| Autor: |
Leonid V. Gening, Alexandr A. Volodin, Konstantin Y. Kazachenko, Irina V. Makarova, Vyacheslav Z. Tarantul |
| Jazyk: |
angličtina |
| Rok vydání: |
2020 |
| Předmět: |
|
| Zdroj: |
Methods and Protocols, Vol 3, Iss 1, p 3 (2020) |
| Druh dokumentu: |
article |
| ISSN: |
2409-9279 |
| DOI: |
10.3390/mps3010003 |
| Popis: |
We propose an improved earlier described “mirror” method for detecting in cell nuclear extracts mutations that arise in DNA during its replication due to the misincorporation of deoxyadenosine-5′-monophosphate (dAMP) opposite 7,8-dihydro-8-oxoguanine (8-oxoG). This method is based on the synthesis of a complementary chain (“mirror”) by nuclear extracts of different mice organs on a template containing 8-oxoG and dideoxycytidine residue (ddC) at the 3′‑end. The “mirror” was amplified by PCR using primers part of which was non-complementary to the template. It allowed obtaining the “framed mirror” products. The misincorporation of dAMP in “framed mirror” products forms an EcoRI restriction site. The restriction analysis of double-stranded “framed mirror” products allows a quantification of the mutation frequency in nuclear extracts. The data obtained show that the mutagenic potential of 8-oxoG markedly varied in different organs of adult mice and embryos. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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