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Related Concept Videos

RNA-seq03:21

RNA-seq

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RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases. 
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while...
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FISH - Fluorescent In-situ Hybridization02:07

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Fluorescence in situ hybridization, or FISH, was developed in the early 1980s and has quickly become one of the most widely used techniques in cytogenetics. Labeled probes are used to bind complementary DNA or RNA sequences on a chromosome or in a region within a cell. Earlier, the probes could only be obtained by cloning or reverse transcription of a DNA template. Currently, the probe oligonucleotides can be synthesized synthetically. Additionally, with the advancement of optical techniques,...
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Ribosome Profiling

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Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
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Highly-Multiplexed Tissue Imaging with Raman Dyes
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Highly specific multiplexed RNA imaging in tissues with split-FISH.

Jolene Jie Lin Goh1, Nigel Chou1, Wan Yi Seow1

  • 1Genome Institute of Singapore, Agency for Science, Technology and Research (A*STAR), Singapore, Singapore.

Nature Methods
|June 17, 2020
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Summary
This summary is machine-generated.

We developed split-FISH, a novel RNA detection method that improves accuracy by reducing background noise. This technique enables precise spatial transcriptome profiling in intact tissues.

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Area of Science:

  • Molecular Biology
  • Genomics
  • Biotechnology

Background:

  • Multiplexed fluorescence in situ hybridization (FISH) is crucial for spatial transcriptomics.
  • Existing methods often suffer from off-target binding and high background fluorescence, limiting accuracy in complex tissues.

Purpose of the Study:

  • To introduce split-FISH, a novel multiplexed FISH method utilizing a split-probe design.
  • To enhance specificity and reduce background noise for accurate RNA profiling in uncleared tissues.

Main Methods:

  • Split-FISH employs a split-probe design to increase probe binding specificity.
  • The method was validated on various mouse tissues.
  • Quantification of 317 genes in single cells was performed.

Main Results:

  • Split-FISH significantly reduces off-target background fluorescence and false positives.
  • Accurate RNA profiling was achieved in uncleared, complex tissues.
  • Diverse spatial localization patterns of gene expression were revealed.

Conclusions:

  • Split-FISH offers enhanced specificity and accuracy for spatial transcriptomics.
  • The method enables detailed analysis of transcriptome organization in intact tissues.
  • Split-FISH is a powerful tool for understanding spatial gene regulation.