鸡miR-10b-5p生物信息学及组织表达分析

doi:10.16431/j.cnki.1671-7236.2022.04.007

摘要/Abstract

摘要： 【目的】通过对苏禽3号肉鸡的miR-10b-5p进行靶基因预测、生物信息学分析及其在鸡不同生长时期组织表达变化规律研究,探索其在鸡肌肉生长发育中作用。【方法】通过文献和miRBase检索脊椎动物的miR-10b-5p序列,利用Ensembl数据库查询并确定miR-10b-5p在基因组中的位置,根据前体序列构建系统进化树;使用miRDB、microT和TargetScan网站预测miR-10b-5p靶基因,并对靶基因进行GO功能和KEGG通路分析,进而揭示miR-10b-5p的基因功能。采用实时荧光定量PCR技术检测不同生长时期鸡miR-10b-5p组织表达量,探索其在大脑、小脑、垂体、下丘脑、胸肌、腿肌、卵巢、心脏、肝脏、脾脏、肺脏和肾脏组织中表达变化。【结果】 miRBase检索共获得59种动物59条miR-10b-5p序列,即每一种动物都只有一种miR-10b-5p形式。基因定位分析发现,鸡miR-10b-5p位于2号染色体上的HOX基因家族中。常见物种miR-10b-5p成熟序列比对分析结果表明,miR-10b-5p的成熟序列相似性较高,物种间较为保守。系统进化树分析发现,miR-10b在哺乳类中的灵长目、啮齿目、鸟类、鱼类中各自先聚为一支,然后再共同聚为一类,这表明miR-10b在进化过程中是保守的。靶基因预测发现,miR-10b-5p共有300个靶基因。GO功能分析发现,靶基因主要富集到染色质组装和基因表达的转录后调控、组蛋白甲基转移酶复合物、转录因子复合物等功能。KEGG通路分析表明,靶基因主要富集到Wnt和p53信号通路上。组织表达分析显示,miR-10b-5p在检测的各个组织中均有表达;3日龄时大脑、小脑、垂体、胸肌和腿肌中miR-10b-5p表达量显著高于心脏、下丘脑、肾脏和肺脏等组织(P<0.05);90日龄时,垂体、胸肌、腿肌和大脑中miR-10b-5p表达量显著高于其他组织(P<0.05)。与3日龄雏鸡相比,90日龄时垂体、胸肌和腿肌中miR-10b-5p表达量均显著上升(P<0.05)。【结论】鸡miR-10b-5p是组织广泛表达的miRNA,miR-10b在进化过程中是保守的,其可能通过Wnt及p53信号通路调控肌肉细胞增殖分化,进而调控鸡肌肉生长发育。

关键词: 鸡; miR-10b-5p; 靶基因; 表达分析; 生物信息学

Abstract: 【Objective】 The target gene prediction,bioinformatics analysis and tissue expression of miR-10b-5p in Suqin No.3 chickens at different growth stages were carried out to explore its role in chicken muscle growth and development.【Method】 The miR-10b-5p sequence of vertebrates were obtained through scientific research and miRBase database,the position of miR-10b-5p was determined using the Ensembl database,the phylogenetic tree was constructed based on the miRNA precursor sequence.The target genes of miR-10b-5p were predicted by using miRDB,microT and TargetScan websites,then GO function and KEGG pathway analysis were performed to reveal the function of miR-10b-5p gene.The expression changes of miR-10b-5p were detected by Real-time quantitative PCR in brain,cerebellum,pituitary gland,chest muscle,leg muscle,ovary,heart,liver,spleen,lung and kidney of chickens at different growth stages.【Result】 A total of 59 miR-10b-5p sequences from 59 species of animals were retrieved by miRBase.Each animal had only one miR-10b-5p form.Gene mapping analysis revealed that chicken miR-10b-5p was located in HOX gene family cluster on chromosome 2.Alignment analysis of the mature sequence of miR-10b-5p in common species showed that the mature sequence of miR-10b-5p had high homology and was relatively conserved among species.The phylogenetic tree analysis showed that miR-10b in primates,rodents,birds,and fishes in mammals were clustered first,and then clustered together into one group,which indicated that miR-10b was conservative in the process of evolution.Target genes prediction revealed that miR-10b-5p had 300 target genes.GO function analysis showed that target genes were mainly enriched mainly to chromatin assembly and post-transcriptional regulation of gene expression,histone methyltransferase complexes,transcription factor complexes and other functions.KEGG pathway analysis showed that target genes were mainly enriched in Wnt and p53 signaling pathways.Tissue expression analysis showed that miR-10b-5p was detective in all tissues tested, the expression of miR-10b-5p in brain, cerebellum, pituitary, chest and leg muscles at 3-day-old were significantly higher than those in heart, hypothalamus, kidney and lung (P<0.05). The expression of miR-10b-5p in pituitary, chest muscle, leg muscle and brain were significantly higher than that in other tissues at 90-day-old (P<0.05).Compared with 3-day-old chicks,the expression of miR-10b-5p in pituitary,chest and leg muscles of chickens at 90-day-old was significantly increased (P<0.05).【Conclusion】 Chicken miR-10b-5p was a miRNA that was widely expressed in tissues,and miR-10b was conserved in the evolutionary process,it might regulate muscle growth and cell proliferation and differentiation through Wnt and p53 signaling pathways.

Key words: chickens; miR-10b-5p; target gene; expression analysis; bioinformatics

中图分类号:

S831

黄正洋, 赵振华, 黄华云, 李春苗, 王钱保, 穆春宇, 黎寿丰. 鸡miR-10b-5p生物信息学及组织表达分析[J]. 中国畜牧兽医, 2022, 49(4): 1262-1270.

HUANG Zhengyang, ZHAO Zhenhua, HUANG Huayun, LI Chunmiao, WANG Qianbao, MU Chunyu, LI Shoufeng. Bioinformatics and Gene Expression Analysis of miR-10b-5p in Different Tissues of Chickens[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(4): 1262-1270.

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