多倍体银鲫nanos2等位多态性、共线性和表达模式分析  

ALLELIC DIVERSIFICATION,SYNTENIC ALIGNMENT AND EXPRESSION PATTERNS OF NANOS2 IN POLYPLOID GIBEL CARP

作　　者：张琴琴 周莉[1,2] 李志 甘瑞海[1,2] 俞兆曦 桂建芳 汪洋[1,2] ZHANG Qin-Qin;ZHOU Li;LI Zhi;GAN Rui-Hai;YU Zhao-Xi;GUI Jian-Fang;WANG Yang(State Key Laboratory of Freshwater Ecology and Biotechnology,Innovation Academy for Seed Design,Institute of Hydrobiology,Chinese Academy of Sciences,Wuhan 430072,China;University of Chinese Academy of Sciences,Beijing 100049,China;Ningxia Fisheries Research Institute,Yinchuan 750001,China)

机构地区：[1]中国科学院水生生物研究所种子创新研究院,淡水生态与生物技术国家重点实验室,武汉430072 [2]中国科学院大学,北京100049 [3]宁夏回族自治区水产研究所(有限公司),银川750001

出　　处：《水生生物学报》2020年第5期1087-1096,共10页Acta Hydrobiologica Sinica

基　　金：国家重点研发计划蓝色粮仓科技创新(2018YFD0900204);国家大宗淡水鱼产业技术体系(CARS-45-07)资助。

摘　　要：为研究生殖干细胞(Germline stem cells, GSCs)的标记基因nanos2的功能,在银鲫(Carassius gibelio)中克隆了两个nanos2的同源基因(Homologs),将其命名为Cgnanos2a和Cgnanos2b,等位、系统进化树和共线性分析表明,在银鲫进化历程中发生了额外的两轮多倍化事件,是一个异源六倍体。qPCR分析表明Cgnanos2a和Cgnanos2b在5月龄银鲫精巢中的表达水平最高,其次是卵巢;对在孵化后25—190d的卵巢中的表达动态分析表明,孵化后25d的卵巢中的Cgnanos2a和Cgnanos2b转录水平最高,随后表达水平急剧下降;并且Cgnanos2a和Cgnanos2b呈现出偏向表达的特征。切片RNA原位杂交实验结果表明, Cgnanos2a和Cgnanos2b均特异地在银鲫卵巢邻近生殖上皮的胞囊(Cyst)中一类直径小于20μm的细胞中表达, Cgnanos2a在精巢精小囊边缘一类单个或两个紧紧相邻的细胞中高表达,推测是银鲫的GSCs。此外,还可在精原细胞和初级精母细胞中检测到Cgnanos2a和Cgnanos2b的转录本。研究通过对nanos2阳性细胞的追踪描绘了银鲫卵巢早期胞囊的发育过程,为后续分离银鲫GSCs奠定了基础;同时对银鲫nanos2两个歧化的部分同源基因的序列特征、进化及表达特征分析,为研究鱼类多次多倍化事件后重复基因的进化提供了一个典型例子。Germline stem cells(GSCs)have the self-renewal ability to continuously give rise to gametes throughout their life.Owing to its unique reproductive and sex determination modes,polyploid gibel carp(Carassius gibelio)has become a promising model organism for reproductive and developmental genetics in vertebrates.Although several germ cell marker genes,such as vasa,dazl and dnd,have been studied in gibel carp,GSC genes has not been identified.Here we identified two divergent nanos2 homeologs(named Cgnanos2a and Cgnanos2b),each of which has three alleles.Phylogenetic and synteny analyses indicated that two rounds of polyploidization occurred during allo-autohexaploid gibel carp evolution.Cgnanos2a and Cgnanos2b were most expressed in testis,followed by ovary.They also highly expressed in ovary at 25 dph,and then decreased sharply.The expression levels of Cgnanos2a in testis and ovaries from 25 dph to 60 dph were higher than those of Cgnanos2b(P<0.05),indicating significant expression bias between Cgnanos2a and Cgnanos2b.Finally,Cgnanos2a and Cgnanos2b specifically expressed in a subset of small germ cells with a diameter of<20μm in the ovary,and abundant Cgnanos2a transcripts were detected in isolated cells or in doublets of testis.Considering these characteristics,Cgnanos2a and Cgnanos2b might be the markers of gibel carp GSCs.In addition,their transcripts were also detected in spermatogonia and spermatocytes.Therefore,we described the development of early ovarian cysts with nanos2 positive cells,which will help to isolate gibel carp GSCs.The sequence characteristics,evolution and expression patterns of two divergent nanos2 genes in polyploid gibel carp provide a paradigm for the evolution of duplicated genes in recurrent polyploidy vertebrates.

关 键 词：生殖干细胞 银鲫 六倍体 nanos2 偏向表达 

分 类 号：Q344.1[生物学—遗传学]