Methylated RNA Immunoprecipitation Assay to Study m5C Modification in Arabidopsis.
J Vis Exp
; (159)2020 05 14.
Article
em En
| MEDLINE
| ID: mdl-32478728
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.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Arabidopsis
/
RNA de Plantas
/
5-Metilcitosina
/
Imunoprecipitação
Idioma:
En
Revista:
J Vis Exp
Ano de publicação:
2020
Tipo de documento:
Article