Reverse strand-displacement amplification strategy for rapid detection of p53 gene

Abstract

The development of rapid approaches to detect prognostic markers is significant in reducing the morbidity and mortality of cancer. In this paper, we describe a rapid and specific biosensing platform for target DNA (p53 gene as a model) detection based on reverse strand displacement amplification (R-SDA). When the p53 gene is added, multifuctional molecular beacon (MMB) is unfolded via the hybridization with p53 gene. With the assist of Klenow fragment (KF) and Nt.BbvCI (the nicking endonuclease), p53 gene recycling could be initiated and considerable amount of complementary sequences for the MMBs (Nicked fragments, NFs) could be formed, generating enhanced fluorescence signal. Using this amplification strategy, the proposed biosensor displays the detection limit of 1 nM and a wide linear range from 1 to 100 nM, even if only one type of probe is involved. Notably, remarkable detection specificity for single-base mismatched target p53 gene is achieved. Moreover, the described biosensor also exhibited the stability in real biological samples (human serum). The rapid detection strategy can be performed less than 30 min without harsh reaction conditions or expensive nanoparticles. This biosensor shows great potential for application in clinic assay, especially, for early cancer diagnosis.