/A520),所得檢出限為1.0×10-11M。與此同時,溶液中的顏色由淡紅色變?yōu)樽仙?然后再變?yōu)榈{色。對于ATP,在適配體與ATP的強的相互作用下,引發(fā)序列(DNA 3的5’端)從捕獲探針(DNA3)釋放,我們所設計的比色法對ATP檢測具有良好的靈敏性,檢測限為1.0×10-8M ATP。該策略對于細胞內核酸和核酸適配體的特異性配體的定性分析與定量分析也顯示出良好的性能。
[Abstract]:In this paper, using fluorescence, ultraviolet absorption and laser scanning confocal microscope, we used functional nucleic acid origami method and colorimetric amplification method based on hybrid chain reaction to carry out combined administration, targeted therapy and the detection of miRNA-ATP. The main contents of this paper are as follows: 1. A Diagnostic and Therapeutic Integration system based on Magnetic RNA Nanoflowers for combined Administration and targeted Therapy. In this part, an integrated diagnostic and therapeutic system based on magnetic RNA nanosplast for combined drug delivery and targeted therapy is developed. Compared with the traditional nucleic acid structure, RNA NF can be separated easily by introducing magnetic nanoparticles into the nucleic acid structure. Folate FA-modified MNP/RNA NF(FA/MNP/RNA NFS as a good biocompatible nano-target carrier overcomes the nonselectivity of MNP/RNA NF. Then, the anticancer drug dox) and the Guang Min agent TMPyP4) combined with RNA NF were used as a common drug transport model for the first time. Therefore, imaging fluorescent labeling, target recognition elements and drug molecules are clustered on the surface of MNP/RNA NF. The results showed that the combined drug delivery platform, FAR / MNPR / RNA NFR / D / T, was more effective than the single drug delivery platform, namely, FAR / MNPR / RNA NFR / DX. In addition, FAP / MNPrRNA NF was used as a probe to detect cancer cells with a detection limit of 500 cell/m / L. In conclusion, for the intracellular quantification of other biomolecules, the co-delivery platform based on FA/MNP/RNA NF and the integrative system of diagnostics and therapy are promising methods. A universal colorimetric amplification method based on hybrid chain reaction for detection of nucleic acid and aptamer specific ligands. We propose a universal amplification colorimetric method based on gold nanoparticles (DNA-GNP-DNA) hybridization chain reaction (HCR) to detect nucleic acid and aptamer specific ligands. The universal array consists of a capture probe and a hairpin DNA-GNP. First, the target molecule is specifically identified by the capture probe, which leads to the release of the initiator sequence. Then, the gold nanoparticles modified by hairpin DNA were synthesized into cross-linked aggregates by HCR initiated by the initiator. The target sequence of miRNA and the aptamer specific ligand ATP) were used as target molecules to verify this method. Under the strong competitiveness of miRNA-203, the initiation sequence DNA2) was released from the capture probe MNP / DNA 1 / 2 conjugate. Hairpin DNA(H _ 1 and H _ 2) could complement DNA _ 2 to form GNP-H1/GNP-H2 aggregates. When the miRNA-203 concentration was 1.0 脳 10 ~ (-11) to 3.0 脳 10 ~ (-10) M, The absorbance ratio of GNP-H1/GNP-H2 aggregates at 620nm and 520nm was recorded. The detection limit was 1.0 脳 10-11M. At the same time, the color in the solution changed from light red to purple and then to pale blue. For ATP, the 5 'end of the initiation sequence of ATP is released from the capture probe DNA3 under the strong interaction between aptamer and ATP. Our colorimetric method is sensitive to ATP detection with a detection limit of 1.0 脳 10 ~ (-8) M ATP. This strategy also shows good performance for qualitative analysis and quantitative analysis of specific ligands of nucleic acid and aptamer.
【學位授予單位】：曲阜師范大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R318;O657.3

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本文編號：1947871