Designing a Fluorescence Padlock Probe-Based Biosensor and Colorimetric Assay for the Detection of G12d Kras Mutation Publisher Pubmed



Mahmoudian F¹ ; Akbariqomi M¹ ; Heidari R¹ ; Ghahremani MH² ; Roshan N³ ; Adabi M⁴ ; Absalan M¹ ; Karimi F⁴ ; Bahrami S⁵ ; Fathi S⁴ ; Tavoosidana G¹
Source: Biomarkers in Medicine Published:2021

Abstract

Aim: Cell-free DNA in the plasma is known to be a potential biomarker for noninvasive diagnosis of oncogenic mutations. The authors aimed to design an optimized padlock probe-based hyperbranched rolling circle amplification biosensor to detect the KRAS G12D mutation using fluorescence and colorimetric methods. Methods: Single-factor experiments, Plackett-Burman design and response surface methodology were applied to optimize the padlock probe-based hyperbranched rolling circle amplification reaction. Results: The maximum fluorescence intensity was achieved at a padlock probe concentration of 1.5 pM and target concentration of 9 pM at 38°C ligation temperature. The proposed biosensor has a low detection limit of 60 fM of target DNA and a linear response in the concentration range of 60 fM to 0.2 pM. Conclusion: The results indicated the power of these assays to detect KRAS point mutations in liquid state reactions. © 2021 Future Medicine Ltd.