本文選題：紅藻 + microRNA　； 參考：《山西大學(xué)》2016年博士論文

【摘要】：MicroRNAs (miRNAs)是一類廣泛存在于真核生物中的內(nèi)源性非編碼小RNAs,其長(zhǎng)度約為22nt,有典型的頸環(huán)狀前體二級(jí)結(jié)構(gòu),通過(guò)與靶位點(diǎn)的特異性結(jié)合,以切割降解靶mRNA或翻譯抑制的方式來(lái)調(diào)控靶基因的轉(zhuǎn)錄后表達(dá)。研究表明,miRNAs在調(diào)節(jié)生物體的生長(zhǎng)發(fā)育、新陳代謝、細(xì)胞分化與凋亡、脅迫應(yīng)答等方面發(fā)揮著重要的作用,是當(dāng)前生命科學(xué)領(lǐng)域研究的熱點(diǎn)。紅藻是一類地球上進(jìn)化時(shí)間較早、結(jié)構(gòu)相對(duì)簡(jiǎn)單、類群較為龐大、功能多樣的藻類。它們不僅具有經(jīng)濟(jì)和工業(yè)價(jià)值,有的還具有醫(yī)藥、保健等多種特殊的生物學(xué)功能。盡管目前在動(dòng)植物中已發(fā)現(xiàn)上萬(wàn)條miRNAs,但在低等植物紅藻中卻鮮有相關(guān)報(bào)道,對(duì)其相關(guān)功能的研究則更少。本文選取皺波角叉菜、細(xì)齒麒麟菜、紫球藻和溫泉紅藻四種不同的紅藻,運(yùn)用高通量測(cè)序技術(shù)聯(lián)合生物信息學(xué)技術(shù)首次對(duì)四種紅藻的miRNAs進(jìn)行了鑒定和特征分析,這對(duì)于進(jìn)一步充實(shí)藻類miRNA數(shù)據(jù),彌補(bǔ)四種紅藻miRNAs研究的空白,以及對(duì)四種紅藻特殊功能的深入研究具有十分重要的理論和現(xiàn)實(shí)意義。本文的主要研究結(jié)果與意義小結(jié)如下：1、運(yùn)用深度測(cè)序技術(shù)聯(lián)合生物信息學(xué)技術(shù)從四種紅藻中共鑒定出miRNAs 645條,屬于596個(gè)miRNA家族,其中保守miRNAs 562條,物種特異miRNAs 83條。2、運(yùn)用統(tǒng)計(jì)學(xué)軟件和生物信息學(xué)技術(shù)從數(shù)量、長(zhǎng)度、堿基偏好性、二級(jí)結(jié)構(gòu)、基因家族保守性、靶位點(diǎn)調(diào)控方式等方面對(duì)四種紅藻miRNAs的特征進(jìn)行了分析。分析結(jié)果表明,本研究鑒定的四種紅藻miRNAs基本可信。3、用Northern blot和stem-loop qRT-PCR兩種不同的實(shí)驗(yàn)方法分別對(duì)部分紅藻miRNAs進(jìn)行驗(yàn)證。驗(yàn)證分析的結(jié)果進(jìn)一步表明,本文鑒定的四種紅藻miRNAs基本可信。4、用psRobot和TargetFinder兩種軟件在四種紅藻miRNAs中共預(yù)測(cè)到上萬(wàn)條的靶基因和靶位點(diǎn),psRNATarget軟件預(yù)測(cè)到切割抑制為紅藻miRNAs與其靶位點(diǎn)間的主要作用方式,這為將來(lái)某些重要miRNA靶基因的表達(dá)驗(yàn)證及其調(diào)控方式的深入研究奠定了基礎(chǔ)。5、基于Gene Ontology平臺(tái),我們發(fā)現(xiàn)四種紅藻中預(yù)測(cè)的靶基因種類繁多、功能各異,多數(shù)可歸為生物學(xué)過(guò)程、分子功能和細(xì)胞組分三大類,這也體現(xiàn)了miRNAs功能的多樣性。某些潛在的,富集程度較高的,可能在紅藻中具有特殊生物學(xué)意義的候選靶基因?qū)⑹墙窈笤敿?xì)闡明紅藻某些重要特征及功能的突破口。6、通過(guò)KEGG pathway分析,我們預(yù)測(cè)到PPAR信號(hào)通路、卟啉及葉綠素代謝通路、FcyR介導(dǎo)的吞噬作用和RNA轉(zhuǎn)運(yùn)通路分別為四種不同紅藻靶基因參與調(diào)控下的顯著富集的代謝通路。這為深入探究各代謝通路,闡明其在紅藻生命活動(dòng)中的重要作用奠定了基礎(chǔ)。7、運(yùn)用Cytoscape軟件繪制的miRNA-靶基因互作網(wǎng)絡(luò)圖,以及基于統(tǒng)計(jì)學(xué)計(jì)算下的四種紅藻間miRNA數(shù)據(jù)的比較分析結(jié)果,將從整體化和個(gè)性化兩個(gè)角度為紅藻miRNAs的深入研究指明方向。此外,miRNA基因家族的進(jìn)化分析將是未來(lái)藻類miRNA研究的新方向。
[Abstract]:MicroRNAs is a class of endogenous non-coding small RNAss widely found in eukaryotes. Its length is about 22nt.It has a typical secondary structure of cervical ring precursor, and it binds specifically to the target site. The post-transcriptional expression of target gene is regulated by cleavage and degradation of target mRNA or translation inhibition. Studies have shown that miRNAs play an important role in regulating the growth and development of organisms, metabolism, cell differentiation and apoptosis, stress response and so on. Red algae is a kind of algae which evolved earlier on the earth, has relatively simple structure, has a large group of species, and has a variety of functions. They not only have economic and industrial value, but also have many special biological functions, such as medicine, health care and so on. Although tens of thousands of miRNAss have been found in plants and animals, there are few reports on the related functions of red algae. In this paper, four different species of red algae, Carrageenia crassifolia, Eucheuma tenuifolia, Chlorella purpurea and red algae from hot springs were selected. The miRNAs of four species of red algae