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银鲫生殖细胞标记基因Dazl的特征及其与Vasa在
Alternative TitleStudies on characterization of germ cell marker gene Dazl and its dynamic expression with Vasa in germ cell development of gibel carp
彭金霞
Subtype博士
Thesis Advisor桂建芳
2009-05-30
Degree Grantor中国科学院水生生物研究所
Place of Conferral水生生物研究所
Keyword性别分化 幼鱼雌雄同体 银鲫 共线性 雌核发育 原始生殖细胞 生殖质 Metro途径
Abstract克隆鉴定了生殖细胞标记基因Dazl的同源基因CagDazl。发现各物种特别是鱼类的Dazl基因具有保守的基因组结构,并且DAZ家族基因与其附近的一批基因具有保守的共线性关系。制备了CagDazl特异的多克隆抗体,通过RT-PCR和Western blot分析,证明CagDazl特异表达于性腺中。追踪了CagDazl基因在雌核发育银鲫生殖细胞的整个生命周期中的动态变化: CagDazl RNA和蛋白都特异表达于卵巢生殖细胞并且在卵子发生中的各期卵母细胞中呈相同的变化动态;CagDazl蛋白和线粒体的共定位揭示在银鲫中存在METRO途径,而CagDazl RNA和蛋白是在卵子发生过程中经METRO途径迁移的生殖质成分。 在成熟卵子中,CagDazl 蛋白集中在植物极的薄层中;当雌核发育开始,CagDazl 蛋白从植物极经卵黄间胞质流迁移到动物极分裂球;至桑椹胚期,CagDazl 蛋白全部迁移到分裂球中并呈均匀分布;随着原始生殖细胞的形成和迁移,CagDazl 蛋白集中到PGC中并维持其在生殖细胞的特异表达;CagDazl 蛋白在幼鱼性腺的卵原细胞和早期卵母细胞中丰富表达。因此,CagDazl可以作为标记基因用于雌核发育银鲫生殖细胞和生殖质的相关研究,为银鲫生殖细胞特化、生殖细胞发育相关研究奠定了基础。 以CagVasa蛋白作为生殖细胞的标记分子,分析银鲫性别分化发现:和斑马鱼中情况相似,银鲫雄鱼性腺分化经历幼鱼雌雄同体阶段,并且雌鱼分化早于雄鱼。
Other AbstractTo investigate germline development and germ plasm specification, we identified a germ cell marker gene Dazl homolog (CagDazl) from gynogenetic gibel carp (Carassius auratus gibelio). Its cDNA sequence and BAC clone sequence analyses revealed the genomic organization conservation and conserved synteny of the Dazl family members and their neighborhood genes among vertebrates, especially in fish. Moreover, a polyclonal antibody specific to CagDazl was produced and used to examine its expression and distribution throughout germline development at protein level. Firstly, gonad-specific expression pattern of CagDazl was confirmed in adult tissues by RT-PCR and Western blot. And, in situ hybridization and immunofluorescence localization demonstrated its specific expression in germ cells, and both its transcript and protein were localized to germ plasm. Then, co-localization of CagDazl and mitochondrial cloud (MC) was found, confirming that CagDazl transcript and its protein are germ plasm component and move via METRO pathway during oogenesis. During embryogenesis, vegetal cortex localized CagDazl protein move toward blastomere in cytoplasm stream before 1k-cell stage and PGC specific expression was detected in 10-somite stage. Afterward, CagDazl protein was restricted to germ cells. The CagDazl is abundant and continuous throughout germline development and germ cell specification including PGC formation, oogonium differentiation, oocyte development, and embryogenesis, and the dynamic distribution occurs at different development stages.Therefore, CagDazl is a useful and specific marker for tracing germ plasm and germ cell development in the gynogenetic gibel carp. We traced the sex differentiation process in both gynogenetic and sexual reproduction population by immunofluorescence localization using Vasa protein as germ cell marker. The results revealed that, like the situation in zebrafish, the differentiation of gibel carp male gonad should undergo an intersex phase in juvenile fish, and the ovary differentiation is earlier than testis.
Pages121
Language中文
Document Type学位论文
Identifierhttp://ir.ihb.ac.cn/handle/342005/12416
Collection学位论文
Recommended Citation
GB/T 7714
彭金霞. 银鲫生殖细胞标记基因Dazl的特征及其与Vasa在[D]. 水生生物研究所. 中国科学院水生生物研究所,2009.
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