Imaging rRNA Methylation in Bacteria by MR-FISH

Martin R Challand

doi:10.1007/978-1-4939-9674-2_7

Imaging rRNA Methylation in Bacteria by MR-FISH

Martin R Challand

School of Biochemistry

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

Abstract

Methylation of RNA is normally monitored in purified cell lysates using next-generation sequencing, gel electrophoresis, or mass spectrometry as readouts. These bulk methods require the RNA from ~104 to 107 cells to be pooled to generate sufficient material for analysis. Here we describe a method—methylation-sensitive RNA in situ hybridization (MR-FISH)—that assays rRNA methylation in bacteria on a cell-by-cell basis, using methylation-sensitive hybridization probes and fluorescence microscopy. We outline step-by-step protocols for designing probes, in situ hybridization, and analysis of data using freely available code

Original language	English
Title of host publication	Imaging Gene Expression
Publisher	Humana Press
Pages	89-107
Number of pages	18
ISBN (Electronic)	978-1-4939-9674-2
ISBN (Print)	978-1-4939-9673-5
DOIs	https://doi.org/10.1007/978-1-4939-9674-2_7
Publication status	Published - 13 Aug 2019

Publication series

Name	Methods in Molecular Biology

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10.1007/978-1-4939-9674-2_7

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Unravelling a Novel Mode of Multiple Antibiotic Resistance: Mechanism and Inhibition of Radical-SAM RNA Methyltransferases
Spencer, J. (Principal Investigator)
1/07/12 → 30/09/15
Project: Research

Cite this

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abstract = "Methylation of RNA is normally monitored in purified cell lysates using next-generation sequencing, gel electrophoresis, or mass spectrometry as readouts. These bulk methods require the RNA from \textasciitilde{}104 to 107 cells to be pooled to generate sufficient material for analysis. Here we describe a method—methylation-sensitive RNA in situ hybridization (MR-FISH)—that assays rRNA methylation in bacteria on a cell-by-cell basis, using methylation-sensitive hybridization probes and fluorescence microscopy. We outline step-by-step protocols for designing probes, in situ hybridization, and analysis of data using freely available code",

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AB - Methylation of RNA is normally monitored in purified cell lysates using next-generation sequencing, gel electrophoresis, or mass spectrometry as readouts. These bulk methods require the RNA from ~104 to 107 cells to be pooled to generate sufficient material for analysis. Here we describe a method—methylation-sensitive RNA in situ hybridization (MR-FISH)—that assays rRNA methylation in bacteria on a cell-by-cell basis, using methylation-sensitive hybridization probes and fluorescence microscopy. We outline step-by-step protocols for designing probes, in situ hybridization, and analysis of data using freely available code

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BT - Imaging Gene Expression

PB - Humana Press

ER -

Imaging rRNA Methylation in Bacteria by MR-FISH

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Unravelling a Novel Mode of Multiple Antibiotic Resistance: Mechanism and Inhibition of Radical-SAM RNA Methyltransferases

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