Indirect Immunofluorescence of Proteins in Oogenic Germ Cells of Caenorhabditis elegans.
Indirect Immunofluorescence of Proteins in Oogenic Germ Cells of Caenorhabditis elegans.
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秀丽隐杆线虫卵子生殖细胞中蛋白质的间接免疫荧光。
DOI：
10.1007/978-1-4939-3795-0_2
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发表时间：
2016
期刊：
影响因子：
--
通讯作者：
Schedl,Tim
中科院分区：
文献类型：
--
作者：
Brenner,JohnL;Schedl,Tim
Formation of full-grown oocytes requires the control and coordination of a number of processes (e.g., oocyte growth) through multiple stages, where disruption at any one step can result in infertility. Numerous proteins are required for the regulation and execution of the various oogenic processes as well as functioning as maternal products needed for embryogenesis. Immunofluorescence microscopy combined with staining using antibodies against specific proteins, or their posttranslationally modified forms, is a standard approach to determine the temporal and spatial location of gene products that function in oocyte development. The simple linear organization of the germline in the model organismCaenorhabditis elegansallows easy correlation of protein localization and germ cell developmental stage, thus aiding in our understanding of protein function during gametogenesis. Here we outline co-immunofluorescence staining for two major regulators ofC. elegansgermline development, the translational repressor GLD-1 and activated form of MPK-1 (dpMPK-1) ERK MAP kinase in dissected gonads from adultC. elegans. Worms are first dissected and the extruded gonads are fixed and permeabilized before being bathed in primary antibodies against GLD-1 and dpMPK-1. Secondary antibodies conjugated to fluorophore dyes and that target the IgG domains of the primary antibody reagents are then used to provide a fluorescent signal that corresponds to the position of GLD-1 and dpMPK-1. The outlined procedure is amenable to many other proteins expressed inC. elegansgerm cells.
