【摘要】 旨在分析猪miR-145-5p的序列特征及其生物学功能，验证其在猪卵巢颗粒细胞(GCs)凋亡中的作用。利用生物信息学方法获取猪miR-145-5p前体序列并分析其序列信息、结构和生物学功能;设计合成miR-145-5p反义寡核苷酸抑制剂，转染至体外培养的猪卵巢GCs中，实时荧光定量PCR(RT-q PCR)检测miR-145-5p抑制效率;通过流式细胞术检测GCs凋亡率。结果:猪miR-145-5p前体序列长度为86 bp，其核苷酸序列在哺乳动物中保守性较高;功能富集分析发现miR-145-5p靶基因主要富集在与细胞增殖、存活、代谢和凋亡调控作用相关的磷脂酰肌醇-3-激酶/蛋白激酶(PI3K-Akt)、丝裂原活化蛋白激酶(MAPK)、叉头框O蛋白(Fox O)、转化生长因子β(TGF-β)等信号通路;转染miR-145-5p抑制剂，流式细胞术分析发现抑制miR-145-5p的表达，能显著降低GCs的凋亡率(P<0.05)。研究表明，miR-145-5p具有促进猪卵巢GCs凋亡的生物学作用。更多还原

【Abstract】 To analyze the sequence characteristics of porcine miR-145-5p and its biological function,and to verify its role in apoptosis of porcine ovarian granulosa cells,in this study,bioinformatic approaches were used to obtain the porcine miR-145-5p precursor sequence and to analyze its sequence information,structure and biological function. Then,an miR-145-5p antisense oligonucleotide inhibitor was designed and synthesized,and was transfected into porcine ovarian granulosa cells cultured in vitro. Next,the knockdown effect was detected by RTq PCR. Finally,the apoptosis rate of the granulosa cells was detected by flow cytometry. The results showed that the length of the porcine miR-145-5p precursor sequence was 86 bp,and its nucleotide sequence was highly conserved among mammals. The functional enrichment analysis revealed that the target genes of miR-145-5p were mainly enriched in the signaling pathways such as PI3K-Akt,MAPK,Fox O,and TGF-β,which were related to the regulatory effects of cell proliferation,survival,metabolism,and apoptosis. After transfection of the miR-145-5p inhibitor,the flow cytometry analysis showed that inhibition of miR-145-5p expression significantly reduced the apoptosis rate of the granulosa cells( P<0. 05). In conclusion,miR-145-5p might promote apoptosis of porcine ovarian granulosa cells.更多还原