A simple and rapid method for combining fluorescent in situ RNA hybridization (FISH) and immunofluorescence in the C. elegans germline

摘要

class="kwd-title">Keywords: Fluorescent in situ hybridization (FISH), Immunofluorescence, C. elegans, Germline class="head no_bottom_margin" id="abs0010title">AbstractImaging of RNAs and proteins in specific tissues has opened ample avenues to understand gene expression during development. Recently, a fluorescent in situ RNA hybridization (FISH) method has been developed to analyze the spatio-temporal expression patterns of endogenous mRNAs. However, combining FISH with immunofluorescence is challenging as the reaction conditions for the two procedures conflict in multiple ways. In this report, we developed a simple and rapid method to detect both RNAs and associated proteins with better preservation of the fine structure in the C. elegans germline. This method will provide new tools for in vivo imaging of RNAs and their associated proteins in the same germline, which also enables simultaneous visualization of RNA/protein complex at the cellular level in vivo. class="first-line-outdent">

• Developing a simple and rapid FISH method with better preservation of the fine structure.

• Combining FISH with immunofluorescence in C. elegans germline.

• Labeling extruded gonads, instead of the whole worms, to prevent non-specific somatic autofluorescence.