Detection of mRNA Transfer Between Mammalian Cells in Coculture by Single-Molecule Fluorescent In Situ Hybridization (smFISH)

Gal Haimovich; Jeffrey E. Gerst

doi:10.1007/978-1-4939-9674-2_8

Detection of mRNA Transfer Between Mammalian Cells in Coculture by Single-Molecule Fluorescent In Situ Hybridization (smFISH)

Gal Haimovich, Jeffrey E. Gerst

Department of Molecular Genetics

Weizmann Institute of Science

Research output: Chapter in Book/Report/Conference proceeding › Chapter

Abstract

In eukaryotic cells, a small percentage of mRNA molecules can undergo transfer from one cell to another. mRNA transfer occurs primarily via membrane nanotubes, which are long thin protrusions that are produced by numerous cell types and can connect cells that can be up to hundreds of microns apart. Potentially, mRNAs might also transfer via extracellular vesicles (EVs). Here we describe a method to detect transferred mRNA in cocultures of two different cell types and to distinguish between nanotube- and EVs-mediated transfer. This method uses single molecule fluorescent in situ hybridization (smFISH) to provide an accurate and quantitative detection of transferred mRNA molecules and their subcellular localization. Following the guidelines presented here will allow the user to investigate mRNA transfer of most transcripts in any co-culture system. In addition, we present modifications that improve nanotube preservation during the smFISH procedure.

Original language	English
Title of host publication	Imaging Gene Expression
Subtitle of host publication	Methods and Protocols
Editors	Yaron Shav-Tal
Place of Publication	New York, NY
Publisher	Humana Press
Pages	109-129
Number of pages	21
ISBN (Electronic)	9781493996742
ISBN (Print)	9781493996735
DOIs	https://doi.org/10.1007/978-1-4939-9674-2_8
State	Published Online - 13 Aug 2019

Publication series

Name	Methods in Molecular Biology
Volume	2038
ISSN (Print)	1064-3745

All Science Journal Classification (ASJC) codes

Molecular Biology
Genetics

Access to Document

10.1007/978-1-4939-9674-2_8

Cite this

Haimovich, G., & Gerst, J. E. (2019). Detection of mRNA Transfer Between Mammalian Cells in Coculture by Single-Molecule Fluorescent In Situ Hybridization (smFISH). In Y. Shav-Tal (Ed.), Imaging Gene Expression: Methods and Protocols (pp. 109-129). (Methods in Molecular Biology; Vol. 2038). Humana Press. Advance online publication. https://doi.org/10.1007/978-1-4939-9674-2_8

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title = "Detection of mRNA Transfer Between Mammalian Cells in Coculture by Single-Molecule Fluorescent In Situ Hybridization (smFISH)",

abstract = "In eukaryotic cells, a small percentage of mRNA molecules can undergo transfer from one cell to another. mRNA transfer occurs primarily via membrane nanotubes, which are long thin protrusions that are produced by numerous cell types and can connect cells that can be up to hundreds of microns apart. Potentially, mRNAs might also transfer via extracellular vesicles (EVs). Here we describe a method to detect transferred mRNA in cocultures of two different cell types and to distinguish between nanotube- and EVs-mediated transfer. This method uses single molecule fluorescent in situ hybridization (smFISH) to provide an accurate and quantitative detection of transferred mRNA molecules and their subcellular localization. Following the guidelines presented here will allow the user to investigate mRNA transfer of most transcripts in any co-culture system. In addition, we present modifications that improve nanotube preservation during the smFISH procedure.",

author = "Gal Haimovich and Gerst, {Jeffrey E.}",

note = "G.H. was funded by the Koshland Foundation and McDonald-Leapman Grant Senior Postdoctoral Fellowships. This work was funded by grants to J.E.G. from the Joel and Mady Dukler Fund for Cancer Research, the Jean-Jacques Brunschwig Fund for the Molecular Genetics of Cancer, a Proof-of-Principle Grant from the Moross Integrated Cancer Center (Weizmann Institute of Science), and the US-Israel Binational Science Foundation-National Science Foundation (#2015846).",

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doi = "10.1007/978-1-4939-9674-2_8",

language = "الإنجليزيّة",

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Detection of mRNA Transfer Between Mammalian Cells in Coculture by Single-Molecule Fluorescent In Situ Hybridization (smFISH). / Haimovich, Gal; Gerst, Jeffrey E.
Imaging Gene Expression: Methods and Protocols. ed. / Yaron Shav-Tal. New York, NY: Humana Press, 2019. p. 109-129 (Methods in Molecular Biology; Vol. 2038).

Research output: Chapter in Book/Report/Conference proceeding › Chapter

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T1 - Detection of mRNA Transfer Between Mammalian Cells in Coculture by Single-Molecule Fluorescent In Situ Hybridization (smFISH)

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AU - Gerst, Jeffrey E.

N1 - G.H. was funded by the Koshland Foundation and McDonald-Leapman Grant Senior Postdoctoral Fellowships. This work was funded by grants to J.E.G. from the Joel and Mady Dukler Fund for Cancer Research, the Jean-Jacques Brunschwig Fund for the Molecular Genetics of Cancer, a Proof-of-Principle Grant from the Moross Integrated Cancer Center (Weizmann Institute of Science), and the US-Israel Binational Science Foundation-National Science Foundation (#2015846).

PY - 2019/8/13

Y1 - 2019/8/13

N2 - In eukaryotic cells, a small percentage of mRNA molecules can undergo transfer from one cell to another. mRNA transfer occurs primarily via membrane nanotubes, which are long thin protrusions that are produced by numerous cell types and can connect cells that can be up to hundreds of microns apart. Potentially, mRNAs might also transfer via extracellular vesicles (EVs). Here we describe a method to detect transferred mRNA in cocultures of two different cell types and to distinguish between nanotube- and EVs-mediated transfer. This method uses single molecule fluorescent in situ hybridization (smFISH) to provide an accurate and quantitative detection of transferred mRNA molecules and their subcellular localization. Following the guidelines presented here will allow the user to investigate mRNA transfer of most transcripts in any co-culture system. In addition, we present modifications that improve nanotube preservation during the smFISH procedure.

AB - In eukaryotic cells, a small percentage of mRNA molecules can undergo transfer from one cell to another. mRNA transfer occurs primarily via membrane nanotubes, which are long thin protrusions that are produced by numerous cell types and can connect cells that can be up to hundreds of microns apart. Potentially, mRNAs might also transfer via extracellular vesicles (EVs). Here we describe a method to detect transferred mRNA in cocultures of two different cell types and to distinguish between nanotube- and EVs-mediated transfer. This method uses single molecule fluorescent in situ hybridization (smFISH) to provide an accurate and quantitative detection of transferred mRNA molecules and their subcellular localization. Following the guidelines presented here will allow the user to investigate mRNA transfer of most transcripts in any co-culture system. In addition, we present modifications that improve nanotube preservation during the smFISH procedure.

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DO - 10.1007/978-1-4939-9674-2_8

M3 - فصل

SN - 9781493996735

T3 - Methods in Molecular Biology

SP - 109

EP - 129

BT - Imaging Gene Expression

A2 - Shav-Tal, Yaron

PB - Humana Press

CY - New York, NY

ER -

Detection of mRNA Transfer Between Mammalian Cells in Coculture by Single-Molecule Fluorescent In Situ Hybridization (smFISH)

Abstract

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