Circle-Seq: Isolation and Sequencing of Chromosome-Derived Circular DNA Elements in Cells
Research output: Chapter in Book/Report/Conference proceeding › Book chapter › Research › peer-review
Chromosome-derived extrachromosomal circular DNA elements (eccDNAs) are detected in all eukaryotes examined so far. Here I describe the Circle-Seq protocol, applicable for physical enrichment of eccDNAs of a broad size range, combined with sequence confirmation of circular structures.Briefly, by concise alkaline treatment and gentle gravity flow-through an ion-exchange column, eccDNAs are enriched in the eluate fraction. EccDNAs are enzymatically isolated by extensive Plasmid-Safe DNase digestion of linear chromosomes and further enriched by φ29 rolling circle amplification. By means of high throughput sequencing of amplified eccDNA and custom eccDNA mapping software, around ten-thousand unique eccDNA types could be detected at nucleotide resolution in a million human muscle nuclei by this method.
Original language | English |
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Title of host publication | DNA Electrophoresis : Methods and Protocols |
Editors | Katsuhiro Hanada |
Number of pages | 17 |
Publisher | Humana Press |
Publication date | 2020 |
Pages | 165-181 |
Chapter | 15 |
ISBN (Print) | 978-1-0716-0322-2 |
ISBN (Electronic) | 978-1-0716-0323-9 |
DOIs | |
Publication status | Published - 2020 |
Series | Methods in Molecular Biology |
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Volume | 2119 |
ISSN | 1064-3745 |
- Circle-Seq, Circular DNA purification, DNA circle, Double minutes, eccDNA, Episomes, Extrachromosomal
Research areas
ID: 235588304