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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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RNA Splicing01:32

RNA Splicing

56.2K
Splicing is the process by which eukaryotic RNA is edited before its translation into protein. The RNA strand transcribed from eukaryotic DNA is called the primary transcript. The primary transcripts that become mRNAs are called precursor messenger RNAs (pre-mRNAs). Eukaryotic pre-mRNA contains alternating sequences of exons and introns. Exons are nucleotide sequences that code for proteins, whereas introns are the non-coding regions. In RNA splicing, introns are removed and exons are bonded...
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Pre-mRNA Processing: RNA Splicing01:36

Pre-mRNA Processing: RNA Splicing

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5.2K
Alternative RNA Splicing02:18

Alternative RNA Splicing

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Alternative RNA splicing is the regulated splicing of exons and introns to produce different mature mRNAs from a single pre-mRNA. Unlike in constitutive splicing where a single gene produces a single type of mRNA, alternative splicing allows an organism to produce multiple proteins from a single gene and plays an important role in protein diversity.
There are five types of alternative RNA splicing that vary in the ways the pre-mRNA segments are removed or retained in the mature mRNA. The first...
21.0K

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Related Experiment Video

Updated: Jun 13, 2025

3' End Sequencing Library Preparation with A-seq2
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3' End Sequencing Library Preparation with A-seq2

Published on: October 10, 2017

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StringTie3 Improves Total RNA-seq Assembly by Resolving Nascent and Mature Transcripts.

Ida Shinder1,2, Geo Pertea3, Richard Hu2,4

  • 1Cross Disciplinary Graduate Program in Biomedical Sciences, Johns Hopkins School of Medicine, Baltimore, MD, United States.

Biorxiv : the Preprint Server for Biology
|June 12, 2025
PubMed
Summary
This summary is machine-generated.

StringTie3 accurately assembles total RNA sequencing (RNA-seq) data by distinguishing nascent from mature transcripts. This new method improves RNA-seq assembly accuracy and reveals novel insights into gene regulation.

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Area of Science:

  • Genomics
  • Molecular Biology
  • Bioinformatics

Background:

  • Accurate transcriptome assembly from total RNA sequencing (RNA-seq) is hindered by the conflation of nascent and mature RNA isoforms.
  • Existing methods lead to misassemblies and quantification errors, impacting downstream biological analyses.

Purpose of the Study:

  • To introduce StringTie3, a novel assembler for total RNA-seq data that addresses the challenge of distinguishing nascent from mature transcripts.
  • To improve the accuracy and sensitivity of RNA-seq assembly, particularly for long-read and hybrid datasets.

Main Methods:

  • StringTie3 incorporates a "nascent mode" to model co-transcriptional splicing and separate nascent from mature transcripts.
  • A refined long-read module differentiates genuine polyadenylation sites from artifacts.
  • Performance was evaluated across short-, long-, and hybrid-read datasets using various biological experiments.

Main Results:

  • StringTie3 significantly reduces assembly errors compared to existing tools.
  • It improves short-read total RNA-seq precision by up to 20% and enhances long-read assembly sensitivity and precision by up to 37% and 75%, respectively.
  • Analysis of Argonaute knockouts and breast cancer samples revealed distinct patterns of nascent and mature RNA regulation.

Conclusions:

  • StringTie3 provides a powerful framework for accurate transcriptome reconstruction from total RNA-seq data.
  • The ability to distinguish nascent from mature RNA uncovers previously hidden layers of transcriptional and post-transcriptional regulation.
  • This tool enhances the investigation of complex RNA processing and regulatory mechanisms.