Measuring the Activity of DNA Repair Enzymes in Isolated Mitochondria

Beatriz Ferrando, Ian Max Møller*, Tinna Stevnsner*

*Corresponding author af dette arbejde

Publikation: Bidrag til bog/antologi/rapport/proceedingBidrag til bog/antologiForskningpeer review

Abstract

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.

OriginalsprogEngelsk
TitelPlant Mitochondria
UdgivelsesstedNew York
ForlagHumana Press
Publikationsdato2022
Sider321-334
ISBN (Trykt)978-1-0716-1652-9
ISBN (Elektronisk)978-1-0716-1653-6
DOI
StatusUdgivet - 2022
NavnMethods in Molecular Biology
Vol/bind2363
ISSN1064-3745

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