Direct and quantitative electrophoretic detection of covalently closed DNA circles formed by in vitro ligation

Research paper by Akiko Mizutani, Masafumi Tanaka

Indexed on: 01 Jun '18Published on: 30 May '18Published in: Analytical Biochemistry: Methods in the Biological Sciences


Publication date: Available online 29 May 2018 Source:Analytical Biochemistry Author(s): Akiko Mizutani, Masafumi Tanaka The typical products of enzymatic circularization of DNA, using DNA ligase or recombinase, are covalently closed and mostly relaxed DNA circles. Because they are difficult to analyze on conventional gels, they are often converted to nicked circles prior to electrophoresis. Herein, we present a sensitive and quantitative procedure for directly analyzing ligated closed circle DNA on agarose gels without additional treatments. Specifically, inclusion of GelStar dye in the gel allowed detection of ligated closed circle DNAs, which were likely super-twisted by being intercalated by GelStar, as discrete bands with good separation from linear DNA of the same sizes. Graphical abstract