Use of the comet-FISH assay to demonstrate repair of the TP53 gene region in two human bladder carcinoma cell lines

Declan J. McKenna, Nor F. Rajab, Stephanie R. McKeown, George McKerr, Valerie J. McKelvey-Martin

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

The alkaline single-cell gel electrophoresis (comet) assay can be combined with fluorescence in situ hybridization (FISH) methodology to investigate the localization of specific gene domains within an individual cell. The position of the fluorescent hybridization spots in the comet head or tail indicates whether the sequence of interest lies within or in the vicinity of a damaged region of DNA. In this study, we used the comet-FISH assay to examine initial DNA damage and subsequent repair in the TP53 gene region of RT4 and RT112 bladder carcinoma cells after 5 Gy γ irradiation. In addition to standard comet parameter measurements, the number and location of TP53 hybridization spots within each comet was recorded at each repair time. The results indicate that the rate of repair of the TP53 gene region was fastest during the first 15 min after damage in both cell lines. When compared to overall genomic repair, the repair of the TP53 gene region was observed to be significantly faster during the first 15 min and thereafter followed a rate similar to that for the overall genome. The data indicate that the TP53 domain in RT4 and RT112 cells is repaired rapidly after γ irradiation. Furthermore, this repair may be preferential compared to the repair of overall genomic DNA, which gives a measure of the average DNA repair response of the whole genome. We suggest that the comet-FISH assay has considerable potential in the study of gene-specific repair after DNA damage.

Original languageEnglish
Pages (from-to)49-56
Number of pages8
JournalRadiation Research
Volume159
Issue number1
DOIs
Publication statusPublished - 1 Jan 2003
Externally publishedYes

Fingerprint

bladder
p53 Genes
fluorescence in situ hybridization
comets
Fluorescence In Situ Hybridization
cultured cells
genes
carcinoma
Urinary Bladder
Comet Assay
deoxyribonucleic acid
cancer
cell lines
Carcinoma
Cell Line
fluorescence
DNA Damage
assays
Genome
genome

ASJC Scopus subject areas

  • Agricultural and Biological Sciences (miscellaneous)
  • Radiology Nuclear Medicine and imaging
  • Biophysics
  • Radiation

Cite this

Use of the comet-FISH assay to demonstrate repair of the TP53 gene region in two human bladder carcinoma cell lines. / McKenna, Declan J.; Rajab, Nor F.; McKeown, Stephanie R.; McKerr, George; McKelvey-Martin, Valerie J.

In: Radiation Research, Vol. 159, No. 1, 01.01.2003, p. 49-56.

Research output: Contribution to journalArticle

McKenna, Declan J. ; Rajab, Nor F. ; McKeown, Stephanie R. ; McKerr, George ; McKelvey-Martin, Valerie J. / Use of the comet-FISH assay to demonstrate repair of the TP53 gene region in two human bladder carcinoma cell lines. In: Radiation Research. 2003 ; Vol. 159, No. 1. pp. 49-56.
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