miR-126-5p在沙子岭猪睾丸组织差异表达分析及功能预测

摘要

In this study,real-time fluorescence quantitative PCR was performed to detect miR-126-5p expression levels in eight different developmental stages (E90,D1,D30,D60,D90,D120,D150,D180) of Shaziling pig testis to explore the role of miR-126-5p in animal testicular development and spermatogenesis.The miRanda,TargetScan and mirWalk databases were used to predict the target genes of miR-126-5p,then the intersection from the three databases were taken to analyze the biological function of target genes by making use of David online software.The Real-time PCR results showed that miR-126-5p highly expressed in embryonic period (E90-D1) and with down-regulated expression in growth periods(D1-D90),while up-regulated in the sexual maturity (D120).The D1 and D90 was the highest and the lowest expression level,and they were extremely significant(P<0.01) with the other seven periods;the differences among D30,E90,D180 were not significant (P>0.05),and they had extremely significant differences (P<0.01) with the rest of the periods;the difference between D120 and D30 was significant (P<0.05),and the difference between D120 and the other periods was extremely remarkable (P<0.01);there had no difference(P>0.05) between D150 and D60,but they had bally distinct difference with the rest of periods respectively (P<0.01).The target genes of miR-126-5p involved in cell proliferation,apoptosis,differentiation,cell communication,migration and protein transport biological processes (P<0.001),the molecular functions including protein dimerization activity and ligand-dependent nuclear receptor activity (P<0.001),KEGG pathway analysis results showed that miR-126-5p involved in TGF-β and MAPK signaling pathway,focal adhesion (P<0.01).Combined the results of miR-126-5p expression differences at different developmental stages of testicular tissue with target genes function prediction,miR-126-5p may be involved in the regulation of Shaziling pig germ cell differentiation and spermatogenesis,and this study could provide a research basis for the function and regulation mechanism of miR-126-5p in spermatogenesis.%利用实时荧光定量PCR(Real-time PCR)检测miR-126-5p在沙子岭猪睾丸组织8个不同发育时期(E90、D1、D30 、D60、D90、D120、D150、D180)表达量,以了解其在动物睾丸发育及精子生成中的作用.利用miRanda、TargetScan和mirWalk 3个数据库预测miR-126-5p靶基因并取交集,利用David在线软件分析miR-126-5p靶基因的生物功能.结果表明,miR-126-5p在胚胎期(E90-D1)睾丸组织中高表达,而在生长期(D1-D90)表达量下调,但在沙子岭猪性成熟(D120)后表达量又上调,其中D1表达量最高,D90表达量最低,且二者与其他7个时期均差异极显著(P<0.01);D30、E90、D180相互之间差异不显著(P>0.05),且分别与其余各时期差异极显著(P<0.01);D120与D30差异显著(P<0.05),与其余各时期差异极显著(P<0.01);D150与D60之间差异不显著(P>0.05),二者分别与其余各时期差异极显著(P<0.01).miR-126-5p靶基因参与细胞增殖、凋亡、分化、细胞通讯、迁移及蛋白质转运等生物学过程(P<0.001);靶基因分子功能包括蛋白二聚体活性、配体依赖性核受体活性(P<0.001);KEGG分析显示miR-126-5p靶基因参与TGF-β、MAPK、细胞黏附等信号通路(P<0.01).结合miR-126-5p在睾丸组织不同发育时期表达差异结果及其靶基因功能预测结果,miR-126-5p可能参与沙子岭猪生殖细胞分化及精子生成调控,本研究为miR-126-5p在动物精子生中的作用及其调控机制的研究提供依据.