ATAC-Seq-based Identification of Extrachromosomal Circular DNA in Mammalian Cells and Its Validation Using Inverse PCR and FISH.

ABSTRACT: Recent studies from multiple labs including ours have demonstrated the importance of extrachromosomal circular DNA (eccDNA) from yeast to humans ( Shibata et al., 2012 ; Dillon et al., 2015 ; Møller et al., 2016 ; Kumar et al., 2017 ; Turner et al., 2017 ; Kim et al., 2020 ). More recently, it has been found that cancer cells obtain a selective advantage by amplifying oncogenes on eccDNA, which drives genomic instability ( Wu et al., 2019 ; Kim et al., 2020 ). Previously, we have purified circular DNA and enriched the population using rolling circle amplification followed by high-throughput sequencing for the identification of eccDNA based on the unique junctional sequence. However, eccDNA identification by rolling circle amplification is biased toward small circles. Here, we report a rolling circle-independent method to detect eccDNA in human cancer cells. We demonstrate a sensitive and robust step-by-step workflow for finding novel eccDNAs using ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) combined with a Circle_finder bioinformatics algorithm to predict the eccDNAs, followed by its validation using two independent methods, inverse PCR and metaphase FISH (Fluorescence in situ Hybridization).

SUBMITTER: Su Z 

PROVIDER: S-EPMC8161110 | biostudies-literature | 2021 May

REPOSITORIES: biostudies-literature