牦牛体外成熟卵母细胞差异转录组学研究  

Differential transcriptome analysis of yak oocytes in vitro maturation

作　　者：兰道亮 熊显荣[2] 陈亚冰[2] 泽让东科 艾鷖 李键 LAN DaoLiang XIONG XianRong CHENG YaBing MIPAM Tserang-donko AI Yi LI Jian(Institute of Tibetan Plateau Research, Southwest University for Nationalities, Chengdu 610041, China College of Life Science and Technology, Southwest University for Nationalities, Chengdu 610041, China)

机构地区：[1]西南民族大学青藏高原研究院,成都610041 [2]西南民族大学生命科学与技术学院,成都610041

出　　处：《中国科学：生命科学》2017年第10期1099-1112,共14页Scientia Sinica(Vitae)

基　　金：西南民族大学中央高校基本科研业务费专项资金(批准号:2015NZYTD01)资助

摘　　要：卵母细胞体外成熟环节是现代繁殖育种技术的基础环节,体外成熟的MⅡ期卵母细胞的质量直接影响了卵母细胞的后续受精及受精卵卵裂.由于卵母细胞的发育成熟是一个涉及大量基因转录调控的复杂过程.因此,转录组学研究是了解卵母细胞发生发育的关键.本研究以高海拔地区的特有牛属动物牦牛(Bos grunniens)卵母细胞作为研究对象,应用RNA-seq技术对牦牛GV期卵母细胞及体外成熟MⅡ期卵母进行转录组学测序及比对分析.经深度测序后,各获得一个包含51380686条过滤测序序列(cleanreads),4624261740碱基(bp)的GV期牦牛卵母细胞测序文库和一个包含50303412条过滤测序序列(clean reads),4527307080碱基(bp)的MⅡ期牦牛卵母细胞测序文库.基因覆盖率统计表明,GV期和MⅡ期文库中分别有16719条和16339条牦牛基因得到转录.通过比较分析MⅡ和GV期转录组数据,共筛选出4767个差异表达基因,其中1418个基因表达量上调和3349个基因表达量下调.GO功能分类注释显示,差异基因与"代谢过程"等生物学过程存在密切关联,同时结合分子功能相关类别等分子事件表现活跃.按表达量上调和下调分别对差异基因进行KEGG通路互作分析,结果表明,凋亡通路、内吞通路及代谢通路是上调的关键调控通路,而代谢通路、丙酮酸代谢、黏着斑及氧化磷酸化是下调的关键调控通路,其中代谢通路在上调及下调通路互作中均起关键调控作用.进一步对代谢通路进行分析,结果表明,组氨酸代谢、甘氨酸、丝氨酸和苏氨酸代谢、乙醛酸和二羧酸代谢及谷胱甘肽代谢是整个代谢通路的关键调控节点。此外,本研究发现,在牦牛卵母细胞体外成熟过程中,与免疫相关的白细胞介素家族及干扰素家族基因发生了显著差异表达.该研究结果为进一步解析牦牛卵母细胞发育的分子机制及全面了解牦牛繁殖的特异性提供了基础.Sound development and maturation of oocytes serve as important indicators in evaluating female fertility. Development and maturation of oocytes are complex processes involving transcriptional regulation of a large number of genes.Transcriptome studies are crucial in understanding such processes. In this study, oocytes obtained from yak—a unique bovine found in high-altitude areas—were used as research subject. RNA-seq was applied in comparative transcriptome analysis of yak oocytes, which were arrested at the germinal vesicle（GV） stage and metaphase Ⅱ（MⅡ）stage after in vitro maturation（IVM）. After deep sequencing, 51380686 clean reads totaling 4624261740 bp were obtained from a GV yak oocyte library, whereas 50303412 clean reads with a total of 4527307080 bp were obtained from a MⅡ yak oocyte library. Gene coverage statistics showed that 16719 and 16339 yak genes were mapped in GV and MⅡ libraries, respectively. Comparative transcriptome analysis of oocytes at MⅡ and GV stages revealed 4767 differentially expressed genes（DEGs）, including 1418 upregulated and 3349 downregulated genes. Gene Ontology（GO） functional classification analysis showed that these DEGs are closely related to ＂metabolism＂ and other biological processes; moreover, molecular events related to molecular binding function are active. The interaction analysis of Kyoto Encyclopedia of Genes and Genome（KEGG） pathways involved DEGs were conducted according to two parts of upregulation and downregulation respectively. Results of analysis of upregulated DEGs revealed that apoptosis,endocytosis, and metabolic pathways are key regulatory pathways. By contrast, downregulated genes are involved in metabolic pathways, pyruvate metabolism, focal adhesion, and oxidative phosphorylation. Therefore, metabolic pathways play critical regulatory roles in upregulation and downregulation of pathways. Further analysis showed that histidine, glycine, serine, and threonine metabolism; glyoxylic acid and dicarboxylic acid metabo

关 键 词：牦牛 卵母细胞 GV期 MⅡ期 体外成熟 RNA-SEQ 转录组 分子机制 

分 类 号：S823.85[农业科学—畜牧学]