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Related Concept Videos

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...
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RNA Splicing01:32

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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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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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Related Experiment Video

Updated: Sep 3, 2025

Quantitative Analysis of Alternative Pre-mRNA Splicing in Mouse Brain Sections Using RNA In Situ Hybridization Assay
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FACS-Based Neuronal Cell Type-Specific RNA Isolation and Alternative Splicing Analysis.

Zicheng Wei1, Yuxin Qin1, Gordon Fishell2

  • 1Department of Physiology, Guangdong Provincial Key Laboratory of Brain Function and Disease, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China.

Methods in Molecular Biology (Clifton, N.J.)
|July 27, 2022
PubMed
Summary

Researchers developed a new method using fluorescence-activated cell sorting (FACS) to isolate specific neuron types. This allows for detailed analysis of alternative splicing in neuronal subpopulations, preserving their transcription state.

Keywords:
Cell type–specificFACSPre-mRNA splicing

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Area of Science:

  • Neuroscience
  • Molecular Biology
  • Genomics

Background:

  • Alternative splicing of pre-mRNAs significantly expands the coding potential of genomes.
  • This process is crucial for generating the complex transcriptome observed in neurons.
  • Neuronal subpopulations exhibit diverse alternative splicing programs and differential expression of splicing regulators.

Purpose of the Study:

  • To develop an efficient protocol for isolating specific neuronal types.
  • To enable cell type-specific analysis of alternative splicing events.
  • To preserve the transcription state of neurons for accurate transcriptomic analysis.

Main Methods:

  • A fluorescence-activated cell sorting (FACS)-based protocol was established.
  • The method isolates RNA from fluorescent protein-labeled neuronal types.
  • Optimization focused on preserving transcription states and improving cell suspension purification efficiency.

Main Results:

  • The protocol successfully isolates fluorescent protein-labeled neurons with high efficiency.
  • The transcription states required for gene expression and alternative splicing analysis are well-preserved.
  • This method allows for cell type-specific analysis of alternative splicing events.

Conclusions:

  • The developed FACS-based protocol provides an efficient means to isolate specific neuronal types.
  • This method facilitates the study of transcriptomic diversity, particularly alternative splicing, in distinct neuronal subpopulations.
  • The protocol is universally applicable for analyzing alternative splicing in fluorescently labeled neurons.