發(fā)布時間：2018-03-03 13:41

  本文選題：miR-203　切入點：miR-217　出處：《浙江理工大學》2017年碩士論文　論文類型：學位論文

【摘要】：miRNA的作用是通過同靶mRNA的3’端靶向結合降解或者是抑制其靶基因翻譯,進而可以對靶基因的轉錄后進行調(diào)控。另mi RNA可識別相應的靶基因且不是單一映射關系,同時存有多個不同miRNA調(diào)控同一靶基因,而miRNA也可以調(diào)控多個不同的靶基因。我們前期預測發(fā)現(xiàn)斑馬魚中nup43是miR-203的靶基因,并且在三唑磷,化學名稱O,O-二乙基-O-(1-苯基-l,2,4-三唑-3-基)硫代磷酸酯的脅迫下nup43的表達受miR-203的調(diào)控。在本實驗中,課題組利用Microcosm Targets軟件預測得出nup43為miR-203與miR-217共同調(diào)控靶基因,通過實驗驗證了這2個miRNAs分別與nup43之間的關系。我們評估了三唑磷暴露下斑馬魚中這2個mi RNAs的表達,研究2種miRNAs能否調(diào)控nup43的表達,結果發(fā)現(xiàn)隨著三唑磷處理濃度的升高,斑馬魚中miR-203基因的表達量顯著上調(diào),斑馬魚中miR-217基因的表達量顯著下調(diào)。mi RNA可作為環(huán)境化學物質或致癌基因得生物標記物。為了解釋三唑磷處理后miRNA表達的變化,進一步驗證生物信息學預測的結果。我們通過將nup43 3’-UTR中miR-203和miR-217結合位點分別敲除,成功構建了2個重組海腎熒光素酶載體,分別命名為pRL/S-K.O-203、pRL/S-K.O-217。通過雙熒光素酶報告基因法驗證了斑馬魚nup43 3’-UTR中存在miR-217的結合位點。我們通過對ZF4細胞轉染miR-217 mimic后,檢測細胞中nup43的表達量,發(fā)現(xiàn)nup43在mRNA水平及蛋白水平都顯著下調(diào),而轉染miR-217 inhibitor后,nup43的mRNA水平及蛋白水平則顯著上調(diào),但對ZF4細胞轉染miR-203 mimic后,其nup43表達量為上調(diào),但轉染miR-203 inhibitor后,其nup43表達量為顯著下調(diào)。依上述可得,nup43是miR-203和miR-217的靶基因,在三唑磷暴露下miR-203與miR-217的變化表明其可以作為暴露于三唑磷環(huán)境的生物標志物。
[Abstract]:The role of miRNA is to degrade or inhibit the translation of target gene by targeting the 3 '-terminal binding of target mRNA, and then to regulate the target gene after transcription. MiRNA can recognize the corresponding target gene and is not a single mapping relationship. At the same time, there are many different miRNA to regulate the same target gene, and miRNA can also regulate many different target genes. We predicted that nup43 in zebrafish is the target gene of miR-203, and in triazophos, The expression of nup43 was regulated by miR-203 under the stress of OO2-diethyl-O-1-phenyl-1-phenyl-1-phenyl-1-phenyl-1-phenyl-1-diethyl-1-triazolyl) phosphate. In this experiment, we predicted that nup43 is a target gene regulated by miR-203 and miR-217 by Microcosm Targets software. We evaluated the expression of miRNAs in zebrafish exposed to triazophos and studied whether the two miRNAs could regulate the expression of nup43. The results showed that the expression of nup43 increased with the concentration of triazophos. The expression of miR-203 gene in zebrafish was upregulated, and the expression of miR-217 gene in zebrafish was down-regulated. Mi RNA could be used as an environmental chemical or a biomarker of oncogene, in order to explain the change of miRNA expression after triazophos treatment. The results of bioinformatics prediction were further verified. By knockout the binding sites of miR-203 and miR-217 in nup43 3, we successfully constructed two recombinant marine kidney luciferase vectors. They were named as pRL / S-K.O-203 and pRL / S-K.O-217.The binding sites of miR-217 in zebrafish nup43 3O -UTR were verified by double luciferase reporter gene method. After transfection of ZF4 cells into miR-217 mimic, we detected the expression of nup43 in ZF4 cells. It was found that nup43 was significantly down-regulated in mRNA and protein levels, while mRNA and protein levels of nup43 were up-regulated after transfection of miR-217 inhibitor, but nup43 expression was up-regulated in ZF4 cells after transfection of miR-203 mimic, but after transfection of miR-203 inhibitor. Nup43 was the target gene of miR-203 and miR-217. The changes of miR-203 and miR-217 under triazophos exposure indicated that it could be used as a biomarker of triazophos exposure.
【學位授予單位】：浙江理工大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：X174;Q78

【參考文獻】

中國期刊全文數(shù)據(jù)庫 前10條

1 王玉倩;薛秀花;;實時熒光定量PCR技術研究進展及其應用[J];生物學通報;2016年02期

2 Fang Yang;Li-Zhi Lv;Qiu-Cheng Cai;Yi Jiang;;Potential roles of EZH2, Bmi-1 and mi R-203 in cell proliferation and invasion in hepatocellular carcinoma cell line Hep3B[J];World Journal of Gastroenterology;2015年47期

3 馮京;徐亮;;核孔復合物與心肌分化、增殖及心臟疾病的研究進展[J];生命科學;2015年10期

4 李紅艷;張U，

本文編號：1561265