麒麟鸡miRNA-1623与靶基因Wnt4的靶标验证分析

发布时间：2018-07-06 13:35

本文选题：麒麟鸡 + 卷羽　；参考：《基因组学与应用生物学》2017年03期

【摘要】：麒麟鸡(frizzle chicken)是我国亚热带地区特有的一种具有卷羽特性的优质肉鸡品种。本课题组前期研究发现在麒麟鸡皮肤中高度表达miRNA-1623(miR-1623),因此推测其参与麒麟鸡毛囊发育及卷羽的形成过程。为研究其独特的毛囊发育及卷羽散热机制,本研究利用生物信息学手段根据其靶标结合位点寻找到其潜在靶标Wnt4基因,并利用qRT-PCR对卷羽麒麟鸡和片羽怀乡鸡中的miR-1623与Wnt4的表达水平进行了检测。进一步通过构建双荧光素酶报告基因pmirGlo载体在细胞水平对miR-1623与Wnt4的靶标关系进行了验证。结果表明:Wnt4 mRNA的3'UTR存在miR-1623靶标结合位点(TGCCTG);相对于怀乡鸡,麒麟鸡皮肤中miR-1623表达水平显著升高(p0.01),Wnt4 mRNA表达水平显著降低(p0.01);双荧光素酶检测分析发现miR-1623靶向负调控Wnt4-3'UTR。本实验可为进一步揭示miRNA-1623在禽类毛囊生长发育过程中的作用提供参考。
[Abstract]:Steamed Sliced Chicken with Mushrooms and Vegetables (frizzle chicken) is a special high quality broiler breed in subtropical region of China. Our previous study found that miRNA-1623 (miR-1623) is highly expressed in the skin of Steamed Sliced Chicken with Mushrooms and Vegetables. Therefore, we speculate that miRNA-1623 is involved in the development of Steamed Sliced Chicken with Mushrooms and Vegetables hair follicles and the formation of curly feathers. In order to study its unique mechanism of hair follicle development and heat dissipation, we used bioinformatics to find its potential target Wnt4 gene according to its target binding site. The expression levels of miR-1623 and Wnt4 were detected by qRT-PCR in curly feather Steamed Sliced Chicken with Mushrooms and Vegetables and Shiyu Huai Xiang chicken. Furthermore, the targeting relationship between miR-1623 and Wnt4 was verified at the cell level by constructing a double luciferase reporter gene pmirGlo vector. The results showed that there was a miR-1623 target binding site (TGCCTG) in 3UTR of Wnt4 mRNA, a significant increase in miR-1623 mRNA expression level in Steamed Sliced Chicken with Mushrooms and Vegetables skin (p0.01) and a significant decrease in Wnt4 mRNA expression level (p0.01) in Steamed Sliced Chicken with Mushrooms and Vegetables skin, and a negative regulation of Wnt4-3UTR targeting miR-1623 was found by double luciferase assay. This study may provide a reference for further revealing the role of miRNA-1623 in the growth and development of poultry hair follicles.
【作者单位】：广东海洋大学动物科学系;中国农业科学院特产研究所特种经济动物分子生物学国家重点实验室;
【基金】：广东省创新强校重大项目(GDOU2013050222) 国家科技基础条件平台-特种经济种质资源共享子平台(2016020)共同资助
【分类号】：S831

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