In nature, plasmid DNA and phage virus DNA often exist in the form of circular DNA, and extrachromosomal circular DNA (eccDNA) is also widely present in eukaryotic cells [1]. Unlike the linear DNA replication mechanism, circular DNA replication follows the rolling circle replication mechanism. For example, phage φX174 produces single-stranded progeny DNA through a simplified form of rolling ring replication mechanism. λ phage, on the other hand, uses the replaced leader strand in rolling circle replication as a template to synthesize the delayed strand, thereby forming double-stranded DNA (dsDNA). Based on this mechanism, researchers developed a novel in vitro nucleic acid amplification technique, rolling circle amplification (RCA) [2, 3]. In the process of rolling circle amplification, the primer is continuously extended along the circular DNA template under the catalysis of polymerase to form long single-stranded DNA (ssDNA)-containing tandem repeats. After linear or exponential RCA, the target molecule can be amplified 105 times [4] or 109 times [2], so the detection of the target molecule can be achieved with high sensitivity using RCA. Compared with the traditional polymerase chain reaction (PCR), RCA can be performed under constant temperature conditions without the need to rely on thermal cycles, thus eliminating the need for precisely controlled thermal cycler and heat-stable DNA polymerase. In addition, RCA can be performed in solutions, solid substrates, or complex biological environments. Since its invention in the mid-1990s, RCA has been widely studied for its unique advantages in detecting a variety of biomolecules, including DNA [5], RNA [6], DNA methylation [7], single-nucleotide polymorphism (SNP) [8, 9], protein [10], and cell [11, 12].