Sep 15, 2019

Multiplexed RNAscope and immunofluorescence on whole-mount skeletal myofibers and their associated stem cells

Development
Allison P Kann, Robert S Krauss

Abstract

Skeletal muscle myofibers are large syncytial cells comprising hundreds of myonuclei, and in situ hybridization experiments have reported a range of transcript localization patterns within them. Although some transcripts are uniformly distributed throughout myofibers, proximity to specialized regions can affect the programming of myonuclei and functional compartmentalization of transcripts. Established techniques are limited by a lack of both sensitivity and spatial resolution, restricting the ability to identify different patterns of gene expression. In this study, we adapted RNAscope fluorescent in situ hybridization technology for use on whole-mount mouse primary myofibers, a preparation that isolates single myofibers with their associated muscle stem cells remaining in their niche. This method can be combined with immunofluorescence, enabling an unparalleled ability to visualize and quantify transcripts and proteins across the length and depth of skeletal myofibers and their associated stem cells. Using this approach, we demonstrate a range of potential uses, including the visualization of specialized transcriptional programming within myofibers, tracking activation-induced transcriptional changes, quantification of stem ce...Continue Reading

  • References51
  • Citations1

Citations

Mentioned in this Paper

Fluorescent Dyes
Study
Patterns
Transcription, Genetic
Spatial Distribution
Muscle Satellite Cell
Gene Expression
Evaluation
Primary Endpoint
Isolate - Microorganism

Related Feeds

Adult Stem Cells

Adult stem cells reside in unique niches that provide vital cues for their survival, self-renewal, and differentiation. They hold great promise for use in tissue repair and regeneration as a novel therapeutic strategies. Here is the latest research.

Related Papers

Methods in Molecular Biology
M HargraveP Koopman
Methods in Molecular Biology
Youtaro ShibayamaMusa M Mhlanga
Developmental Dynamics : an Official Publication of the American Association of Anatomists
Stanley J NeufeldJohn Cobb
Methods in Molecular Biology
J de Almeida EnglerG Engler
© 2020 Meta ULC. All rights reserved