miR-122在吉富罗非鱼肝脏脂肪沉积中的作用及其调控机制研究

项目名称： miR-122在吉富罗非鱼肝脏脂肪沉积中的作用及其调控机制研究

项目编号： No.31502143

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 农业科学

项目作者： 强俊

作者单位： 中国水产科学研究院淡水渔业研究中心

项目金额： 20万元

中文摘要： 肝脏是重要的脂代谢调控器官，长期摄食高脂肪饲料易引发罗非鱼肝脏脂肪沉积，影响生长、肉质与抗病力。miRNA是一种重要的基因调控分子，参与了多种信号传导过程。关于miRNA调控罗非鱼脂肪沉积的分子机制尚未清楚。前期通过miRNA表达谱差异分析，发现miR-122与罗非鱼肝脏脂肪沉积密切相关。本项目拟以miR-122为研究目标，首先构建高脂应激评价体系，通过生物信息学、荧光素酶报告基因系统与凝胶阻滞技术确定miRNA的上游调控基因；采用qRT-PCR、Northern blot和Western blot分析miRNA与靶基因表达特征；采用免疫组化和免疫荧光技术研究靶基因的组织定位与细胞定位；通过过表达与抑制目标miRNA从细胞与整体水平深入分析潜在的信号调控通路。研究结果将为揭示miRNA调控罗非鱼肝脏脂肪沉积的分子机制提供有益线索，为研究脂代谢相关调控途径，解决脂代谢类疾病提供新的学术资料。

中文关键词： 水产动物；胁迫；信号转导；基因表达调控；响应

英文摘要： Liver is an important organ for lipid metabolism regulation. The tilapia fed high lipid diets for a long term is subjected to lipid deposition in liver, which can affect growth, meat quality and disease resistance. miRNA has been proven to be crucial regulation factor and involved in a variety of signal transduction. Up to now, the regulation mechanism of miRNA related to lipid deposition in tilapia remains unclear. The comparative results of miRNA expression profiling between the tilapia fed normal diets and high lipid diets showed that miR-122 is closely related to lipid deposition. In this study, by taking miR-122 as candidate, we attempt to build evaluation system of stress in tilapia fed high lipid diets firstly; the potential target gene of miRNA will be verified by bioinformatics analysis, the luciferase reporter gene system and electrophoretic mobility shift assay(EMSA); the expression characteristics of miRNA and its target gene will be analyzed through real time polymerase chain reaction (qRT-PCR), Northern blot and Western blot; the protein distribution in tissues and in hepatocytes will be studied through immunohistochemistry (IHC) and immunofluorescence (IF) technology; potential signal transduction pathways will be deeply investigated by over-expression and locked nucleic acid (LNA)-expression of miRNA at the cell and overall levels. The results of this project will provide useful clues for the miRNA regulation on the mechanisms of lipid deposition in tilapia liver. Also, the project will benefit the investigation of regulation pathway, and adding new and valuable academic information for solving the disease in lipid metabolism.

英文关键词： aquatic animal;stress;signal transduction;regulation of gene expression;response