Production of zebrafish offspring from cultured female germline stem cells.
Production of zebrafish offspring from cultured female germline stem cells.
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DOI：
10.1371/journal.pone.0062660
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Collodi P
中科院分区：
文献类型：
--
作者：
Wong TT;Tesfamichael A;Collodi P
在生殖细胞特异性启动子ziwi [Tg(ziwi: Neo);Tg(ziwi:DsRed)]的控制下，通过转基因鱼种系产生了斑马鱼雌性生殖系干细胞（FGSC）。通过G418筛选建立了均匀的FGSC培养物，并在6周以上的时间内持续表达ziwi，同时表达生殖细胞标记物nanos3、dnd、dazl和vasa。细胞培养系统的一个关键组成部分是使用从转基因鱼系[Tg(gsdf:neo)]的卵巢启动的饲养细胞系，该细胞系表达由斑马鱼性腺体细胞衍生因子（gsdf）启动子控制的neo。在G418中选择饲养细胞系，通过工程改造表达斑马鱼白血病抑制因子（liff）、碱性成纤维细胞生长因子（Fgf2）和胶质细胞系衍生神经营养因子（Gdnf）。这些因子在培养中显著提高了FGSC的生长、存活和种系能力。细胞移植实验结果表明，培养的FGSCs能够成功地定植在不育受体鱼的性腺上并产生功能配子。在注射了培养的FGSCs的受体鱼中，高达20%的存活鱼是可育的，并且在至少6个月的时间里产生了多批正常后代。FGSC培养物将为斑马鱼生殖细胞生长和分化的研究提供一个体外系统，它们在移植后的高频率生殖系传播可以形成干细胞介导的基因转移和斑马鱼基因组操作策略的基础。
Zebrafish female germline stem cell (FGSC) cultures were generated from a transgenic line of fish that expresses Neo and DsRed under the control of the germ cell specific promoter, ziwi [Tg(ziwi:neo);Tg(ziwi:DsRed)]. Homogeneous FGSC cultures were established by G418 selection and continued to express ziwi for more than 6 weeks along with the germ cell markers nanos3, dnd, dazl and vasa. A key component of the cell culture system was the use of a feeder cell line that was initiated from ovaries of a transgenic line of fish [Tg(gsdf:neo)] that expresses Neo controlled by the zebrafish gonadal soma derived factor (gsdf) promoter. The feeder cell line was selected in G418 and engineered to express zebrafish leukemia inhibitory factor (Lif), basic fibroblast growth factor (Fgf2) and glial-cell-line derived neurotrophic factor (Gdnf). These factors were shown to significantly enhance FGSC growth, survival and germline competency in culture. Results from cell transplantation experiments revealed that the cultured FGSCs were able to successfully colonize the gonad of sterile recipient fish and generate functional gametes. Up to 20% of surviving recipient fish that were injected with the cultured FGSCs were fertile and generated multiple batches of normal offspring for at least 6 months. The FGSC cultures will provide an in vitro system for studies of zebrafish germ cell growth and differentiation and their high frequency of germline transmission following transplantation could form the basis of a stem cell-mediated strategy for gene transfer and manipulation of the zebrafish genome.
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影响因子：
1.2
作者：
Shibata, Yasushi;Paul-Prasanth, Bindhu;Nagahama, Yoshitaka
通讯作者：
Nagahama, Yoshitaka
影响因子：
2.5
作者：
Leu, David H.;Draper, Bruce W.
通讯作者：
Draper, Bruce W.
影响因子：
2.7
作者：
Hofmann, MC;Braydich-Stolle, L;Dym, M
通讯作者：
Dym, M
影响因子：
64.5
作者：
MATSUI, Y;ZSEBO, K;HOGAN, BLM
通讯作者：
HOGAN, BLM
DOI：
10.1073/pnas.0308668101
发表时间：
2004-05-25
影响因子：
11.1
作者：
Hong, YH;Liu, TM;Gui, JF
通讯作者：
Gui, JF