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Two-Dimensional Agarose Gel Analysis of DNA Replication Intermediates

 Last Update: 2021-01-24
 Source: Internet
 Author: User

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The neutral/neutral (N/N) two-dimensional (2-D) agarose gel technique is a useful tool for understanding the mechanisms leading to the complete duplication of linear eukaryotic chromosomes. For the yeast
Saccharomyces cerevisiae
, it has been used to localize and characterize origins of replication as well as fork progression characteristics in a variety of experimental settings. The method involves running a first-dimension gel in order to separate restriction-digested replication intermediates of different mass. A gel slice containing the continuum of replicating
DNA
is then cut and subjected to a second-dimension gel, such as to resolve replication intermediates of varying topology. The 2-D gel is then blotted and probed to allow an examination of replication intermediates in specific DNA regions.

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