A nonradioactive restriction enzyme-mediated assay to detect DNA repair by Fe(II)/2-oxoglutarate-dependent dioxygenase

S, Gururaj and K, Naveena and Roy, Anindya (2014) A nonradioactive restriction enzyme-mediated assay to detect DNA repair by Fe(II)/2-oxoglutarate-dependent dioxygenase. Analytical Biochemistry, 465. pp. 35-37. ISSN 0003-2697

[img]
Preview
Text (Author verson post-print)
Analytical Biochemistry_465_15_35-37_2014.pdf - Accepted Version

Download (6MB) | Preview
[img]
Preview
Text
168_mmc1.pdf - Supplemental Material

Download (1MB)
[img] Text
168_mmc2.pdf - Supplemental Material
Restricted to Registered users only

Download (4MB) | Request a copy

Abstract

The Escherichia coli DNA repair enzyme AlkB belongs to the Fe(II)/2-oxoglutarate-dependent dioxygenase family. It removes methyl groups from 1-methyl adenine (1-meA) and 3-methyl cytosine (3-meC) lesions present in single-stranded DNA by oxidative decarboxylation. In the current article, we describe an in vitro assay that permits rapid detection of AlkB activity. To achieve this, we generated methylated oligonucleotide using methyl methanesulfonate and then monitored DNA repair using a methylation-sensitive restriction enzyme and novel agarose gel electrophoresis system capable of resolving small oligonucleotides. Our approach overcomes several drawbacks of NAD+-dependent formaldehyde dehydrogenase-coupled assay and radioisotope-based assay for determining AlkB DNA repair activity.

[error in script]
IITH Creators:
IITH CreatorsORCiD
Roy, Anindyahttp://orcid.org/0000-0001-8561-907X
Item Type: Article
Uncontrolled Keywords: Fe(II)/2-oxoglutarate-dependent dioxygenase; AlkB; Methyl methanesulfonate; DNA repair
Subjects: Others > Biotechnology
Divisions: Department of Biotechnology
Depositing User: Mr. Siva Shankar K
Date Deposited: 07 Oct 2014 05:24
Last Modified: 05 Dec 2017 04:52
URI: http://raiith.iith.ac.in/id/eprint/168
Publisher URL: https://doi.org/10.1016/j.ab.2014.07.003
OA policy: http://www.sherpa.ac.uk/romeo/issn/0003-2697/
Related URLs:

Actions (login required)

View Item View Item
Statistics for RAIITH ePrint 168 Statistics for this ePrint Item