Quantitative analysis of global 5-methyl- and 5-hydroxymethylcytosine in TET1 expressed HEK293T cells.
Biosens Bioelectron
; 167: 112472, 2020 Nov 01.
Article
em En
| MEDLINE
| ID: mdl-32763827
ABSTRACT
DNA methylation at the 5-position of cytosine bases (5-methylcytosine, 5mC) in genomic DNA is representative epigenetic modification and is involved in many cellular processes, including gene expression and embryonic development. The hydroxylation of 5mC provide 5-hydroxymethylcytosine (5hmC), the so-called sixth base rediscovered recently in mammalian cells, is also considered to act as an epigenetic regulator. We report herein the immunochemical assessment of 5hmC achieved by an enzyme-linked immunosorbent assay (ELISA) using our linker technology. The keys to this assay are 1) the immobilization of genomic DNA with the bifunctional linker molecule, and 2) quantitative analysis by using guaranteed standard samples containing defined amounts of 5hmC. We succeeded in the sensitive and quantitative detection of 5hmC as well as 5mC in HEK293T cells transfected with TET1, and also monitored the effect of ascorbate on the TET1 catalyzed conversion of 5mC to 5hmC. Our linker technology enables the rapid and stable immobilization of genomic samples and thus contributes to the realization of a reproducible 5hmC evaluation method.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Técnicas Biossensoriais
/
5-Metilcitosina
Limite:
Animals
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Humans
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article