A Practical Site-specific Method for the Detection of Bulky DNA Damages.
J Mol Biol
; 436(6): 168450, 2024 03 15.
Article
en En
| MEDLINE
| ID: mdl-38246411
ABSTRACT
Helix-distorting DNA damages block RNA and DNA polymerase, compromising cell function and fate. In human cells, these damages are removed primarily by nucleotide excision repair (NER). Here, we describe damage-sensing PCR (dsPCR), a PCR-based method for the detection of these DNA damages. Exposure to DNA damaging agents results in lower PCR signal in comparison to non-damaged DNA, and repair is measured as the restoration of PCR signal over time. We show that the method successfully detects damages induced by ultraviolet (UV) radiation, by the carcinogenic component of cigarette smoke benzo[a]pyrene diol epoxide (BPDE) and by the chemotherapeutic drug cisplatin. Damage removal measured by dsPCR in a heterochromatic region is less efficient than in a transcribed and accessible region. Furthermore, lower repair is measured in repair-deficient knock-out cells. This straight-forward method could be applied by non-DNA repair experts to study the involvement of their gene-of-interest in repair. Furthermore, this method is fully amenable for high-throughput screening of DNA repair activity.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Daño del ADN
/
Aductos de ADN
/
Reparación del ADN
Tipo de estudio:
Diagnostic_studies
Límite:
Humans
Idioma:
En
Revista:
J Mol Biol
Año:
2024
Tipo del documento:
Article
País de afiliación:
Israel
Pais de publicación:
Países Bajos