Rolling circle amplification (RCA) catalyzed by φ29 DNA polymerase offers a simple method for DNA amplification in the presence of a circular DNA template and its complimentary primer. RCA continuously produces long single-strand DNA using the strand displacement activity of polymerase during DNA synthesis. This property allows one to monitor the progress of a reaction by means of electrophoresis or fluorescence measurements, and has eventually allowed the application of RCA to signal increments in the sensing of a variety of molecular species. Originally, RCA was successfully applied for the detection of specific DNA, such as single nucleotide polymorphisms. In addition, the conjugation of an antibody with a primer achieves efficient signal enhancement in antigen detection, and mRNA can also be specifically detected. Since RCA is a carry-over contamination-resistant, cost-effective, and user-friendly method of DNA amplification, RCA could be a universal technology for biosensing in fields of medical- and food-related industries.