Investigation of the Nodavirus lifecycle in vivo

描述（由申请人提供）： 鸡群屋病毒（FHV）是一种感染昆虫的无包膜二十面体RNA病毒。FHV已经在遗传学、生物药理学和结构上进行了彻底的研究，这使得它成为一个很好的系统，用于扩展这些研究以原位研究病毒的生命周期（病毒进入、拆卸、组装和细胞退出）。为了原位研究FHV的生命周期，将用双砷荧光团特异性标记含有四半胱氨酸标签的病毒外壳蛋白，所述双砷荧光团在荧光显微镜和电子显微镜中均可见，从而允许不同的图像相关联。此外，在光学脉冲追踪实验中，FHV将连续标记不同颜色的双砷荧光团，以区分较老和较年轻的外壳蛋白。为了以更高的分辨率研究细胞内FHV的空间关系，将使用断层扫描电子显微镜图像来产生病毒感染细胞的三维重建。

DESCRIPTION (provided by applicant): Flock House Virus (FHV) is a non-enveloped, icosahedral RNA virus that infects insects. FHV has been thoroughly studied genetically, biophysically, and structurally, which makes it an excellent system for extending these studies to investigate the virus lifecycle (virus entry, disassembly, assembly, and cellular exit) in situ. To study the FHV lifecycle in situ the virus coat protein containing the tetracysteine tag will be labeled specifically with biarsenical fluorophores, which are visible in both the fluorescent and electron microscopes allowing the different images to be correlated. Additionally, in optical pulse-chase experiments the FHV will be successively labeled with different colored biarsenical fluorophores to distinguish older and younger coat protein. To investigate at higher resolution the spatial relationship of FHV within cells, tomographic electron microscope images will be used to produce three-dimensional reconstructions of the virus infected cells.