黄河鲤 nanog 基因克隆及其表达特征研究
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摘要：
转录因子Nanog在干细胞多能性维持、胚胎早期发育等方面具有重要作用.利用PCR技术克隆黄河鲤nanog基因(Ccnanog),通过实时荧光定量PCR或半定量PCR分别对其在黄河鲤不同胚胎发育时期以及雌、雄成体不同组织中的表达特征进行了分析,并利用荧光原位杂交和免疫组织化学方法进一步检测了该基因在黄河鲤卵巢和精巢中的亚细胞定位.结果显示,黄河鲤nanog基因ORF为1158bp,编码385个氨基酸残基;Nanog蛋白包含保守的HOX结构域,与斑马鱼的序列相似性最高.在胚胎中,Ccnanog在未受精卵、早期胚胎的Ⅰ细胞期至原肠胚期高表达,在神经胚期表达量显著降低,之后在眼囊期至出膜期基本检测不出;在组织中,Ccnanog短片段在以性腺为首的多个组织中均有不同程度的表达,Ccnanog长片段仅在雌鱼卵巢和皮肤中表达,其中在雌鱼卵巢中的表达量最高,其组织表达具有一定的性别二态性;在性腺中进一步检测到,Ccnanog主要分布在卵巢的卵原细胞Ⅰ期、Ⅱ期和Ⅲ期初级卵母细胞的细胞质中,以及精巢的精原细胞、精母细胞和精子细胞中,且随着生殖细胞的发育其表达量逐渐下降.以上结果表明Ccnanog可能参与了黄河鲤早期胚胎发育过程与生殖细胞的发育,为后续功能研究提供了参考.
The transcription factorNanog plays an importantrole in maintenanceofpluripotencyofstem cells and early embryonic development. In this study, thenanog geneofthe Yellow Rivercarp(Ccnanog) was cloned byPCR technology. The expression characteristicsofCcnanog genein differentembryonicdevelopmentstagesand in differenttissuesoffemaleand male adultsoftheYellow Rivercarp wereanalyzed by real-timefluorescencequantitativePCR orsemi-quantitativePCR. The subcel- lularlocalization ofCcnanog gene in ovary and testisofthe Yellow Rivercarp werefurtherdetected byfluorescenceinsitu hy- bridization and immunohistochemistry. The results showed thatthe ORF ofthenanog geneofthe Yellow Rivercarp was 1158 bp, encoding385amino acid residues. Nanog protein contains a conserved HOX domain, which has the highestsequence simi- larity with zebrafish. In theembryo, Ccnanog washighlyexpressed in theunfertilized eggs, from the1-cellstageto thegastru- lastage, and the expression level significantly decreased in the neurula stage, then was basically undetectable from the optic capsule stage to the hatching stage. In tissues, shortsegments ofCcnanog are expressed to varying degrees in multiple tissues headed bythegonads, whilelong segmentsofCcnanog areonlyexpressed in theovaryand skin offemalefish, with thehighest expression levelin theovary, and theirtissueexpression hasa certain genderdimorphism. Itwasfurtherdetected in thegonads thatCcnanog was mainlydistributed in the oogonia, cytoplasm ofthe phaseI, IIand IIIprimary oocytes ofthe ovary, as well as in the spermatogonia, spermatocytes and sperm cells ofthe testis, and their expression gradually decreased with the devel- opmentoftheoocytesorspermatocytes. TheaboveresultssuggestthatCcnanog mayparticipatein theearlyembryosand germ cells developmentofthe Yellow River carp, providing a reference for the follow-up study ofits function.
作者：
于淼,刚会会,王方圆，李木子,王园，江红霞，王磊，张猛，乔志刚
Yu Miao, Gang Huihui, Wang Fangyuan, LiMuzi, Wang Yuan, Jiang Hongxia, WangLei, Zhang Meng, Qiao Zhigang
机构地区：
betway官方app 水产学院 ;水产动物疾病控制河南省工程实验室 ;河南省水产动物养殖工程技术研究中心
引用本文：
于淼,刚会会,王方圆等.黄河鲤nanog基因克隆及其表达特征研究[J].betway官方app 学报（自然科学版）,2024,52(4):63-70.
Yu Miao, Gang Huihui, Wang Fangyuan, et al.Cloning and expression characteristics of nanog gene in the Yellow Rivercarp[J] .Journalof Henan NormalUniversity(NaturalScience Edition) ,2024, 52(4) :63-70. DOI:10. 16366/j. cnki.1000-2367. 2023. 03. 29. 0003.
基金：
国家自然科学基金；河南省高等学校重点科研项目；河南省科技攻关项目;betway官方app 博士科研启动基金项目
关键词：
黄河鲤 ;nanog;基因表达 ;性腺
Yellow River carp; nanog; gene expression; gonad
分类号：
S917. 4
