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RNA and Protein Detection by Single-Molecule Fluorescent in Situ Hybridization (smFISH) Combined with Immunofluorescence in the Budding Yeast S. cerevisiae

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Fluorescence In Situ Hybridization (FISH)

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2784))

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Abstract

The inherent stochastic processes governing gene expression give rise to heterogeneity across individual cells, highlighting the importance of single-cell studies. The emergence of single-molecule fluorescent in situ hybridization (smFISH) enabled gene expression analysis at the single-cell level while including the spatial dimension through the visualization and quantification of mRNAs in intact fixed cells. By combining smFISH with immunofluorescence (IF), a comprehensive approach takes shape facilitating the study of mRNAs and proteins to correlate gene expression profiles to different cellular states. This chapter serves as a comprehensive guide to a smFISH-IF protocol optimized for gene expression analysis in the budding yeast S. cerevisiae. We utilize smFISH to visualize the mRNA localization pattern of the CLB2 cyclin over the course of the cell cycle inferred by alpha-tubulin IF.

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Authors and Affiliations

  1. Department of Cell Biology, Albert Einstein College of Medicine, Bronx, NY, USA

    Anna Maekiniemi & Robert H. Singer

Authors
  1. Anna Maekiniemi
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  2. Robert H. Singer
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Corresponding author

Correspondence to Robert H. Singer .

Editor information

Editors and Affiliations

  1. Department of Molecular Genetics, Weizmann Institute of Science, Rehovot, Israel

    Gal Haimovich

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© 2024 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Maekiniemi, A., Singer, R.H. (2024). RNA and Protein Detection by Single-Molecule Fluorescent in Situ Hybridization (smFISH) Combined with Immunofluorescence in the Budding Yeast S. cerevisiae. In: Haimovich, G. (eds) Fluorescence In Situ Hybridization (FISH). Methods in Molecular Biology, vol 2784. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3766-1_3

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  • DOI: https://doi.org/10.1007/978-1-0716-3766-1_3

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3765-4

  • Online ISBN: 978-1-0716-3766-1

  • eBook Packages: Springer Protocols

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