A linear DNA probe as an alternative to a molecular beacon for improving the sensitivity of a homogenous fluorescence biosensing platform for DNA detection using target-primed rolling circle amplification

Herein, we report a simple and homogenous fluorescence method for ultrasensitive DNA detection. It is based on rolling circle amplification (RCA) and fluorescence resonance energy transfer (FRET). As an alternative to a molecular beacon (MB), a linear single-labeled DNA probe was used in this RCA-based fluorescence strategy for DNA detection. The performance of linear DNA probes was compared with that of the MB probe in an RCA-based fluorescence strategy. The results showed that the linear DNA probes could effectively avoid the fluorescence quenching between neighboring signal probes, which would significantly improve the sensitivity of the RCA-based fluorescence strategy. This method exhibited a high sensitivity toward target DNA with a detection limit of 0.7 aM, which was about 100-fold lower than that of the RCA-based fluorescence strategy with MB as the signal probe. This method provides a simple, isothermal, and low-cost approach for sensitive detection of DNA and holds great potential for early diagnosis in gene-related diseases.