基于转录组测序挖掘奶牛乳脂代谢关键候选基因

doi:10.11843/j.issn.0366-6964.2022.10.015

摘要/Abstract

摘要： 旨在对高、低乳脂率奶牛乳腺上皮细胞（BMECs）转录组测序的mRNA表达谱数据进行深入分析，挖掘影响奶牛乳脂代谢的关键候选基因。本研究采用Illumina PE150方法对乳脂率具有极端差异的荷斯坦奶牛（高、低乳脂组各4头）的BMECs进行转录组测序，以P<0.05和|log₂FoldChange|≥1.5为阈值筛选差异表达基因，并利用KOBAS在线网址进行功能富集分析，最后通过实时荧光定量PCR （qRT-PCR）技术分析测序结果的准确性及乳脂代谢相关差异表达基因的组织表达谱。结果表明，在高、低乳脂组之间共发现578个差异表达基因，包括332个上调差异表达基因，246个下调差异表达基因。功能富集分析共确定了包含生物学过程（BP）、细胞组分（CC）和分子功能（MF）的366个显著富集的GO条目（P<0.05），其中与脂代谢密切相关的GO条目有长链脂肪酸的运输、脂肪细胞分化的正向调节、乳腺肺泡发育、花生四烯酸的结合等。差异表达基因显著富集到47条KEGG通路（P<0.05），参与脂代谢的通路有15条，分别为脂肪细胞内脂解的调节、磷脂酶D信号通路、河马信号通路等，其中ID2、PRKKA2、FABP4和ADCY5为可能调控乳脂代谢的关键候选基因。组织表达谱分析发现，FABP4在乳腺组织中的表达水平相对最高，ID2、PRKKA2和ADCY5相对于其它组织在乳腺中的表达也均处于较高水平。本研究筛选得到了4个影响奶牛乳脂代谢的重要候选基因，为今后奶牛乳脂代谢的分子调控机制研究提供了重要的理论依据。

关键词: 荷斯坦奶牛, 转录组, 差异表达基因, 乳脂, 组织表达谱

Abstract: The aim of this study was to conduct in-depth analysis of mRNA expression profile data from transcriptome sequencing in bovine mammary epithelial cells (BMECs) of dairy cows with high and low milk fat percentage, and to identify key candidate genes affecting milk fat metabolism in dairy cows. Transcriptome sequencing of BMECs from Holstein dairy cows (4 cows in high and low milk fat percentage groups,respectively) was performed by Illumina PE150 method, and the differentially expressed genes were screened with P<0.05 and|log₂FoldChange| ≥ 1.5. KOBAS website was used for functional enrichment analysis. Finally, the accuracy of the sequencing results and the tissue expression profiles of differentially expressed genes related to milk fat metabolism were analyzed by real-time quantitative PCR (qRT-PCR) technology. The results show that there were 578 differentially expressed genes between the high and low milk fat groups, including 332 differentially expressed up-regulated genes, 246 differentially expressed down-regulated genes. Functional enrichment analysis identified 366 significantly enriched GO items including biological process (BP), cellular component (CC) and molecular function (MF) (P<0.05). Among them, GO items closely related to lipid metabolism included long-chain fatty acid transport, positive regulation of adipocyte differentiation, mammary alveolar development, arachidonic acid binding, etc. The differential genes were significantly enriched in 47 KEGG pathways (P<0.05), and there were 15 pathways involved in lipid metabolism, including regulation of lipolysis in adipocytes, phospholipase D signaling pathway, Hippo signaling pathway, etc. Among them, ID2, PRKAA2, FABP4 and ADCY5 were the key candidate genes regulating milk lipid metabolism. Tissue expression profile analysis showed that the expression level of FABP4 was the highest in mammary gland tissue, and the expression levels of ID2, PRKAA2 and ADCY5 were also at a high level compared with other tissues. In this study, 4 important candidate genes affecting milk fat metabolism in dairy cows were screened, which provided an important theoretical basis for studying on the molecular regulation mechanism of milk fat metabolism in dairy cows in the future.

Key words: Holstein dairy cattle, transcriptome, differentially expressed genes, milk fat, tissue expression profiling

中图分类号:

王川川, 母童, 冯小芳, 禹保军, 张娟, 顾亚玲. 基于转录组测序挖掘奶牛乳脂代谢关键候选基因[J]. 畜牧兽医学报, 2022, 53(10): 3421-3433.

WANG Chuanchuan, MU Tong, FENG Xiaofang, YU Baojun, ZHANG Juan, GU Yaling. Identification of Key Candidate Genes for Milk Fat Metabolism in Dairy Cows Based on Transcriptome Sequencing[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(10): 3421-3433.

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