Measuring the Activity of DNA Repair Enzymes in Isolated Mitochondria.
Base excision repair
DNA repair
Plant mitochondria
mtDNA
Journal
Methods in molecular biology (Clifton, N.J.)
ISSN: 1940-6029
Titre abrégé: Methods Mol Biol
Pays: United States
ID NLM: 9214969
Informations de publication
Date de publication:
2022
2022
Historique:
entrez:
21
9
2021
pubmed:
22
9
2021
medline:
11
1
2022
Statut:
ppublish
Résumé
Nuclear, mitochondrial and plastidic DNA is constantly exposed to conditions, such as ultraviolet radiation or reactive oxygen species, which will induce chemical modifications to the nucleotides. Unless repaired, these modifications can lead to mutations, so the nucleus, mitochondria and plastids each contains a number of DNA repair systems. We here describe assays for measuring the enzyme activities associated with the base-excision repair pathway in potato tuber mitochondria. As the name implies, this pathway involves removing a modified base and replacing it with an undamaged base. Activity of each of the enzymes involved, DNA glycosylase, apurinic/apyrimidinic endonuclease, DNA polymerase and DNA ligase can be measured by incubating a mitochondrial extract with a specifically designed oligonucleotide. After incubation, the reaction mixture is separated on a polyacrylamide gel, and the amounts of specific products formed is estimated by autoradiography, which makes it possible to calculate the enzymatic activity.
Identifiants
pubmed: 34545501
doi: 10.1007/978-1-0716-1653-6_21
doi:
Substances chimiques
DNA, Mitochondrial
0
DNA Glycosylases
EC 3.2.2.-
DNA-(Apurinic or Apyrimidinic Site) Lyase
EC 4.2.99.18
DNA Repair Enzymes
EC 6.5.1.-
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
321-334Informations de copyright
© 2022. Springer Science+Business Media, LLC, part of Springer Nature.
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