敖汉细毛羊的粗毛返祖性状相关lncRNA-mRNA的联合测序分析

doi:10.11843/j.issn.0366-6964.2025.06.014

摘要/Abstract

摘要：

本研究从长链非编码RNA (long non-coding RNA, lncRNA)切入，旨在探究敖汉细毛羊群体中粗毛返祖(ancestral-like coarse, ALC)个体的转录组特性。本研究采集了3只30日龄ALC羔羊和3只30日龄非返祖(称为MF)细毛羔羊的皮肤样本，进行RNA测序(RNA-seq)，挖掘与细毛羊“返祖”现象存在潜在关联的mRNA和lncRNA，通过生物信息学分析探究lncRNA在细毛羊毛囊发育中的作用，并进行lncRNA与mRNA的联合分析。本研究筛选出1 540个差异表达lncRNA (differentially expressed lncRNA, DE-lncRNA)和513个差异表达基因(differentially expressed genes, DEGs)，富集到Wnt、PPAR、MAPK等多个与毛囊发育相关的信号通路中，均存在显著下调。对差异显著(P < 0.05)的DE-lncRNA进行分析，其中包含被认为与初级毛囊早期发育密切相关的lncRNA，如GTL2等。对DE-lncRNA靶向基因和DEGs进行GO和KEGG富集分析，DE-lncRNA靶基因和DEGs富集到WNT、PPAR、MAPK等多个与毛囊发育相关的信号通路，以及与TGF-β负调控相关的生物学过程。此外，DE-lncRNA和DEGs富集分析都显示，ALC羔羊有多个物质和能量代谢通路上调。DEGs与DE-lncRNA相关性分析筛选出90个可能受DE-lncRNA调控的基因，ADIPOQ、BRCA1、CCAR2、KIF20B、SOAT1等基因涉及与细胞增殖、细胞命运决定、细胞周期调节、物质结合、催化活性等有关的过程。结果显示，ALC羔羊在毛囊相关的lncRNA方面与MF羔羊存在显著差异，ALC羔羊的初级毛囊可能处在全面退化的初始阶段；MAPK和PI3K/AKT信号通路在ALC绵羊幼龄返祖现象比较重要；ALC羔羊代谢相较于MF羔羊更旺盛，GTL2等DE-lncRNA可能与ALC羊较强的适应性存在关联。本研究通过挖掘细毛羊ALC性状相关的lncRNA，为后期深入研究细毛羊毛囊发育机制提供参考线索，对细毛羊抗逆育种有重要参考价值。

关键词: 敖汉细毛羊, 粗毛返祖性状, lncRNA, mRNA, 生物信息学

Abstract:

This study started with long non-coding RNA (lncRNA) and aimed to explore the transcriptome characteristics of ancestral-like coarse (ALC) individuals in the Aohan fine wool sheep population. Skin samples were collected from 3 ALC lambs and 3 non ancestral modern fine (MF) wool lambs, all 30 days old. RNA sequencing (RNA-seq) was performed to identify mRNA and lncRNA potentially associated with the "ALC" phenomenon in fine wool sheep. Through bioinformatics analysis, we explored the role of lncRNA in the development of fine wool follicles and conducted joint analysis of lncRNA and mRNA. The total of 1 540 differentially expressed lncRNAs (DE-lncRNAs) and 513 differentially expressed genes (DEGs) were screened, which were enriched in multiple signaling pathways related to hair follicle development, such as WNT, PPAR, MAPK, and all showed significant downregulation. DE-lncRNAs with significant differences (P < 0.05) was analyzed, including lncRNAs closely related to early development of primary hair follicles, such as GTL2. GO and KEGG enrichment analysis was performed on DE-lncRNA target genes and DEGs. DE-lncRNA target genes and DEGs were enriched in multiple signaling pathways related to hair follicle development, such as WNT, PPAR, MAPK, as well as biological processes related to TGF-β negative regulation. In addition, DE-lncRNA and DEGs enrichment analysis showed that ALC lambs had multiple upregulated pathways for substance and energy metabolism. The correlation analysis between DEGs and DE-lncRNA identified 90 genes that might be regulated by DE-lncRNA, including ADIPOQ, BRCA1, CCAR2, KIF20B, SOAT1, which are involved in processes related to cell proliferation, cell fate determination, cell cycle regulation, substance binding, catalytic activity, etc. The results showed that ALC lambs had significant differences in hair follicle-associated lncRNAs compared to MF lambs, indicating that the primary hair follicles of ALC lambs might be in the initial stage of comprehensive degeneration. The MAPK and PI3K/AKT signaling pathways were potentially important for the phenomenon of juvenile atavism in ALC sheep. The metabolism of ALC lamb was more vigorous than that of MF lamb. GTL2 and other DE-lncRNAs might be associated with stronger adaptability in ALC sheep. This study aims to explore lncRNAs related to ALC traits in fine wool sheep, providing reference clues for further research on the development mechanism of fine wool sacs and having important reference value for stress resistant breeding of fine wool sheep.

