Imaging retinal progenitor lineages in developing zebrafish embryos.
Imaging retinal progenitor lineages in developing zebrafish embryos.
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DOI：
10.1101/pdb.prot073544
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发表时间：
2013-03-01
影响因子：
--
通讯作者：
Poggi, Lucia
中科院分区：
文献类型：
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作者：
Jusuf, Patricia;Harris, William A;Poggi, Lucia
在本协议中，我们描述了如何制作和分析体内斑马鱼胚胎视网膜谱系的四维（4D）电影。 4D 由共焦平面堆栈和一个时间维度重建的三个空间维度 (3D) 组成。我们的成像是在细胞特异性启动子控制下表达荧光蛋白的转基因细胞或在特定视网膜细胞祖细胞中瞬时表达此类报告基因的细胞上进行的。谱系追踪的一个重要方面是跟踪单个细胞经历多次细胞分裂、最终迁移和分化的能力。这可能意味着许多小时的 4D 成像，需要细胞保持健康并维持在适合正常发育的条件下。我们制作的最长的电影有 50 小时。通过分析这些电影，我们可以看到特定细胞的出生时间以及它的姐妹是谁，从而使我们能够在体内重建其视网膜谱系。
In this protocol, we describe how to make and analyze four dimensional (4D) movies of retinal lineage in the zebrafish embryo in vivo. 4D consists of three spatial dimensions (3D) reconstructed from stacks of confocal planes plus one time dimension. Our imaging is performed on transgenic cells that express fluorescent proteins under the control of cell-specific promoters or on cells that transiently express such reporters in specific retinal cell progenitors. An important aspect of lineage tracing is the ability to follow individual cells as they undergo multiple cell divisions, final migration, and differentiation. This may mean many hours of 4D imaging, requiring that cells be kept healthy and maintained under conditions suitable for normal development. The longest movies we have made are 50 h. By analyzing these movies, we can see when a specific cell was born and who its sister was, allowing us to reconstruct its retinal lineages in vivo.