were identified and characterized for the first time by using high-throughput sequencing technique and bioinformatics technique. It is of great theoretical and practical significance to further enrich the miRNA data of algae, to make up for the blank in the study of four species of red algae miRNAs, and to further study the special functions of the four species of red algae. The main results and significance of this study are summarized as follows: 1. A total of 645 miRNAs fragments were identified from four species of red algae by using deep sequencing technique and bioinformatics technique, belonging to 596 miRNA families, among which 596 were conserved miRNAs 562. The characteristics of four species of red algae miRNAs were analyzed by statistical software and bioinformatics from the aspects of quantity, length, base preference, secondary structure, gene family conservatism and target site regulation. The results showed that the miRNAs of four species of red algae identified in this study was basically credible. The Northern blot and stem-loop qRT-PCR methods were used to verify the miRNAs of some red algae. The results of the verification analysis further show that, Four species of red algae miRNAs were identified in this paper. PsRobot and TargetFinder software were used to predict tens of thousands of target genes and target sites in four species of red algae miRNAs. The software predicted that the cleavage inhibition was the main mode of interaction between miRNAs and its target sites. This laid a foundation for the further study of the expression and regulation of some important miRNA target genes in the future. Based on the Gene Ontology platform, we found that the predicted target genes in the four species of red algae have various types and functions, most of which can be classified as biological processes. Molecular function and cell composition are divided into three categories, which also reflect the diversity of miRNAs functions. Some potential, highly enriched candidate target genes, which may have special biological significance in red algae, will be a breakthrough to clarify some important characteristics and functions of red algae in the future. By KEGG pathway analysis, we can predict the PPAR signaling pathway. The phagocytosis mediated by FcyR and the RNA transport pathway by porphyrin and chlorophyll metabolism pathway were four highly enriched metabolic pathways regulated by target genes of red algae respectively. This laid a foundation for further exploring the metabolic pathways and elucidating their important role in the life activities of red algae. The miRNA-target gene interaction network map made by Cytoscape software was used to map the interaction of miRNA-target genes. The results of comparison and analysis of miRNA data among four species of red algae based on statistical calculation will point out the direction of further research on miRNAs from the aspects of integration and individuation. In addition, phylogenetic analysis of miRNA gene family will be a new direction of algal miRNA research in the future.
【學(xué)位授予單位】：山西大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：Q943.2

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本文編號(hào)：1950218