Key words: Aohan fine wool sheep, ancestral-like coarse, lncRNA, mRNA, bioinformatics

中图分类号:

S826.2

罗睿杰, 王建魁, 曹素英. 敖汉细毛羊的粗毛返祖性状相关lncRNA-mRNA的联合测序分析[J]. 畜牧兽医学报, 2025, 56(6): 2685-2700.

LUO Ruijie, WANG Jiankui, CAO Suying. Integrated Sequencing Analysis of lncRNA and mRNA Related to Ancestral-like Coarse in Aohan Fine Wool Sheep[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(6): 2685-2700.

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标记 Code	软件中的描述 Description	可能的lncRNA类型 Potential lncRNA type
=	Complete match of intron chain
c	Contained
j	Potentially novel isoform (fragment): at least one splice junction is shared with a reference transcript
e	Single exon transfrag overlapping a reference exon and at least 10 bp of a reference intron, indicating a possible pre-mRNA fragment
i	A transfrag falling entirely within a reference intron	intronic lncRNA^[31]
o	Generic exonic overlap with a reference intron	sense lncRNA^[32]
p	Possible polymerase run-on fragment (within 2 kb of a reference transcript)
r	Repeat. Currently determined by looking at the soft-masked reference sequence and applied to transcripts where at least 50% of the bases are lower case
u	Unknown, intergenic transcript	intergenic lncRNA^[33]
x	Exonic overlap with reference on the opposite strand	anti-sense lncRNA^[34]
s	An intron of the transfrag overlaps a reference intron on the opposite strand (likely due to read mapping errors)
.	Tracking file only, indicates multiple classifications

样本 Sample	原始序列 Raw reads	过滤后序列 Clean reads	Q30/%	GC含量/% GC content
15180	90 832 836	89 431 798	97.06	50.00
15188	93 026 342	91 784 550	97.25	49.00
15190	92 599 986	91 039 240	96.85	49.50
15264	95 508 670	94 033 082	97.05	49.00
15268	88 211 386	86 928 822	97.17	48.50
15270	90 455 404	89 110 872	97.18	48.50

样本 Sample	过滤后序列总数 Total clean reads	比对到参考基因组上的Reads数 Total mapped reads	在参考基因组上比对到多个位置的Reads数 Multiple mapped reads	在参考基因组上比对到唯一位置的Reads数 Uniquely mapped reads
15180	89 431 798	76 136 143 (85.13%)	7 542 762 (8.43%)	68 593 381 (76.70%)
15188	91 784 550	77 309 464 (84.23%)	6 219 587 (6.78%)	71 089 877 (77.45%)
15190	91 039 240	75 357 994 (82.78%)	5 924 476 (6.51%)	69 433 518 (76.27%)
15264	94 033 082	79 377 054 (84.41%)	6 917 846 (7.36%)	72 459 208 (77.06%)
15268	86 928 822	71 416 612 (82.16%)	5 356 182 (6.16%)	66 060 430 (75.99%)
15270	89 110 872	75 382 070 (84.59%)	6 598 558 (7.40%)	68 783 512 (77.19%)

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