title: Fluorescent in situ hybridization on comets: FISH comet.
abstract: The DNA in eukaryotic cells is organized into loop domains that represent basic structural and functional units of chromatin packaging. The comet assay, a sensitive method for monitoring DNA damage and repair, involves electrophoresis of nucleoids comprising supercoiled DNA attached to the nuclear matrix. Breaks in the DNA relax the supercoiling and allow DNA loops to expand, and on electrophoresis to move towards the anode, giving the appearance of a comet tail. We use fluorescent in situ hybridization (FISH) to investigate the structure of the chromatin within comet preparations and to study specific DNA sequences within comets. In this chapter we describe our FISH comets protocols, deal with some technical questions and outline the theory. FISH with comets should be useful to researchers interested in the structural organization of DNA and chromatin, the localization of DNA damage, and the kinetics of repair of damage.
authors.affiliation: NorGenoTech AS, Totenvegen 2049, Skreia, 2848, Norway, sas@norgenotech.no.
authors.forename: Sergey
authors.initials: S
authors.lastname: Shaposhnikov
authors.forename: Naouale
authors.initials: N
authors.lastname: El Yamani
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Collins
mesh_terms: D020552:Comet Assay / Q000379:methods*; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*