牛胚胎期肌肉发育关键lncRNA的筛选及功能分析

doi:10.11843/j.issn.0366-6964.2025.08.021

摘要/Abstract

摘要：

旨在分析胚胎时期牛肌肉组织中的lncRNA表达谱，进而筛选出可能与肌肉发育相关的lncRNA。本试验使用通过胚胎分割技术获得的遗传背景高度一致的和牛3个时期肌肉组织样品(3月龄胎儿、6月龄胎儿和9月龄出生个体)进行转录组测序，对鉴定到的lncRNA进行差异分析，筛选胚胎肌肉发育过程中的关键lncRNA，并对其功能和靶标基因进行生物信息学分析。测序结果显示，在不同时期差异表达的lncRNAs有9 913条，在不同时期差异表达的mRNA转录本有3 079条，对两者进行聚类分析发现在牛3月龄胎儿肌肉组织中差异表达的mRNA和lncRNA均最多。筛选差异lncRNA上下游10 kb及以内的蛋白编码基因为lncRNA的靶基因，发现lncRNA共有2 064个顺势调控的靶基因，将其与差异mRNA进行韦恩分析共得到448个靶基因，通过对交集靶基因进行GO和KEGG富集分析，最终筛选出与肌肉发育密切相关的99个lncRNAs和27个候选靶标mRNAs，结合测序中两者的表达量数据分别对其进行STEM分析，发现lncRNA和mRNA分别聚类到一个显著的profile，包含12个mRNAs和68个lncRNAs，且两者的表达量均属于先上升再下降的趋势。最后，使用Cytoscape通过顺式调控关系对这一表达趋势的12个mRNAs和68个lncRNAs进行调控网络构建，发现大多数lncRNA可能对SERP1、FXR1和DVL3具有调控功能。通过研究结果发现3月龄可能是牛肌肉发育的关键时期，且筛选出的68个lncRNAs可能通过竞争性吸附对应的miRNA进而影响相应靶基因的表达，从而在牛不同时期骨骼肌生长发育中起调控作用。

关键词: RNA测序, 生物信息学分析, 牛肌肉组织, lncRNA

Abstract:

This study aimed to analyze the expression profiles of long non-coding RNAs (lncRNAs) in bovine muscle tissues during embryonic development, with the goal of identifying lncRNAs potentially involved in muscle development. Transcriptome sequencing was conducted on muscle tissue samples from 3 developmental stages (3-month-old fetuses, 6-month-old fetuses, and 9-month-old newborns obtained from genetically homogeneous Japanese Black cattle using embryo splitting techniques). Differential expression analysis was performed on the identified lncRNAs to screen key lncRNAs involved in embryonic muscle development. Subsequent bioinformatic analyses were carried out to predict the functions and target genes of these lncRNAs. Sequencing data revealed 9 913 differentially expressed lncRNAs and 3 079 differentially expressed mRNA transcripts across the developmental stages. Clustering analysis indicated that the highest number of differentially expressed lncRNAs and mRNAs occurred in the 3-month-old fetal muscle tissue. By identifying protein-coding genes located within 10 kb upstream or downstream of the differentially expressed lncRNAs, 2 064 cis-regulated target genes were predicted. A Venn diagram analysis comparing these target genes with the differentially expressed mRNAs identified 448 overlapping genes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of these overlapping genes highlighted 99 lncRNAs and 27 candidate target mRNAs closely associated with muscle development. Short time-series expression miner (STEM) analysis of expression data revealed that both lncRNAs and mRNAs clustered into a significant profile characterized by an initial increase followed by a decrease in expression levels, encompassing 68 lncRNAs and 12 mRNAs. Finally, a regulatory network was constructed using Cytoscape based on cis-regulatory relationships based on the 68 lncRNAs and 12 mRNAs, suggesting that the majority of these lncRNAs may regulate genes such as SERP1, FXR1, and DVL3. The results showed that the 3-month-old period may be a critical period for bovine muscle development, and the 68 lncRNAs screened may affect the expression of corresponding target genes through competitive adsorption of corresponding miRNAs, thereby playing a regulatory role in the growth and development of bovine skeletal muscles at different stages.

Key words: RNA-Seq, bioinformatics analysis, bovine muscle tissue, lncRNA

中图分类号:

S823.2

万琼飞, 石闪闪, 郭若楠, 吕航, 胡德宝, 郭益文, 张林林, 丁向彬, 郭宏, 李新. 牛胚胎期肌肉发育关键lncRNA的筛选及功能分析[J]. 畜牧兽医学报, 2025, 56(8): 3802-3812.

WAN Qiongfei, SHI Shanshan, GUO Ruonan, Lü Hang, HU Debao, GUO Yiwen, ZHANG Linlin, DING Xiangbin, GUO Hong, LI Xin. Screening and Functional Analysis of Key lncRNAs of Bovine Embryonic Muscle Development[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(8): 3802-3812.

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样本 Sample	有效数据 Valid reads	比对上的数据 Mapped reads	唯一比对 Unique mapped	多个比对 Multi mapped
3月龄胎儿ET3_SM	225 995 560	190 748 763(84.40%)	179 083 290(79.24%)	11 665 473(5.16%)
6月龄胎儿ET6_SM	133 685 384	82 960 660(62.06%)	56 607 053(42.34%)	26 353 607(19.71%)
9月龄出生个体ET9_SM	180 918 318	84 074 062(46.47%)	77 060 274(42.59%)	7 013 788(3.88%)

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