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Method Article
Purification and Sequencing of Large Circular DNA from Human Cells
Anton Henssen
Department of Pediatrics, Division of Oncology & Hematology, Charité Universitätsmedizin Berlin, Berlin, Germany.
Corresponding Author
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This work is licensed under a CC BY 4.0 License. Read Full License
This method is adapted from Møller et. al. (2015)1 and optimized to extract large circular DNA from human cancer cells. Previous methods of isolating circular DNA species have involved CsCl-EtBr density gradients and 2D gel electrophoresis of genomic DNA2. While Møller et. al. (2015) use column purification to isolate circular DNA from yeast, our protocol uses magnetic bead-based purification to extract low and high molecular weight circular DNA from human cells. Although the biological significance of circular DNA is incompletely understood, recent work has shown that it is prevalent in yeast1 and cancer cells3, has demonstrated that circular DNA amplification can drive intratumoral heterogeneity3, and has shown that circular DNA may form via microdeletions4 and intrachromatid homologous recombination5.
Keywords
Circular DNA, eccDNA, ecDNA, Cancer
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Protocol as PDF Protocol as PDF This is the entire protocol in PDF format for download.
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This is a preprint. It has not completed peer review.
Method Article
Purification and Sequencing of Large Circular DNA from Human Cells
Anton Henssen
Department of Pediatrics, Division of Oncology & Hematology, Charité Universitätsmedizin Berlin, Berlin, Germany.
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 22 Jan, 2019
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
This method is adapted from Møller et. al. (2015)1 and optimized to extract large circular DNA from human cancer cells. Previous methods of isolating circular DNA species have involved CsCl-EtBr density gradients and 2D gel electrophoresis of genomic DNA2. While Møller et. al. (2015) use column purification to isolate circular DNA from yeast, our protocol uses magnetic bead-based purification to extract low and high molecular weight circular DNA from human cells. Although the biological significance of circular DNA is incompletely understood, recent work has shown that it is prevalent in yeast1 and cancer cells3, has demonstrated that circular DNA amplification can drive intratumoral heterogeneity3, and has shown that circular DNA may form via microdeletions4 and intrachromatid homologous recombination5.