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RNA editing is a post-transcriptional modification where a precursor mRNA (pre-mRNA) nucleotide sequence is changed by base insertion, deletion, or modification. The extent of RNA editing varies from a few hundred bases, in mitochondrial DNA of trypanosomes, to a just single base, in nuclear genes of mammals. Even a single base change in the pre-mRNA can convert a codon for one amino acid into the codon for another amino acid or a stop codon. This type of re-coding can significantly affect the...
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Efficient Genome Editing of Mice by CRISPR Electroporation of Zygotes
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RNA Editing in Human and Mouse Tissues.

Harini Srinivasan1,2, Eng Piew Louis Kok2, Meng How Tan3,4

  • 1School of Chemical and Biomedical Engineering, Nanyang Technological University, Singapore, Singapore.

Methods in Molecular Biology (Clifton, N.J.)
|July 31, 2020
PubMed
Summary
This summary is machine-generated.

Adenosine-to-inosine (A-to-I) RNA editing diversifies the transcriptome and impacts health. This chapter details methods for accurately quantifying A-to-I RNA editing levels in human and mouse tissues using next-generation sequencing.

Keywords:
ADARAmplicon sequencingEpitranscriptomeRNA editingRNA-seq

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Area of Science:

  • Molecular Biology
  • Genomics
  • Posttranscriptional Regulation

Background:

  • Adenosine-to-inosine (A-to-I) RNA editing is a crucial posttranscriptional mechanism in mammals.
  • A-to-I RNA editing significantly impacts transcriptome diversity, normal development, physiology, and various human diseases.
  • Accurate quantification of RNA editing levels is essential due to their downstream effects on cellular behavior.

Purpose of the Study:

  • To describe a comprehensive workflow for examining RNA editing in human and mouse tissues.
  • To detail the application of next-generation sequencing technologies for high-throughput RNA editing site analysis.
  • To provide guidance on computational analysis of sequencing data generated from Illumina platforms.

Main Methods:

  • Harvesting of high-quality RNA samples from mammalian tissues.
  • Construction of various high-throughput sequencing libraries.
  • Computational analysis of sequencing data using established bioinformatics pipelines for Illumina platforms.

Main Results:

  • The described workflow enables accurate measurement of numerous RNA editing sites simultaneously.
  • Next-generation sequencing technologies facilitate unprecedented precision in quantifying editing levels.
  • The methodology covers both experimental and computational aspects of RNA editing analysis.

Conclusions:

  • This chapter provides a detailed protocol for studying A-to-I RNA editing in mammalian tissues.
  • The presented methods leverage advanced sequencing technologies for comprehensive transcriptome analysis.
  • Accurate RNA editing quantification is vital for understanding its role in biological processes and diseases.