An Enzyme-Free Cyclic Amplification Strategy for Intracellular Telomerase Activity Detection

Abstract

Sensitive detection of telomerase and its inhibitor are of great significance to early diagnosis and treatment of malignant tumors. A cascade signal amplification strategy based on cyclic strand displacement reaction was designed to detect telomerase activity in cell lysates and living tumor cells. The primer can be elongated with repeated sequences of (TTAGGG) <sub xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">n</sub> due to the presence of telomerase. The telomerase extension product (TEP) competes C-DNA with G-DNA by forming more stable duplex. Thus, G-DNA was released and trigger the cyclic strand displacement reaction. Two molecular beacons (MB1 and MB2) were applied to fulfill the cyclic amplification reaction and report the fluorescence signal. Compared with unamplified OMB strategy, the limit of detection for telomerase activity with amplified TMB strategy was enhanced by two orders of magnitude. It provides a new enzyme-free and PCR-free amplification method for quantification and monitoring of telomerase activity in living tumor cells. It can also be utilized for distinguishing tumor cells from normal cells by in situ imaging. The monitoring of telomerase activity with inhibitor demonstrated its potential applications in the discovery of anti-cancer drugs.