In vivo Live Imaging of Calcium Waves and Other Cellular Processes during Fertilization in Caenorhabditis elegans
In vivo Live Imaging of Calcium Waves and Other Cellular Processes during Fertilization in Caenorhabditis elegans
复制标题
DOI：
10.21769/bioprotoc.2205
复制
发表时间：
2017-04-05
期刊：
影响因子：
0.8
通讯作者：
Onami, Shuichi
中科院分区：
文献类型：
--
作者：
Takayama, Jun;Fujita, Masashi;Onami, Shuichi
受精钙波是动物发育的保守触发器；然而，由于体内受精成像的困难，这些波的遗传分析受到限制。在这里，我们描述了在遗传动物模型秀丽隐杆线虫体内受精过程中图像钙动力学的协议。该方案包括化学钙指示剂的种系显微注射，蠕虫固定化，实时成像和图像处理，量化排卵期间在视野中移动的卵母细胞区域的钙荧光。该成像方案也可用于成像秀丽隐杆线虫体内受精过程中的其他细胞过程，如膜融合和细胞骨架动力学。
Fertilization calcium waves are a conserved trigger for animal development; however, genetic analysis of these waves has been limited due to the difficulty of imaging in vivo fertilization. Here we describe a protocol to image calcium dynamics during in vivo fertilization in the genetic animal model Caenorhabditis elegans. This protocol consists of germline microinjection of a chemical calcium indicator, worm immobilization, live imaging, and image processing that quantifies the calcium fluorescence in the oocyte region moving in the field-of-view during ovulation. This imaging protocol can also be used to image other cellular processes during in vivo fertilization in C. elegans, such as membrane fusion and cytoskeletal dynamics.