Manipulation and Imaging of Kryptolebias marmoratus Embryos
Manipulation and Imaging of Kryptolebias marmoratus Embryos
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DOI：
10.1093/icb/ics076
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发表时间：
2012-12-01
影响因子：
2.6
通讯作者：
Kudoh, Tetsuhiro
中科院分区：
文献类型：
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作者：
Mourabit, Sulayman;Kudoh, Tetsuhiro
自花红树林鳉，Kryptolebias marmoratus，是一个即将到来的模式物种的一系列生物学科。为了在发育生物学领域进一步建立这种模型，我们研究了几种用于胚胎操作和成像的技术，这些技术可以用于一系列实验设计。这些方法可分为两类：处理有绒毛膜和无绒毛膜的胚胎。仍然封闭在绒毛膜中的胚胎可以使用琼脂糖床或甲基纤维素系统进行操作，将它们固定在适当的位置，并允许它们旋转到更特定的角度和位置。使用这些方法，我们证明显微注射胚胎和荧光卵黄合胞体核（YSN）的监测，使用立体和复合显微镜。对于使用复合显微镜的更高放大率成像以及延时分析，将胚胎脱氯并包埋在低熔点琼脂糖中。为了证明这种包埋技术，我们进一步研究了荧光YSN，并分析了K。marmoratus胚胎后者被观察到为研究胚胎发育期间细胞的行为和形态提供了一个极好的成像平台，用于各种类型的细胞。我们的数据表明，K。marmoratus是发育生物学研究的一个极好的模式物种，因为胚胎操作和成像的方法学方法是有效和容易获得的。
The self-fertilizing mangrove killifish, Kryptolebias marmoratus, is an upcoming model species for a range of biological disciplines. To further establish this model in the field of developmental biology, we examined several techniques for embryonic manipulation and for imaging that can be used in an array of experimental designs. These methodological approaches can be divided into two categories: handling of embryos with and without their chorionic membrane. Embryos still enclosed in their chorion can be manipulated using an agarose bed or a methyl cellulose system, holding them in place and allowing their rotation to more specific angles and positions. Using these methods, we demonstrate microinjection of embryos and monitoring of fluorescent yolk syncytial nuclei (YSN) using both stereo and compound microscopes. For higher magnification imaging using compound microscopes as well as time-lapse analyses, embryos were dechorionated and embedded in low-melting-point agarose. To demonstrate this embedding technique, we further examined fluorescent YSN and also analyzed the yolk surface of K. marmoratus embryos. The latter was observed to provide an excellent imaging platform for study of the behavior and morphology of cells during embryonic development, for various types of cells. Our data demonstrate that K. marmoratus is an excellent model species for research in developmental biology, as methodological approaches for the manipulation and imaging of embryos are efficient and readily available.