Exponential rolling circle amplification and its sensing application for highly sensitive DNA detection of tumor suppressor gene

• Published in: Sensors and actuators. B, Chemical Chemical, 2017-05, Vol.243, p.1240-1247
• Main Authors: Xu, Huo, Xue, Chang, Zhang, Rongbo, Chen, Yanru, Li, Feng, Shen, Zhifa, Jia, Lee, Wu, Zai-Sheng
• Format: Article
• Language: English
• Subjects: Amplification; Binding sites; Biosensors; Cancer; Deoxyribonucleic acid; Diagnosis; DNA; Exponential rolling circle amplification (E-RCA); Genes; Optical communication; Polymerization; Primer-containing molecular beacon (PMB); Tumor suppressor p53 gene; Tumors
• ISSN: 0925-4005; 1873-3077
• DOI: 10.1016/j.snb.2016.12.107

•We propose a new concept, exponential rolling circle amplification (E-RCA), for circular oligonucleotide-mediated p53 gene detection.•Besides a wide linear response range, the E-RCA exhibits high assay sensitivity and capability to recognize the point mutation.•The sensing strategy could be applied to screen cancer cells to confirm the feasibility in the analysis of real samples.•The newly-proposed E-RCA sensing system is a promising candidate for disease-related gene detection in early clinical diagnostics. The development of robust biosensing platforms for accurate detection of cancer-related gene is of great significance for early diagnosis and treatment of cancer. In this paper, we proposed a new concept, exponential rolling circle amplification (E-RCA), for the circular oligonucleotide-mediated tumor suppressor p53 gene detection. Specifically, PMB (primer-containing molecular beacon), PPT (primer-containing polymerization template) and padlock probe are involved. PMB is capable of recognizing target DNA and serves as signal reporter, while PPT is designed to harbor the half binding site of nicking endonuclease and is used as the primary polymerization primer. After the target-activated hybridization between PMB and PPT, besides the as-designed PPT can promote the cyclical target strand-displacement polymerization (CTDP) and nicking-mediated strand-displacement polymerization (NSDP), the nicked primer-induced RCA can make subsequent NSDPs and RCAs proceed repeatedly and exponentially, increasing dramatically the optical signal due to the opening of a large amount of PMBs. The p53 gene can be detected down to 1 pM, and point mutation can be easily discriminated. The exciting breakthrough in the genetic diagnosis concept, desirable analytical characteristics and measured data for real sample assay indicate the potential application in biomedical research and early clinical diagnostics.