PCR-based detection of a rare linear DNA in cell culture

The described method allows for detection of rare linear DNA fragments generated during genomic deletions. The predicted limit of the detection is one DNA molecule per 10(7) or more cells. The method is based on anchor PCR and involves gel separation of the linear DNA fragment and chromosomal DNA be...

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Bibliographic Details
Published in:Biological procedures online 2002-11, Vol.4 (1), p.70-80
Main Author: Saveliev, Sergei V.
Format: Article
Language:English
Subjects:
cell culture
DNA
Polymerase Chain Reaction
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Summary:The described method allows for detection of rare linear DNA fragments generated during genomic deletions. The predicted limit of the detection is one DNA molecule per 10(7) or more cells. The method is based on anchor PCR and involves gel separation of the linear DNA fragment and chromosomal DNA before amplification. The detailed chemical structure of the ends of the linear DNA can be defined with the use of additional PCR-based protocols. The method was applied to study the short-lived linear DNA generated during programmed genomic deletions in a ciliate. It can be useful in studies of spontaneous DNA deletions in cell culture or for tracking intracellular modifications at the ends of transfected DNA during gene therapy trials.
ISSN:1480-9222
1480-9222
DOI:10.1251/bpo36