Increasingly branched rolling circle amplification for the cancer gene detection

• Published in: Biosensors & bioelectronics Vol. 86; pp. 1067 - 1073
• Main Authors: Li, Hongling, Xu, Jianguo, Wang, Zhenmeng, Wu, Zai-Sheng, Jia, Lee
• Format: Journal Article
• Language: English
• Published: England Elsevier B.V 15.12.2016
• Subjects: Amplification; Biomarkers, Tumor - genetics; Biotechnology; Cancer; Circularity; Deoxyribonucleic acid; DNA Probes - genetics; DNA, Neoplasm - genetics; Equipment Design; Equipment Failure Analysis; Fluorescence; Genes; Genes, Neoplasm - genetics; HCT116 Cells; Humans; In Situ Hybridization, Fluorescence - instrumentation; Increasingly branched rolling circle amplification (IB-RCA); Molecular Diagnostic Techniques - instrumentation; Nucleic Acid Amplification Techniques - instrumentation; Polypropylenes; Proto-Oncogene Proteins p21(ras) - analysis; Proto-Oncogene Proteins p21(ras) - genetics; Reproducibility of Results; Sensitivity and Specificity; Increasingly branched rolling circle amplification (IB-RCA)
• ISSN: 0956-5663; 1873-4235
• DOI: 10.1016/j.bios.2016.07.095

An increasingly branched rolling circle amplification (IB-RCA) which contains a padlock probe (PP) and a structurally tailored molecular beacon (MB) was innovatively developed for highly sensitive detection of cancer gene, Kras gene codon 12. In this system, the PP can be circularized after hybridization with the precisely-matched target DNA, while the stem of MB can be also opened by target DNA, resulting in hybridization with the circularized PP to generate a long tandem single-stranded DNA (ssDNA) product. Since the MB is also designed to hybridize with ssDNA product, the newly-opened MBs are able to trigger the next RCA reactions, therapy producing branched rolling circle amplification (RCA) products and in turn leading to the increasingly branched RCA (IB-RCA). This alternately and continuously operates hybridization-based MB opening and opened MBs-triggered RCA. As a result, a great number of MBs are opened that is associated with a dramatically amplified fluorescent signal, enabling to quantify target DNA down to 100 fM. This sensing method demonstrates a new concept of IB-RCA amplification even in a simple way to efficiently transduce the fluorescence signal, accomplishing the highly sensitive and selective detection of cancer gene. •Increasingly branched rolling circle amplification (IB-RCA) for cancer diagnosis was developed.•The novel IB-RCA possesses several merits, indicating great promise for cancer gene detection.•The sensing strategy has been applied to screen the target in the presence of human serum.