Kemp Hilary A, Carmany-Rampey Amanda, Moens Cecilia
HHMI and Division of Basic Sciences, Fred Hutchinson Cancer Research Center.
J Vis Exp. 2009 Jul 17(29):1394. doi: 10.3791/1394.
One of the most powerful tools used to gain insight into complex developmental processes is the analysis of chimeric embryos. A chimera is defined as an organism that contains cells from more than one animal; mosaics are one type of chimera in which cells from more than one genotype are mixed, usually wild-type and mutant. In the zebrafish, chimeras can be readily made by transplantation of cells from a donor embryo into a host embryo at the appropriate embryonic stage. Labeled donor cells are generated by injection of a lineage marker, such as a fluorescent dye, into the one-cell stage embryo. Labeled donor cells are removed from donor embryos and introduced into unlabeled host embryos using an oil-controlled glass pipette mounted on either a compound or dissecting microscope. Donor cells can in some cases be targeted to a specific region or tissue of the developing blastula or gastrula stage host embryo by choosing a transplantation site in the host embryo based on well-established fate maps.
用于深入了解复杂发育过程的最强大工具之一是嵌合胚胎分析。嵌合体被定义为包含来自不止一种动物细胞的生物体；镶嵌体是嵌合体的一种类型，其中来自不止一种基因型的细胞混合在一起，通常是野生型和突变型。在斑马鱼中，可以通过在适当的胚胎阶段将供体胚胎的细胞移植到宿主胚胎中来轻松制造嵌合体。通过将谱系标记物（如荧光染料）注射到单细胞阶段的胚胎中产生标记的供体细胞。使用安装在复式显微镜或解剖显微镜上的油控玻璃移液管从供体胚胎中取出标记的供体细胞，并将其引入未标记的宿主胚胎中。在某些情况下，通过根据已建立的命运图谱在宿主胚胎中选择移植部位，可以将供体细胞靶向发育中的囊胚或原肠胚阶段宿主胚胎的特定区域或组织。
