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Related Concept Videos

Labeling DNA Probes03:31

Labeling DNA Probes

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In situ hybridization (ISH) is a technique used to detect and localize specific DNA or RNA molecules in cells, tissue, or tissue sections using a labeled probe. The technique was first used in 1969 for the investigation of nucleic acids. It is currently an essential tool in scientific research and clinical settings, especially for diagnostic purposes.
Types of probes and labels
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Related Experiment Video

Updated: Apr 29, 2026

Fluorescent End-Labeling and Encapsulation of Long RNAs for Single-Molecule FRET-TIRF Microscopy
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Enzyme-Mediated Covalent Labeling Enables In Situ Imaging of RNA Modification States.

Andrew H Ryu1, Neal K Devaraj1

  • 1Department of Chemistry and Biochemistry, University of California San Diego, 9500 Gilman Drive, La Jolla, San Diego, California 92093, United States.

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Researchers developed a new method to visualize transfer RNA (tRNA) modifications within cells. This technique uses fluorescent labeling to track the hypomodification status of tRNAs, offering new insights into gene expression regulation.

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Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Post-transcriptional tRNA modifications regulate gene expression but their spatial and dynamic aspects are poorly understood due to a lack of in-cell tracking methods.
  • Current population-level sequencing and biochemical methods require RNA extraction, losing spatial information and obscuring cellular heterogeneity.

Purpose of the Study:

  • To develop a novel chemoenzymatic strategy for imaging specific tRNA modification states in live mammalian cells.
  • To enable spatial and dynamic analysis of tRNA modifications, overcoming limitations of existing techniques.

Main Methods:

  • A chemoenzymatic RNA fluorescent labeling strategy utilizing the bacterial enzyme tRNA guanine transglycosylase (TGT).
  • Selective incorporation of fluorophore-conjugated preQ1 analogues into hypomodified tRNAs lacking the queuine base.
  • Visualization of queuine-hypomodified tRNAs with subcellular resolution in fixed cells.

Main Results:

  • Demonstrated the ability to visualize tRNA hypomodification status in mammalian cells.
  • Tracked queuine incorporation and loss kinetics, analyzed genetic factors controlling modification, and differentiated cytosolic and mitochondrial tRNA populations.
  • Established a generalizable chemical framework for spatial analysis of endogenous RNA modifications in intact cells.

Conclusions:

  • The developed fluorescent labeling strategy provides a powerful tool for studying tRNA modification dynamics and spatial organization in cells.
  • This method opens new avenues for investigating the roles of tRNA modifications in translation, stress responses, and overall mammalian physiology.
  • The approach offers a versatile chemical framework for the spatial analysis of diverse RNA modifications within intact cellular systems.