Identification of Extrachromosomal Circular DNA in Hop via Rolling Circle Amplification

During a survey for new viruses affecting hop plants, a circular DNA molecule was identified via rolling circle amplification (RCA) and later characterized. A small region of the 5.7-kb long molecule aligned with a microsatellite region in the Humulus lupulus genome, and no coding sequence was identified. Sequence analysis and literature review suggest that the small DNA molecule is an extranuclear DNA element, specifically, an extrachromosomal circular DNA (eccDNA), and its presence was confirmed by electron microscopy. This work is the first report of eccDNAs in the family Cannabaceae. Additionally, this work highlights the advantages of using RCA to study extrachromosomal DNA in higher plants.