Rolling circle amplification or RCA is an isothermal nucleic acid amplification where a DNA or RNA strand displacement polymerase uses a primer-initiated amplification to make copies of a single-stranded circular (i.e., without a free terminus) oligonucleotide template . Similar to other isothermal amplifications such as loop-mediated isothermal amplification (2) (LAMP), recombinase polymerase amplification , and helicase-dependent isothermal amplification (4), the main advantage of RCA as a bioanalytical technique is its independence from thermal cycler. In addition, 29 polymerase-based RCA assays are operable at 30C, making them suitable for ambient condition benchtop applications at limited-resource settings. However, RCA is perhaps also unique in the aspect that it can be subjected to extensive nucleic acid engineering, and can be conjugated with proteins , nanomaterials , and polymers . Along with conventional nucleic acid detection, RCA, therefore, has seen versatile applications in the ultrasensitive detection of small molecules , proteins , and carbohydrates , DNA methylation assay , topoisomerase functionality , generation of programmable mRNA detection probes , and nucleic acid logic gates . As a result, RCA-based assays are only next to LAMP in publication number among the isothermal amplification techniques . Although fluorescence remains the primary method of choice for readout in these techniques, the flexibility of RCA has been utilized in the development of several alternatives such as optomagnetic , electrochemical , surface plasmon resonance , and colorimetrybased readout platforms . When coupled with branching techniques such as hyperbranched RCA (HRCA) , endonuclease-assisted primer regeneration RCA (PG-RCA) , or circleto-circle RCA , zeptomolar to attomolar concentration of the target can be detected within 1 -3 h.