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  Methylated RNA Immunoprecipitation Assay to Study m5C Modification in Arabidopsis
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 Creators:
Saplaoura, E.1, Author              
Perrera, Valentina 2, Author
Kragler, F.1, Author              
Affiliations:
1Intercellular Macromolecular Transport, Department Stitt, Max Planck Institute of Molecular Plant Physiology, Max Planck Society, ou_1753331              
2external, ou_persistent22              

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Free keywords: Genetics, RNA methylation, 5-methylcytosine (m5C), immunoprecipitation (IP), Arabidopsis, epitranscriptomics, mRNA, RNA sequencing
 Abstract: Secondary base modifications on RNA, such as m5C, affect the structure and function of the modified RNA molecules. Methylated RNA Immunoprecipitation and sequencing (MeRIP-seq) is a method that aims to enrich for methylated RNA and ultimately identify modified transcripts. Briefly, sonicated RNA is incubated with an antibody for 5-methylated cytosines and precipitated with the assistance of protein G beads. The enriched fragments are then sequenced and the potential methylation sites are mapped based on the distribution of the reads and peak detection. MeRIP can be applied to any organism, as it does not require any prior sequence or modifying enzyme knowledge. In addition, besides fragmentation, RNA is not subjected to any other chemical or temperature treatment. However, MeRIP-seq does not provide single-nucleotide prediction of the methylation site as other methods do, although the methylated area can be narrowed down to a few nucleotides. The use of different modification-specific antibodies allows MeRIP to be adjusted for the different base modifications present on RNA, expanding the possible applications of this method.

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Language(s): eng - English
 Dates: 2020
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Type: -
 Degree: -

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Title: Journal of Visualized Experiments
  Other : Journal of Visualized Experiments: JoVE
  Abbreviation : J. Vis. Exp.
Source Genre: Journal
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Publ. Info: Rockville Pike, Bethesda MD : JoVE
Pages: - Volume / Issue: (159) Sequence Number: e61231 Start / End Page: - Identifier: ISSN: 1940-087X
CoNE: https://pure.mpg.de/cone/journals/resource/1940087X