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Ribosome Profiling02:24

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.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique...
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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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Modern Molecular Taxonomy

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Advancements in molecular biology have revolutionized the identification and characterization of bacteria, with multiple methods leveraging DNA sequencing for enhanced precision. As sequencing technologies improve and costs decline, these approaches are increasingly used in clinical, environmental, and evolutionary studies.Multilocus Sequence Typing (MLST) examines several housekeeping genes, essential chromosomal genes encoding cellular functions, to distinguish strains. Approximately...
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Translational Regulation01:29

Translational Regulation

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Translational regulation in prokaryotes ensures efficient protein synthesis by controlling ribosome access to mRNA. This regulation is mediated by secondary RNA structures, including translational riboswitches, RNA thermometers, and small RNAs (sRNAs), which respond to intracellular and environmental signals to modulate gene expression.Translational RiboswitchesRiboswitches in the leader region of mRNAs can regulate translation by altering the accessibility of the Shine-Dalgarno (SD) sequence,...
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Leaky Scanning02:28

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During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA.  Marilyn Kozak discovered that the sequence RCCAUGG (where R...
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Related Experiment Video

Updated: Dec 17, 2025

RIBO-seq in Bacteria: a Sample Collection and Library Preparation Protocol for NGS Sequencing
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RIBO-seq in Bacteria: a Sample Collection and Library Preparation Protocol for NGS Sequencing

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MetaRibo-Seq measures translation in microbiomes.

Brayon J Fremin1, Hila Sberro1,2, Ami S Bhatt3,4

  • 1Department of Genetics, Stanford University, Stanford, CA, 94305, USA.

Nature Communications
|July 1, 2020
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Summary

A new method, MetaRibo-Seq, enables large-scale measurement of gene translation in uncultured microbiome organisms. This technique identifies thousands of previously unknown small proteins and reveals translation differences in microbial communities.

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Genomics

Background:

  • Measuring gene translation in uncultured microbiome organisms is challenging.
  • Existing methods like RNA-Seq cannot definitively confirm protein production.

Purpose of the Study:

  • To develop a method for simultaneous ribosome profiling of multiple organisms in microbiome samples.
  • To identify translated genes, particularly small proteins, and analyze translational differences.

Main Methods:

  • Developed MetaRibo-Seq, a novel ribosome profiling technique for microbiome samples.
  • Benchmarked MetaRibo-Seq against Ribo-Seq in a mock microbial community.
  • Applied MetaRibo-Seq to human fecal samples and combined with RNA-Seq data.

Main Results:

  • MetaRibo-Seq successfully profiled translation across tens to hundreds of organisms simultaneously.
  • Identified 2,091 translated, unannotated small protein families in human fecal samples, more than doubling known small proteins.
  • Revealed differences in transcript translation when combining MetaRibo-Seq with RNA-Seq.

Conclusions:

  • MetaRibo-Seq provides a comprehensive approach for translational profiling in complex microbiomes.
  • The method significantly expands the known repertoire of small proteins in the human gut microbiome.
  • Enables deeper understanding of microbial gene expression and protein synthesis in situ.