Base-resolution analysis of 5-hydroxymethylcytidine by selective oxidation and reverse transcription arrest.
Org Biomol Chem
; 19(29): 6478-6486, 2021 07 28.
Article
em En
| MEDLINE
| ID: mdl-34241617
ABSTRACT
While 5-hydroxymethylcytidine in RNA (hm5C) is associated with cellular development and differentiation, its distribution and biological function remain largely unexplored because suitable detection methods are lacking. Here, we report a base-resolution sequencing method for hm5C in RNA by applying peroxotungstate-mediated chemical conversion of hm5C to trihydroxylated thymine (thT). Reverse transcription by SuperScript III terminated at the thT site, probably because of its unnatural nucleobase structure producing truncated cDNA. Consequently, base-resolution analysis of the hm5C sites in RNA was achieved with both Sanger sequencing and Illumina sequencing analysis by comparing sequencing data before and after peroxotungstate treatment.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Citidina
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article